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Sample records for escherichia coli bacillus

  1. Enantioselective bioconversion using Escherichia coli cells expressing Saccharomyces cerevisiae reductase and Bacillus subtilis glucose dehydrogenase.

    PubMed

    Park, Hyun Joo; Jung, Jihye; Choi, Hyejeong; Uhm, Ki-Nam; Kim, Hyung Kwoun

    2010-09-01

    Ethyl (R, S)-4-chloro-3-hydroxybutanoate (ECHB) is a useful chiral building block for the synthesis of L-carnitine and hypercholesterolemia drugs. The yeast reductase, YOL151W (GenBank locus tag), exhibits an enantioselective reduction activity, converting ethyl-4-chlorooxobutanoate (ECOB) exclusively into (R)-ECHB. YOL151W was generated in Escherichia coli cells and purified via Ni- NTA and desalting column chromatography. It evidenced an optimum temperature of 45 degrees C and an optimum pH of 6.5-7.5. Bacillus subtilis glucose dehydrogenase (GDH) was also expressed in Escherichia coli, and was used for the recycling of NADPH, required for the reduction reaction. Thereafter, Escherichia coli cells co-expressing YOL151W and GDH were constructed. After permeablization treatment, the Escherichia coli whole cells were utilized for ECHB synthesis. Through the use of this system, the 30 mM ECOB substrate could be converted to (R)-ECHB.

  2. Creating new genes by plasmid recombination in Escherichia coli and Bacillus subtilis.

    PubMed

    Gomez, Ana; Galic, Tatjana; Mariet, Jean-François; Matic, Ivan; Radman, Miroslav; Petit, Marie-Agnès

    2005-11-01

    Gene shuffling is a way of creating proteins with interesting new characteristics, starting from diverged sequences. We tested an alternative to gene shuffling based on plasmid recombination and found that Bacillus subtilis efficiently recombines sequences with 4% divergence, and Escherichia coli mutS is more appropriate for sequences with 22% divergence.

  3. Creating New Genes by Plasmid Recombination in Escherichia coli and Bacillus subtilis

    PubMed Central

    Gomez, Ana; Galic, Tatjana; Mariet, Jean-François; Matic, Ivan; Radman, Miroslav; Petit, Marie-Agnès

    2005-01-01

    Gene shuffling is a way of creating proteins with interesting new characteristics, starting from diverged sequences. We tested an alternative to gene shuffling based on plasmid recombination and found that Bacillus subtilis efficiently recombines sequences with 4% divergence, and Escherichia coli mutS is more appropriate for sequences with 22% divergence. PMID:16269814

  4. [Factor of salinity and adaptive capacity of recombinant strains of Escherichia coli and Bacillus subtilis].

    PubMed

    Boiandin, A N; Lobova, T I; Krylova, T Iu; Kargatova, T V; Popova, L Iu; Pechurkin, N S

    2000-01-01

    Effect of different concentrations of salts on natural and recombinant strains of Bacillus subtilis and Escherichia coli was studied. The recombinant strain of B. subtilis was found to be more osmotolerant than the wild-type strain of this bacterium, whereas the opposite situation was observed for the recombinant and wild-type strains of E. coli. Some salts exerted a bacteriostatic effect on E. coli and B. subtilis. The adaptive capacity of recombinant strains depended on the number of plasmid copies in the cells. The introduction of recombinant bacteria into model ecosystems resulted in the generation of their variants with increased osmotolerance.

  5. Cloning and expression of the lepidopteran toxin produced by Bacillus thuringiensis var Sotto in Escherichia coli.

    PubMed

    Rady, M H

    1991-01-01

    During sporulation, Bacillus thuringiensis var. Sotto produces a parasporal crystalline protein which is toxic for the silk-worm, Bombyx mori and the cotton leaf worm, Spodoptera littoralis. The gene coding this crystal protein is present in a single plasmid. The plasmid DNA was isolated, purified and physically mapped using restriction endonuclease enzymes (R.E.). The gene coding the delta-endotoxin was inserted into Escherichia coli-Jm103, using cloning vector pUC8. Transformed E. coli cells were found to synthesize the delta-endotoxin as demonstrated by the pathogenicity of the transformed cells against 4th instar larvae of S. littoralis.

  6. Autolysis of Escherichia coli and Bacillus subtilis cells in low gravity

    NASA Technical Reports Server (NTRS)

    Kacena, M. A.; Smith, E. E.; Todd, P.

    1999-01-01

    The role of gravity in the autolysis of Bacillus subtilis and Escherichia coli was studied by growing cells on Earth and in microgravity on Space Station Mir. Autolysis analysis was completed by examining the death phase or exponential decay of cells for approximately 4 months following the stationary phase. Consistent with published findings, the stationary-phase cell population was 170% and 90% higher in flight B. subtilis and E. coli cultures, respectively, than in ground cultures. Although both flight autolysis curves began at higher cell densities than control curves, the rate of autolysis in flight cultures was identical to that of their respective ground control rates.

  7. Autolysis of Escherichia coli and Bacillus subtilis cells in low gravity

    NASA Technical Reports Server (NTRS)

    Kacena, M. A.; Smith, E. E.; Todd, P.

    1999-01-01

    The role of gravity in the autolysis of Bacillus subtilis and Escherichia coli was studied by growing cells on Earth and in microgravity on Space Station Mir. Autolysis analysis was completed by examining the death phase or exponential decay of cells for approximately 4 months following the stationary phase. Consistent with published findings, the stationary-phase cell population was 170% and 90% higher in flight B. subtilis and E. coli cultures, respectively, than in ground cultures. Although both flight autolysis curves began at higher cell densities than control curves, the rate of autolysis in flight cultures was identical to that of their respective ground control rates.

  8. Expression of the Arthrobacter viscosus penicillin G acylase gene in Escherichia coli and Bacillus subtilis.

    PubMed Central

    Ohashi, H; Katsuta, Y; Nagashima, M; Kamei, T; Yano, M

    1989-01-01

    The penicillin G acylase gene cloned from Arthrobacter viscosus 8895GU was subcloned into vectors, and the recombinant plasmids were transferred into Escherichia coli or Bacillus subtilis. Both E. coli and B. subtilis transformants expressed the A. viscosus penicillin G acylase. The enzyme activity was found in the intracellular portion of the E. coli transformants or in the cultured medium of the B. subtilis transformants. Penicillin G acylase production in the B. subtilis transformants was 7.2 times higher than that in the parent A. viscosus. The A. viscosus penicillin G acylase was induced by phenylacetic acid in A. viscosus, whereas the enzyme was produced constitutively in both the E. coli and B. subtilis transformants carrying the A. viscosus penicillin G acylase gene. Images PMID:2504107

  9. Overexpression and characterization of thermostable chitinase from Bacillus atrophaeus SC081 in Escherichia coli.

    PubMed

    Cho, Eun Kyung; Choi, In Soon; Choi, Young Ju

    2011-03-01

    The chitinase-producing strain SC081 was isolated from Korean traditional soy sauce and identified as Bacillus atrophaeus based on a phylogenetic analysis of the 16S rDNA sequence and a phenotypic analysis. A gene encoding chitinase from B. atrophaeus SC081 was cloned in Escherichia coli and was named SCChi-1 (GQ360078). The SCChi-1 nucleotide sequences were composed of 1788 base pairs and 596 amino acids, which were 92.6, 89.6, 89.3, and 78.9% identical to those of Bacillus subtilis (ABG57262), Bacillus pumilus (ABI15082), Bacillus amyloliquefaciens (ABO15008), and Bacillus licheniformis (ACF40833), respectively. A recombinant SCChi-1 containing a hexahistidine tag at the amino- terminus was constructed, overexpressed, and purified in E. coli to characterize SCChi-1. H(6)SCChi-1 revealed a hydrolytic band on zymograms containing 0.1% glycol chitin and showed the highest lytic activity on colloidal chitin and acidic chitosan. The optimal temperature and pH for chitinolytic activity were 50°C and pH 8.0, respectively.

  10. Thymine auxotrophy is associated with increased UV sensitivity in Escherichia coli and Bacillus subtilis.

    PubMed

    Lojo, M M

    1995-06-01

    Thymine auxotrophy was shown to be associated with an increase in UV sensitivity both in Bacillus subtilis and in Escherichia coli. This UV sensitization became clearly evident in polA5 mutants of Bacillus subtilis: at UV doses of 16 J/m2, a reduction of more than 10-fold in the survivor population is observed in thymine requiring spontaneous mutants (polA5 thyA thyB) compared to the parental strains (polA5). Reversion of either thyA or thyB mutation led to a partial recovery in the UV resistance. This result suggests that DNA repair polymerization might be improved by the biosynthesis of thymidylate or some effect associated with such activity.

  11. Sonodynamic action of curcumin on foodborne bacteria Bacillus cereus and Escherichia coli.

    PubMed

    Wang, Xinna; Ip, Margaret; Leung, Albert Wingnang; Yang, Zhengrong; Wang, Pan; Zhang, Baoting; Ip, Siupo; Xu, Chuanshan

    2015-09-01

    Bacterial contamination is an important cause of foodborne diseases. The present study aimed to investigate sonodynamic action of curcumin on foodborne bacteria Bacillus cereus (B. cereus) and Escherichia coli (E. coli). The uptake of curcumin was measured for optimizing the concentration incubation time before ultrasound sonication, and colony forming units (CFU) were counted after ultrasound treatment. The chromosomal DNA fragmentation of bacteria was analyzed and the effect of hypoxic condition on the antibacterial efficacy of sonodynamic action of curcumin was also assessed in this study. The results showed that the maximum uptake of curcumin in B. cereus and E. coli occurred in 50min after curcumin incubation. Curcumin had sonodynamic bactericidal activity in a curcumin dose-dependent manner, and 5.6-log reduction in CFU of B. cereus was observed after curcumin treatment (2.0μM), however, only 2-log reduction in CFU of E. coli after 40μM curcumin treatment. No significant change in chromosomal DNA was found after the combined treatment of curcumin and ultrasound. The survival of B. cereus and E. coli after sonodynamic treatment in hypoxic group was significantly higher than that in normal oxygen group. These findings indicated that sonodynamic action of curcumin had significant inactivation effect on foodborne bacteria, and B. cereus was more sensitive to sonodynamic treatment of curcumin than E. coli. Sonodynamic antibacterial activity of curcumin might be dependent on the oxygen environment.

  12. CdSe quantum dot internalization by Bacillus subtilis and Escherichia coli

    NASA Astrophysics Data System (ADS)

    Kloepfer, Jeremiah A.; Mielke, Randall E.; Nadeau, Jay L.

    2004-06-01

    Biological labeling has been demonstrated with CdSe quantum dots in a variety of animal cells, but bacteria are harder to label because of their cell walls. We discuss the challenges of using minimally coated, bare CdSe quantum dots as luminescent internal labels for bacteria. These quantum dots were solubilized with mercaptoacetic acid and conjugated to adenine. Significant evidence for the internal staining of Bacillus subtilis (Gram positive) and Escherichia coli (Gram negative) using these structures is presented via steady-state emission, epifluorescence microscopy, transmission electron microscopy, and energy dispersive spectroscopy. In particular, the E. coli adenine auxotroph, and not the wild type, took up adenine coated quantum dots, and this only occurred in adenine deficient growth media. Labeling strength was enhanced by performing the incubation under room light. This process was examined with steady-state emission spectra and time-resolved luminescence profiles obtained from time-correlated-single-photon counting.

  13. Surface display of recombinant proteins on Escherichia coli by BclA exosporium of Bacillus anthracis

    PubMed Central

    2013-01-01

    Background The anchoring motif is one of the most important aspects of cell surface display as well as efficient and stable display of target proteins. Thus, there is currently a need for the identification and isolation of novel anchoring motifs. Results A system for the display of recombinant proteins on the surface of Escherichia coli was developed using the Bacillus anthracis exosporal protein (BclA) as a new anchoring motif. For the surface display of recombinant proteins, the BAN display platform was constructed in which a target protein is linked to the C-terminus of N-terminal domain (21 amino acids) of BclA. The potential application of BAN platform for cell surface display was demonstrated with two model proteins of different size, the Bacillus sp. endoxylanase (XynA) and monooxygenase (P450 BM3m2). Through experimental analysis including outer membrane fractionation, confocal microscopy and activity assay, it was clearly confirmed that both model proteins were successfully displayed with high activities on the E. coli cell surface. Conclusions These results of this study suggest that the strategy employing the B. anthracis BclA as an anchoring motif is suitable for the display of heterologous proteins on the surface of E. coli and consequently for various biocatalytic applications as well as protein engineering. PMID:24053632

  14. Design and construction of a synthetic Bacillus thuringiensis Cry4Aa gene: hyperexpression in Escherichia coli.

    PubMed

    Hayakawa, Tohru; Howlader, Mohammad Tofazzal Hossain; Yamagiwa, Masashi; Sakai, Hiroshi

    2008-10-01

    Cry4Aa produced by Bacillus thuringiensis is a dipteran-specific toxin and is, therefore, of great interest for developing a bioinsecticide to control mosquitoes. However, the expression of Cry4Aa in Escherichia coli is relatively low, which is a major disadvantage in its development as a bioinsecticide. In this study, to establish an effective production system, a 1,914-bp modified gene (cry4Aa-S1) encoding Cry4Aa was designed and synthesized in accordance with the G + C content and codon preference of E. coli genes without altering the encoded amino acid sequence. The cry4Aa-S1 gene allowed a significant improvement in expression level, over five-fold, compared to that of the original cry4Aa gene. The product of the cry4Aa-S1 gene showed the same level of insecticidal activity against Culex pipiens larvae as that from cry4Aa. This suggested that unfavorable codon usage was one of the reasons for poor expression of cry4Aa in E. coli, and, therefore, changing the cry4Aa codons to accord with the codon usage in E. coli led to efficient production of Cry4Aa. Efficient production of Cry4Aa in E. coli can be a powerful measure to prepare a sufficient amount of Cry4Aa protein for both basic analytical and applied researches.

  15. Antibacterial Effects of Cissus welwitschii and Triumfetta welwitschii Extracts against Escherichia coli and Bacillus cereus

    PubMed Central

    2015-01-01

    Antibiotic resistance has increased sharply, while the pace for the development of new antimicrobials has slowed down. Plants provide an alternative source for new drugs. This study aimed to screen extracts from Cissus welwitschii and Triumfetta welwitschii for antibacterial activity against Escherichia coli and Bacillus cereus. The tests conducted included a susceptibility determination test, analysis of the effect of T. welwitschii on cell wall integrity, and transport across the membrane. It was found that the T. welwitschii methanol extracts were more effective than the water extracts and had the lowest minimum inhibitory concentration and minimum bactericidal concentration at 0.125 mg/mL and 0.5 mg/mL, respectively, against E. coli and B. cereus. The C. welwitschii extract caused the most drug accumulation in E. coli. In B. cereus, no significant drug accumulation was observed. Nucleic acid leakage in B. cereus and E. coli and protein leakage in E. coli were observed after exposure to the T. welwitschii extract. The extracts from T. welwitschii had greater antibacterial activity than the extracts from C. welwitschii. T. welwitschii may be a potential source of lead compounds for that could be developed into antibacterial agents. PMID:26904744

  16. Differential Actions of Chlorhexidine on the Cell Wall of Bacillus subtilis and Escherichia coli

    PubMed Central

    Cheung, Hon-Yeung; Wong, Matthew Man-Kin; Cheung, Sau-Ha; Liang, Longman Yimin; Lam, Yun-Wah; Chiu, Sung-Kay

    2012-01-01

    Chlorhexidine is a chlorinated phenolic disinfectant used commonly in mouthwash for its action against bacteria. However, a comparative study of the action of chlorhexidine on the cell morphology of Gram-positive and Gram-negative bacteria is lacking. In this study, the actions of chlorhexidine on the cell morphology were identified with the aids of electron microscopy. After exposure to chlorhexidine, numerous spots of indentation on the cell wall were found in both Bacillus subtilis and Escherichia coli. The number of indentation spots increased with time of incubation and increasing chlorhexidine concentration. Interestingly, the dented spots found in B. subtilis appeared mainly at the hemispherical caps of the cells, while in E. coli the dented spots were found all over the cells. After being exposed to chlorhexidine for a prolonged period, leakage of cellular contents and subsequent ghost cells were observed, especially from B subtilis. By using 2-D gel/MS-MS analysis, five proteins related to purine nucleoside interconversion and metabolism were preferentially induced in the cell wall of E. coli, while three proteins related to stress response and four others in amino acid biosynthesis were up-regulated in the cell wall materials of B. subtilis. The localized morphological damages together with the biochemical and protein analysis of the chlorhexidine-treated cells suggest that chlorhexidine may act on the differentially distributed lipids in the cell membranes/wall of B. subtilis and E. coli. PMID:22606280

  17. Analysis of insecticidal proteins from Bacillus thuringiensis and recombinant Escherichia coli by capillary electrokinetic chromatography.

    PubMed

    Luong, John H T; Male, Keith B; Mazza, Alberto; Masson, Luke; Brousseau, Roland

    2004-10-01

    Bacillus thuringiensis and recombinant Escherichia coli proteinaceous protoxins were subject to proteolysis and analyzed by capillary electrokinetic chromatography. Three resulting toxins (65 kDa) were baseline-resolved within 22 min using a 10 mM borate, pH 11 separation buffer consisting of 25 mM sodium dodecyl sulfate (SDS) and 30 mM phytic acid. The toxins displayed differential interactions with the SDS and phytic acid phases to effect their separation. The ion-pairing interaction between the analyte and phytic acid was also useful in preventing adsorption to the capillary walls and thus enhanced separation resolution and efficiency. The use of electrokinetic chromatography allows achievement of the separation in a significantly shorter time than conventional high-performance liquid chromatography (HPLC) using a diethylaminoethyl (DEAE) weak-anion exchanger.

  18. Effects of Aronia melanocarpa constituents on biofilm formation of Escherichia coli and Bacillus cereus.

    PubMed

    Bräunlich, Marie; Økstad, Ole A; Slimestad, Rune; Wangensteen, Helle; Malterud, Karl E; Barsett, Hilde

    2013-12-05

    Many bacteria growing on surfaces form biofilms. Adaptive and genetic changes of the microorganisms in this structure make them resistant to antimicrobial agents. Biofilm-forming organisms on medical devices can pose serious threats to human health. Thus, there is a need for novel prevention and treatment strategies. This study aimed to evaluate the ability of Aronia melanocarpa extracts, subfractions and compounds to prevent biofilm formation and to inhibit bacterial growth of Escherichia coli and Bacillus cereus in vitro. It was found that several aronia substances possessed anti-biofilm activity, however, they were not toxic to the species screened. This non-toxic inhibition may confer a lower potential for resistance development compared to conventional antimicrobials.

  19. Functional and Structural Characterization of Four Glutaminases from Escherichia coli and Bacillus subtilis†

    PubMed Central

    Brown, Greg; Singer, Alex; Proudfoot, Michael; Skarina, Tatiana; Kim, Youngchang; Chang, Changsoo; Dementieva, Irina; Kuznetsova, Ekaterina; Gonzalez, Claudio F.; Joachimiak, Andrzej; Savchenko, Alexei; Yakunin, Alexander F.

    2008-01-01

    Glutaminases belong to the large superfamily of serine-dependent β-lactamases and penicillin-binding proteins, and they catalyze the hydrolytic deamidation of l-glutamine to l-glutamate. In this work, we purified and biochemically characterized four predicted glutaminases from Escherichia coli (YbaS and YneH) and Bacillus subtilis (YlaM and YbgJ). The proteins demonstrated strict specificity to l-glutamine and did not hydrolyze d-glutamine or l-asparagine. In each organism, one glutaminase showed higher affinity to glutamine (E. coli YbaS and B. subtilis YlaM; Km 7.3 and 7.6 mM, respectively) than the second glutaminase (E. coli YneH and B. subtilis YbgJ; Km 27.6 and 30.6 mM, respectively). The crystal structures of the E. coli YbaS and the B. subtilis YbgJ revealed the presence of a classical β-lactamase-like fold and conservation of several key catalytic residues of β-lactamases (Ser74, Lys77, Asn126, Lys268, and Ser269 in YbgJ). Alanine replacement mutagenesis demonstrated that most of the conserved residues located in the putative glutaminase catalytic site are essential for activity. The crystal structure of the YbgJ complex with the glutaminase inhibitor 6-diazo-5-oxo-l-norleucine revealed the presence of a covalent bond between the inhibitor and the hydroxyl oxygen of Ser74, providing evidence that Ser74 is the primary catalytic nucleophile and that the glutaminase reaction proceeds through formation of an enzyme–glutamyl intermediate. Growth experiments with the E. coli glutaminase deletion strains revealed that YneH is involved in the assimilation of l-glutamine as a sole source of carbon and nitrogen and suggested that both glutaminases (YbaS and YneH) also contribute to acid resistance in E. coli. PMID:18459799

  20. Cloning of a fibrinolytic enzyme (subtilisin) gene from Bacillus subtilis in Escherichia coli.

    PubMed

    Ghasemi, Younes; Dabbagh, Fatemeh; Ghasemian, Abdollah

    2012-09-01

    Several investigations are being pursued to enhance the efficacy and specificity of fibrinolytic therapy. In this regard, microbial fibrinolytic enzymes attracted much more medical interests during these decades. Subtilisin, a member of subtilases (the superfamily of subtilisin-like serine proteases) and also a fibrinolytic enzyme is quite common in Gram-positive bacteria, and Bacillus species stand out in particular, as many extracellular and even intracellular variants have been identified. In the present work, the subtilisin gene from Bacillus subtilis PTCC 1023 was cloned into the vector pET-15b and expressed in Escherichia coli strain BL21 (DE3). Total genomic DNA were isolated and used for PCR amplification of the subtilisin gene by means of the specific primers. SDS-PAGE and enzyme assay were done for characterizing the expressed protein. A ~1,100 bp of the structural subtilisin gene was amplified. The DNA and amino acid sequence alignments resulting from the BLAST search of subtilisin showed high sequence identity with the other strains of B. subtilis, whereas significantly lower identity was observed with other bacterial subtilisins. The recombinant enzyme had the same molecular weight as other reported subtilisins and the E. coli transformants showed high subtilisin activity. This study provides evidence that subtilisin can be actively expressed in E. coli. The commercial availability of subtilisin is of great importance for industrial applications and also pharmaceutical purposes as thrombolytic agent. Thus, the characterization of new recombinant subtilisin and the development of rapid, simple, and effective production methods are not only of academic interest, but also of practical importance.

  1. High-level expression of the thermoalkalophilic lipase from Bacillus thermocatenulatus in Escherichia coli.

    PubMed

    Rúa, M L; Atomi, H; Schmidt-Dannert, C; Schmid, R D

    1998-04-01

    An efficient expression system for the previously only weakly expressed thermophilic lipase BTL2 (Bacillus thermocatenulatus lipase 2) was developed for the production of large amounts of lipase in Escherichia coli. Therefore, the gene was subcloned in the pCYT-EXP1 (pT1) expression vector downstream of the temperature-inducible lambda promoter PL. Three different expression vectors were constructed: (i) pT1-BTL2 containing the mature lipase gene, (ii) pT1-preBTL2 containing the prelipase gene and (iii) pT1-OmpABTL2 containing the mature lipase gene fused to the signal peptide of the OmpA protein, the major outer membrane protein of E. coli. With pT1-BTL2 and pT1-preBTL2, comparable expression levels of 7000-9000 U/g cells were obtained independently of the E. coli host. In contrast, with E. coli JM105 harbouring pT1-OmpABTL2, 660,000 soluble lipase U/g cells was produced, whereas, with E. coli DH5 alpha and BL321, production levels of 30,000 U/g cells were achieved. However, most of the lipase remained insoluble but active after cell breakage because of the unprocessed OmpA signal peptide. A simple cholate extraction followed by proteinase K cleavage and ultrafiltration allowed the isolation of 1.15 x 10(6) units of 90% pure mature lipase/wet cells.

  2. Online monitoring of Escherichia coli and Bacillus thuringiensis spore inactivation after advanced oxidation treatment.

    PubMed

    Sherchan, Samendra P; Snyder, Shane A; Gerba, Charles P; Pepper, Ian L

    2014-01-01

    Various studies have shown that advanced oxidation processes (AOPs) such as UV light in combination with hydrogen peroxide is an efficient process for the removal of a large variety of emerging contaminants including microorganisms. The mechanism of destruction in the presence of hydrogen peroxide (H2O2) is the enhanced formation of hydroxyl (·OH) radicals, which have a high oxidation potential. The goal of this study was to utilize in-line advanced oxidation to inactivate microbes, and document the inactivation via an in-line, real-time sensor. Escherichia coli cells and Bacillus thuringiensis spores were exposed to UV/H2O2 treatment in DI water, and the online sensor BioSentry(®) was evaluated for its potential to monitor inactivation in real-time. B. thuringiensis was selected as a non-pathogenic surrogate for B. anthracis, the causative agent of anthrax and a proven biological weapon. UV radiation and UV/H2O2 exposure resulted in a >6 log10 reduction of the viable culturable counts of E. coli vegetative cells, and a 3 log10 reduction of B. thuringiensis spores. Scanning electron microscopy of the treated samples revealed severe damage on the surface of most E. coli cells, yet there was no significant change observed in the morphology of the B. thuringiensis spores. Following AOP exposure, the BioSentry sensor showed an increase in the categories of unknown, rod and spores counts, but overall, did not correspond well with viable count assays. Data from this study show that advanced oxidation processes effectively inactivate E. coli vegetative cells, but not B. thuringiensis spores, which were more resistant to AOP. Further, the BioSentry in-line sensor was not successful in documenting destruction of the microbial cells in real-time.

  3. Control of Initiation of DNA Replication in Bacillus subtilis and Escherichia coli

    PubMed Central

    Jameson, Katie H.; Wilkinson, Anthony J.

    2017-01-01

    Initiation of DNA Replication is tightly regulated in all cells since imbalances in chromosomal copy number are deleterious and often lethal. In bacteria such as Bacillus subtilis and Escherichia coli, at the point of cytokinesis, there must be two complete copies of the chromosome to partition into the daughter cells following division at mid-cell during vegetative growth. Under conditions of rapid growth, when the time taken to replicate the chromosome exceeds the doubling time of the cells, there will be multiple initiations per cell cycle and daughter cells will inherit chromosomes that are already undergoing replication. In contrast, cells entering the sporulation pathway in B. subtilis can do so only during a short interval in the cell cycle when there are two, and only two, chromosomes per cell, one destined for the spore and one for the mother cell. Here, we briefly describe the overall process of DNA replication in bacteria before reviewing initiation of DNA replication in detail. The review covers DnaA-directed assembly of the replisome at oriC and the multitude of mechanisms of regulation of initiation, with a focus on the similarities and differences between E. coli and B. subtilis. PMID:28075389

  4. Control of Initiation of DNA Replication in Bacillus subtilis and Escherichia coli.

    PubMed

    Jameson, Katie H; Wilkinson, Anthony J

    2017-01-10

    Initiation of DNA Replication is tightly regulated in all cells since imbalances in chromosomal copy number are deleterious and often lethal. In bacteria such as Bacillus subtilis and Escherichia coli, at the point of cytokinesis, there must be two complete copies of the chromosome to partition into the daughter cells following division at mid-cell during vegetative growth. Under conditions of rapid growth, when the time taken to replicate the chromosome exceeds the doubling time of the cells, there will be multiple initiations per cell cycle and daughter cells will inherit chromosomes that are already undergoing replication. In contrast, cells entering the sporulation pathway in B. subtilis can do so only during a short interval in the cell cycle when there are two, and only two, chromosomes per cell, one destined for the spore and one for the mother cell. Here, we briefly describe the overall process of DNA replication in bacteria before reviewing initiation of DNA replication in detail. The review covers DnaA-directed assembly of the replisome at oriC and the multitude of mechanisms of regulation of initiation, with a focus on the similarities and differences between E. coli and B. subtilis.

  5. Current knowledge on isobutanol production with Escherichia coli, Bacillus subtilis and Corynebacterium glutamicum

    PubMed Central

    Eikmanns, Bernhard J

    2011-01-01

    Due to steadily rising crude oil prices great efforts have been made to develop designer bugs for the fermentative production of higher alcohols, such as 2-methyl-1-butanol, 3-methyl-1-butanol and 2-Methyl-1-propanol (isobutanol), which all possess quality characteristics comparable to traditional oil based fuels. The common metabolic engineering approach uses the last two steps of the Ehrlich pathway, catalyzed by 2-ketoacid decarboxylase and an alcohol dehydrogenase converting the branched chain 2-ketoacids of L-isoleucine, L-leucine and L-valine into the respective alcohols. This strategy was successfully used to engineer well suited and industrially employed bacteria, such as Escherichia coli, Bacillus subtilis and Corynebacterium glutamicum for the production of higher alcohols. Among these alcohols, isobutanol is currently the most promising one regarding final titer and yield. This article summarizes the current knowledge and achievements on isobutanol production with E. coli, B. subtilis and C. glutamicum regarding the metabolic engineering approaches and process conditions. PMID:22008938

  6. Current knowledge on isobutanol production with Escherichia coli, Bacillus subtilis and Corynebacterium glutamicum.

    PubMed

    Blombach, Bastian; Eikmanns, Bernhard J

    2011-01-01

    Due to steadily rising crude oil prices great efforts have been made to develop designer bugs for the fermentative production of higher alcohols, such as 2-methyl-1-butanol, 3-methyl-1-butanol and 2-Methyl-1-propanol (isobutanol), which all possess quality characteristics comparable to traditional oil based fuels. The common metabolic engineering approach uses the last two steps of the Ehrlich pathway, catalyzed by 2-ketoacid decarboxylase and an alcohol dehydrogenase converting the branched chain 2-ketoacids of L-isoleucine, L-leucine, and L-valine into the respective alcohols. This strategy was successfully used to engineer well suited and industrially employed bacteria, such as Escherichia coli, Bacillus subtilis and Corynebacterium glutamicum for the production of higher alcohols. Among these alcohols, isobutanol is currently the most promising one regarding final titer and yield. This article summarizes the current knowledge and achievements on isobutanol production with E. coli, B. subtilis and C. glutamicum regarding the metabolic engineering approaches and process conditions.

  7. In vitro transcription of the Bacillus subtilis phage phi 29 DNA by Bacillus subtilis and Escherichia coli RNA polymerases.

    PubMed Central

    Sogo, J M; Lozano, M; Salas, M

    1984-01-01

    The Escherichia coli RNA polymerase bound to phage phi 29 DNA has been visualized by electron microscopy. Thirteen specific binding sites have been observed at 1.7,2.6,5.5,10.4,13.7,25.2,25.7,26.3,33.5,59.5,69.2,91.7 and 99.6 DNA length units and they have been named A1,A1I,A1II,A1III,A1IV,A2,A2I, A3, A4,B1,B1I,C1 and C2, respectively. The binding sites A1,A2,A3,B1,C1 and C2 coincide with those found with Bacillus subtilis RNA polymerase. The transcription of phage phi 29 DNA with B. subtilis or E. coli RNA polymerases has been studied. With the B. subtilis RNA polymerase eight transcripts were found, starting at positions corresponding to the binding sites A1, A1III, A2,A3,B1I,B2,C1 and C2, respectively. With the E. coli RNA polymerase the same transcripts were found and a new one starting at position corresponding to the A4 binding site. The RNAs starting at binding sites A1,A1III,A2,B1I, B2,C1 and C2 are transcribed from right to left, as expected for early RNA. The RNAs which initiate at positions A3 and A4 are transcribed from left to right and probably correspond to late RNAs. Images PMID:6322128

  8. Combining Genes from Multiple Phages for Improved Cell Lysis and DNA Transfer from Escherichia coli to Bacillus subtilis

    PubMed Central

    Juhas, Mario; Wong, Christine; Ajioka, James W.

    2016-01-01

    The ability to efficiently and reliably transfer genetic circuits between the key synthetic biology chassis, such as Escherichia coli and Bacillus subtilis, constitutes one of the major hurdles of the rational genome engineering. Using lambda Red recombineering we integrated the thermosensitive lambda repressor and the lysis genes of several bacteriophages into the E. coli chromosome. The lysis of the engineered autolytic cells is inducible by a simple temperature shift. We improved the lysis efficiency by introducing different combinations of lysis genes from bacteriophages lambda, ΦX174 and MS2 under the control of the thermosensitive lambda repressor into the E. coli chromosome. We tested the engineered autolytic cells by transferring plasmid and bacterial artificial chromosome (BAC)-borne genetic circuits from E. coli to B. subtilis. Our engineered system combines benefits of the two main synthetic biology chassis, E. coli and B. subtilis, and allows reliable and efficient transfer of DNA edited in E. coli into B. subtilis. PMID:27798678

  9. Escherichia Coli

    ERIC Educational Resources Information Center

    Goodsell, David S.

    2009-01-01

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

  10. Escherichia Coli

    ERIC Educational Resources Information Center

    Goodsell, David S.

    2009-01-01

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

  11. Expression in Escherichia coli of a cloned crystal protein gene of Bacillus thuringiensis subsp. israelensis.

    PubMed Central

    McLean, K M; Whiteley, H R

    1987-01-01

    A ca. 10-kilobase (kb) HindIII fragment of plasmid DNA from Bacillus thuringiensis subsp. israelensis was cloned into plasmid pUC9 and transformed into Escherichia coli. Extracts of the recombinant strain contained a 27-kilodalton (kDa) peptide that reacted with antibodies to a 27-kDa peptide isolated from crystals produced by B. thuringiensis subsp. israelensis. Extracts of the recombinant strain were hemolytic and toxic to Aedes aegypti larvae. Full expression of the 27-kDa peptide required the presence of a ca. 0.8-kb region of DNA located 4 kb upstream from the structural gene; the 0.8-kb region could be present in cis or trans relative to the gene and apparently acted post-transcriptionally. Analysis of maxicells showed that the 10-kb insert also coded for peptides of 67, 20, and 16 kDa; data obtained with different subclones suggest that the 20-kDa peptide is encoded in the 0.8-kb DNA region. Images PMID:3546262

  12. Mixed culture models for predicting intestinal microbial interactions between Escherichia coli and Lactobacillus in the presence of probiotic Bacillus subtilis.

    PubMed

    Yang, J J; Niu, C C; Guo, X H

    2015-01-01

    Bacillus has been proposed as a probiotic due to its in vivo effectiveness in the gastrointestinal tract through antimicrobial activities. The present study investigates the effects of Lactobacillus alone or in the presence of Bacillus subtilis MA139 on the inhibition of pathogenic Escherichia coli K88. Mixed cultures were used to predict the possible interactions among these bacteria within the intestinal tract of animals. B. subtilis MA139 was first assayed for its inhibition against E. coli K88 both under shaking and static culture conditions. A co-culture assay was employed under static conditions to test the inhibitory effects of Lactobacillus reuteri on E. coli K88, with or without addition of B. subtilis MA139. The results showed that B. subtilis MA139 had marked inhibition against E. coli K88 under shaking conditions and weak inhibition under static conditions. Lactobacillus alone as well as in combination with B. subtilis MA139 spores exerted strong inhibition against E. coli K88 under static conditions. However, the inhibition by Lactobacillus in combination with B. subilis spores was much higher than that by Lactobacillus alone (P<0.01). B. subtilis MA139 significantly decreased the pH and oxidation-reduction potential values of the co-culture broth compared to that of Lactobacillus alone (P<0.05). The viability of Lactobacillus increased when co-cultured with B. subtilis MA139 because of significantly higher Lactobacillus counts and lower pH values in the broth (P<0.05). The role of Bacillus in the mixed culture models suggests that Bacillus may produce beneficial effects by increasing the viability of lactobacilli and subsequently inhibiting the growth of pathogenic E. coli. Therefore, the combination of Bacillus and Lactobacillus species as a probiotic is recommended.

  13. Peptide-based communication system enables Escherichia coli to Bacillus megaterium interspecies signaling.

    PubMed

    Marchand, Nicholas; Collins, Cynthia H

    2013-11-01

    The use of mixtures of microorganisms, or microbial consortia, has the potential to improve the productivity and efficiency of increasingly complex bioprocesses. However, the use of microbial consortia has been limited by our ability to control and coordinate the behaviors of microorganisms in synthetic communities. Synthetic biologists have previously engineered cell-cell communication systems that employ machinery from bacterial quorum-sensing (QS) networks to enable population-level control of gene expression. However, additional communication systems, such as those that enable communication between different species of bacteria, are needed to enable the use of diverse species in microbial consortia for bioprocessing. Here, we use the agr QS system from Staphylococcus aureus to generate an orthogonal synthetic communication system between Gram-negative Escherichia coli and Gram-positive Bacillus megaterium that is based on the production and recognition of autoinducing peptides (AIPs). We describe the construction and characterization of two types of B. megaterium "receiver" cells, capable of AIP-dependent gene expression in response to AIPs that differ by a single amino acid. Further, we observed interspecies communication when these receiver cells were co-cultured with AIP-producing E. coli. We show that the two AIP-based systems exhibit differences in sensitivity and specificity that may be advantageous in tuning communication-dependent networks in synthetic consortia. These peptide-based communication systems will enable the coordination of gene expression, metabolic pathways and growth between diverse microbial species, and represent a key step towards the use of microbial consortia in bioprocessing and biomanufacturing.

  14. Mosquito larvicidal activity of Escherichia coli with combinations of genes from Bacillus thuringiensis subsp. israelensis.

    PubMed Central

    Ben-Dov, E; Boussiba, S; Zaritsky, A

    1995-01-01

    The genes cryIVA and cryIVD, encoding 134- and 72-kDa proteins, respectively, and the gene for a regulatory 20-kDa polypeptide of Bacillus thuringiensis subsp. israelensis (serovar H14) were cloned in all seven possible combinations by the Escherichia coli expression vectors pT7 and pUHE. The four combinations containing cryIVA (cryIVA alone, with cryIVD, with the 20-kDa-protein gene, and with both) displayed high levels of mosquito larvicidal activity in pUHE. The toxicity of the combination of cryIVA and cryIVD, with or without the 20-kDa-protein gene, was higher than has ever been achieved with delta-endotoxin genes in recombinant E. coli. Fifty percent lethal concentrations against third-instar Aedes aegypti larvae for these clones decreased (i.e., toxicity increased) continuously to about 3 x 10(5) cells ml-1 after 4 h of induction. Larvicidal activities, obtained after 30 min of induction, were lower for clones in pT7 and decreased for an additional 3.5 h. Induction of either cryIVD or the 20-kDa-protein gene alone resulted in no larvicidal activity in either pT7 or pUHE20. Cloned together, these genes were slightly toxic in pT7 but not in pUHE20. Five minutes of induction of this combination (cryIVD with the 20-kDa-protein gene) in pT7 yielded a maximal mortality of about 40%, which decreased rapidly and disappeared completely after 50 min. CryIVD is thus apparently degraded in E. coli and partially stabilized by the 20-kDa regulatory protein. Larvicidal activity of the combination of cryIVA and cryIVD was sevenfold higher than that of cryIVA alone, probably because of the cross-stabilization of the polypeptides or the synergism between their activities. PMID:7751296

  15. Fine Encapsulated ZnO Nanophosphors And Their Potential Antibacterial Evaluation On The Gram Negative Bacillus Escherichia coli

    NASA Astrophysics Data System (ADS)

    Dutta, Ranu K.; Sharma, Prashant K.; Pandey, Avinash C.

    2009-06-01

    The spherical nanophosphors of ZnO of mean size 10-20 nm were synthesized at room temperature by simple co-precipitation method. Their size and shape were governed by several encapsulations using compounds like biotin, citric acid and Poly vinyl alcohol (PVA). Besides playing a critical role in determining their size and shape, these biomolecules served as effective capping agents. The antibacterial behavior of the suspension of fine ZnO nanophosphors was investigated against the gram negative bacillus Escherichia coli. The growth curve showed bacteriostatic activity against Escherichia coli. The antibacterial activities of the ZnO nanophosphors can be attributed to their total surface area, as increasing surface to volume ratio of nanophosphors provides more efficient means for enhanced antibacterial activity.

  16. Purification and characterization of a Bacillus polymyxa beta-glucosidase expressed in Escherichia coli.

    PubMed Central

    Painbeni, E; Valles, S; Polaina, J; Flors, A

    1992-01-01

    The beta-glucosidase encoded by the bglA gene from Bacillus polymyxa was overproduced in Escherichia coli by using a plasmid in which bglA is under control of the lacI promoter. Induction with isopropyl-beta-D-thiogalactopyranoside allowed an increase in the specific activity of the enzyme of about 100 times the basal level of gene expression. The enzyme was purified by a two-step procedure involving salting out with ammonium sulfate and ion-exchange chromatography with DEAE-cellulose. Fractions of beta-glucosidase activity recovered by this procedure, after electrophoresis in an acrylamide gel and staining with silver nitrate, yielded a single band of protein. This band was shown by a zymogram to correspond to beta-glucosidase activity. The purified protein showed an apparent molecular mass of 50 kDa and an isoelectric point of 4.6, values in agreement with those expected from the nucleotide sequence of the gene. Km values of the enzyme, with either cellobiose or p-nitrophenyl-beta-D-glucoside as the substrate, were determined. It was shown that the enzyme is competitively inhibited by glucose. The effects of different metallic ions and other agents were studied. Hg2+ was strongly inhibitory, while none of the other cations tested had any significant effect. Ethanol did not show the stimulating effect observed with other beta-glucosidases. The mechanism of enzyme action was investigated. High-pressure liquid chromatography analysis with cellobiose as the substrate confirmed previous data revealing the formation of two products, glucose and another, unidentified, compound. Results presented here indicate that this compound is cellotriose formed by transglycosylation. Images PMID:1569036

  17. Cloning and characterization of two genes from Bacillus polymyxa expressing beta-glucosidase activity in Escherichia coli.

    PubMed Central

    González-Candelas, L; Aristoy, M C; Polaina, J; Flors, A

    1989-01-01

    DNA fragments from Bacillus polymyxa which encode beta-glucosidase activity were cloned in Escherichia coli by selection of yellow transformants able to hydrolyze the artificial chromogenic substrate p-nitrophenyl-beta-D-glucopyranoside. Restriction endonuclease maps and Southern analysis of the cloned fragments showed the existence of two different genes. Expression of either one of these genes allowed growth of E. coli in minimal medium with cellobiose as the only carbon source. One of the two enzymes was found in the periplasm of E. coli, hydrolyzed arylglucosides more actively than cellobiose, and rendered glucose as the only product upon cellobiose hydrolysis. The other enzyme was located in the cytoplasm, was more active toward cellobiose, and hydrolyzed this disaccharide, yielding glucose and another, unidentified compound, probably a phosphorylated sugar. Images PMID:2515802

  18. Acetolactate synthase from Bacillus subtilis serves as a 2-ketoisovalerate decarboxylase for isobutanol biosynthesis in Escherichia coli.

    PubMed

    Atsumi, Shota; Li, Zhen; Liao, James C

    2009-10-01

    A pathway toward isobutanol production previously constructed in Escherichia coli involves 2-ketoacid decarboxylase (Kdc) from Lactococcus lactis that decarboxylates 2-ketoisovalerate (KIV) to isobutyraldehyde. Here, we showed that a strain lacking Kdc is still capable of producing isobutanol. We found that acetolactate synthase from Bacillus subtilis (AlsS), which originally catalyzes the condensation of two molecules of pyruvate to form 2-acetolactate, is able to catalyze the decarboxylation of KIV like Kdc both in vivo and in vitro. Mutational studies revealed that the replacement of Q487 with amino acids with small side chains (Ala, Ser, and Gly) diminished only the decarboxylase activity but maintained the synthase activity.

  19. Primary structure, expression in Escherichia coli, and properties of S-adenosyl-L-methionine:uroporphyrinogen III methyltransferase from Bacillus megaterium.

    PubMed Central

    Robin, C; Blanche, F; Cauchois, L; Cameron, B; Couder, M; Crouzet, J

    1991-01-01

    A Bacillus megaterium DNA fragment encoding S-adenosyl-L-methionine:uroporphyrinogen III methyltransferase (SUMT) activity was subcloned and sequenced. The encoded polypeptide showed more than 43.5% strict homology to Pseudomonas denitrificans SUMT (F. Blanche, L. Debussche, D. Thibaut, J. Crouzet, and B. Cameron, J. Bacteriol. 171:4222-4231, 1989). The B. megaterium polypeptide was overexpressed in Escherichia coli, partially purified, and shown to exhibit, like P. denitrificans SUMT, substrate inhibition at uroporphyrinogen III concentrations above 0.5 microM, suggesting a common regulation for aerobic cobalamin-producing organisms. Images PMID:1906874

  20. [Purification of recombinant Bacillus cereus ResD-ResE proteins expressed in Escherichia coli strains].

    PubMed

    Shapyrina, E V; Shadrin, A M; Solonin, A S

    2013-01-01

    Recombinant E. coli strains expressing the Bacillus cereus ATCC 14579T resD and resEgenes fused with the ubiquitin gene were constructed, and purification of the ResD and ResE proteins was performed. The approach used in the study allowed us to increase the protein yield of the electrophoretic homogeneous ResD andResE proteins without denaturation steps up to 150 mg per gram of wet cell weight.

  1. Differential analysis of bactericidal systems of blood serum with recombinant luminescent Escherichia coli and Bacillus subtilis strains.

    PubMed

    Deryabin, D G; Karimov, I F; Manukhov, I V; Tolmacheva, N A; Balabanov, V P

    2012-11-01

    Luminescence intensity of recombinant Escherichia coli and Bacillus subtilis strains with cloned luxCD(AB)E genes of the natural luminescent microorganism Photobacterium leiognathi was studied under the influence of 30 individual samples of human blood serum of different component composition. A relationship was found between the level of residual bioluminescence and degree of the bactericidal effect. Moreover, the inhibition of E. coli lux+ luminescence was shown to be related to activity of the complement-lysozyme system. The reaction of B. subtilis lux+ primarily depended on the presence of β-lysin in the blood serum. These data provide an experimental substantiation of a new method of differential analysis of humoral factors of nonspecific innate immunity with recombinant luminescent bacteria.

  2. Structural and catalytic properties of CMP kinase from Bacillus subtilis: a comparative analysis with the homologous enzyme from Escherichia coli.

    PubMed

    Schultz, C P; Ylisastigui-Pons, L; Serina, L; Sakamoto, H; Mantsch, H H; Neuhard, J; Bârzu, O; Gilles, A M

    1997-04-01

    CMP kinases from Bacillus subtilis and from Escherichia coli are encoded by the cmk gene (formerly known as jofC in B. subtilis and as mssA in E. coli). Similar in their primary structure (43% identity and 67% similarity in amino acid sequence), the two proteins exhibit significant differences in nucleotide binding and catalysis. ATP, dATP, and GTP are equally effective as phosphate donors with E. coli CMP kinase whereas GTP is a poor substrate with B. subtilis CMP kinase. While CMP and dCMP are the best phosphate acceptors of both CMP kinases, the specific activity with these substrates and ATP as donor are 7- to 10-fold higher in the E. coli enzyme; the relative Vm values with UMP and CMP are 0.1 for the B. subtilis CMP kinase and 0.01 for the E. coli enzyme. CMP increased the affinity of E. coli CMP kinase for ATP or for the fluorescent analog 3'-anthraniloyl dATP by one order of magnitude but had no effect on the B. subtilis enzyme. The differences in the catalytic properties of B. subtilis and E. coli CMP kinases might be reflected in the structure of the two proteins as inferred from infrared spectroscopy. Whereas the spectrum of B. subtilis CMP kinase is dominated by a band at 1633 cm-1 (representing beta type structures), the spectrum of the E. coli enzyme is dominated by two bands at 1653 and 1642 cm-1 associated with alpha-helical and unordered structures, respectively. CMP induced similar spectral changes in both proteins with a rearrangement of some of the beta-structures. ATP increases the denaturation temperature of B. subtilis CMP kinase by 9.3 degrees C, whereas in the case of the E. coli enzyme, binding of ATP has only a minor effect.

  3. Synthesis and processing of Escherichia coli TEM-beta-lactamase and Bacillus licheniformis alpha-amylase in E. coli: the role of signal peptidase I.

    PubMed

    van Dijl, J M; Smith, H; Bron, S; Venema, G

    1988-09-01

    A mutant of Escherichia coli, in which signal peptidase I synthesis can be regulated, was constructed. The mutant was used to study the effects of signal peptidase I limitation on the synthesis and efficiency of processing of two proteins: the periplasmic E. coli TEM-beta-lactamase and Bacillus licheniformis alpha-amylase, which also accumulates in the periplasm of E. coli. Signal peptidase I limitation resulted in reduced rates of processing of pre-beta-lactamase and in strong inhibition of synthesis of alpha-amylase. The data suggest that beta-lactamase is processed post-translationally and that an intimate relationship exists between the synthesis and processing of alpha-amylase.

  4. Cloning and nucleotide sequencing of a novel 7 beta-(4-carboxybutanamido)cephalosporanic acid acylase gene of Bacillus laterosporus and its expression in Escherichia coli and Bacillus subtilis.

    PubMed

    Aramori, I; Fukagawa, M; Tsumura, M; Iwami, M; Ono, H; Kojo, H; Kohsaka, M; Ueda, Y; Imanaka, H

    1991-12-01

    A strain of Bacillus species which produced an enzyme named glutaryl 7-ACA acylase which converts 7 beta-(4-carboxybutanamido)cephalosporanic acid (glutaryl 7-ACA) to 7-amino cephalosporanic acid (7-ACA) was isolated from soil. The gene for the glutaryl 7-ACA acylase was cloned with pHSG298 in Escherichia coli JM109, and the nucleotide sequence was determined by the M13 dideoxy chain termination method. The DNA sequence revealed only one large open reading frame composed of 1,902 bp corresponding to 634 amino acid residues. The deduced amino acid sequence contained a potential signal sequence in its amino-terminal region. Expression of the gene for glutaryl 7-ACA acylase was performed in both E. coli and Bacillus subtilis. The enzyme preparations purified from either recombinant strain of E. coli or B. subtilis were shown to be identical with each other as regards the profile of sodium dodecyl sulfate-polyacrylamide gel electrophoresis and were composed of a single peptide with the molecular size of 70 kDa. Determination of the amino-terminal sequence of the two enzyme preparations revealed that both amino-terminal sequences (the first nine amino acids) were identical and completely coincided with residues 28 to 36 of the open reading frame. Extracellular excretion of the enzyme was observed in a recombinant strain of B. subtilis.

  5. Cloning and nucleotide sequencing of a novel 7 beta-(4-carboxybutanamido)cephalosporanic acid acylase gene of Bacillus laterosporus and its expression in Escherichia coli and Bacillus subtilis.

    PubMed Central

    Aramori, I; Fukagawa, M; Tsumura, M; Iwami, M; Ono, H; Kojo, H; Kohsaka, M; Ueda, Y; Imanaka, H

    1991-01-01

    A strain of Bacillus species which produced an enzyme named glutaryl 7-ACA acylase which converts 7 beta-(4-carboxybutanamido)cephalosporanic acid (glutaryl 7-ACA) to 7-amino cephalosporanic acid (7-ACA) was isolated from soil. The gene for the glutaryl 7-ACA acylase was cloned with pHSG298 in Escherichia coli JM109, and the nucleotide sequence was determined by the M13 dideoxy chain termination method. The DNA sequence revealed only one large open reading frame composed of 1,902 bp corresponding to 634 amino acid residues. The deduced amino acid sequence contained a potential signal sequence in its amino-terminal region. Expression of the gene for glutaryl 7-ACA acylase was performed in both E. coli and Bacillus subtilis. The enzyme preparations purified from either recombinant strain of E. coli or B. subtilis were shown to be identical with each other as regards the profile of sodium dodecyl sulfate-polyacrylamide gel electrophoresis and were composed of a single peptide with the molecular size of 70 kDa. Determination of the amino-terminal sequence of the two enzyme preparations revealed that both amino-terminal sequences (the first nine amino acids) were identical and completely coincided with residues 28 to 36 of the open reading frame. Extracellular excretion of the enzyme was observed in a recombinant strain of B. subtilis. Images FIG. 2 FIG. 5 FIG. 6 PMID:1744041

  6. Genetic determinant of Bacillus pumilus lipase lethality and its application as positive selection cloning vector in Escherichia coli.

    PubMed

    Mabizela-Mokoena, Nobalanda Betty; Limani, Shonisani Wendy; Ncube, Ignatious; Piater, Lizelle Ann; Litthauer, Derek; Nthangeni, Mulalo Bethuel

    2017-09-01

    Positive selection vectors carry genes that upon expression produce proteins that cause host cell deaths. Insertion of foreign DNA fragments within the ORF of the gene disrupts the lethal effect of the expressed protein. This study described the cloning of Family I.4 Bacillus pumilus lipase gene whose expressed protein is toxic and lethal to Escherichia coli JM109 (DE3) cells. The determinant of toxicity was identified through Error-prone PCR to be the nature of amino acid residue resident at position 28 of the mature lipase protein. The presence of Thr/Ser28 within the mature lipases of B. pumilus and B. licheniformis resulted in lethality to E. coli cells. However, the Thr28Ala or Thr28Gly mutations relieved the lethal phenotype of mature Family I.4 Bacillus lipases. The toxic effect of the expressed mature B. pumilus lipase protein was exploited in the development of a positive selection cloning vector. The B. pumilus lipase gene was synthesised to contain 13 unique silent restriction sites within the ORF, and placed under the regulation of T7 promoter of the pET expression system. Insertional inactivation of the gene's toxic protein was achieved by cloning DNA fragments of different sizes within the designed multiple cloning sites. The toxic effect of the lipase protein was disrupted indicating the potential of the gene for application in suicidal positive selection cloning vectors. The results revealed that protein expression and engineering studies aimed at optimal production of mature Family I.4 Bacillus lipases in E. coli should take into consideration the nature of amino acid 28 resident. Copyright © 2017 Elsevier Inc. All rights reserved.

  7. The effect of ribosomal protein S1 from Escherichia coli and Micrococcus luteus on protein synthesis in vitro by E. coli and Bacillus subtilis.

    PubMed

    Farwell, M A; Roberts, M W; Rabinowitz, J C

    1992-11-01

    We have designed a set of nine plasmids containing the Bacillus pumilis cat gene with one of three Shine-Dalgarno (SD) sequences (weak, strong or stronger) and one of three initiation codons (AUG, GUG or UUG). These constructions have been used to determine the effect of ribosomal protein S1, SD and initiation codon sequences and Escherichia coli ribosomal protein S1 on translation in vitro by E. coli and B. subtilis ribosomes. Translation of these nine constructions was determined with three types of ribosomes: E. coli containing ribosomal protein S1, E. coli depleted of S1, and B. subtilis which is naturally free of S1. E. coli ribosomes were able to translate all nine transcripts with variable efficiencies. B. subtilis and S1-depleted E. coli ribosomes were similar to each other and differed from non-depleted E. coli ribosomes in that they required strong or stronger SD sequences and were unable to translate any of the weak transcripts. Addition of S1 from either E. coli or Micrococcus luteus, a Gram-positive bacterium, enabled S1-depleted E. coli ribosomes to translate mRNAs with weak SD sequences but had no effect on B. subtilis ribosomes. AUG was the preferred initiation codon for all ribosome types; however, B. subtilis ribosomes showed greater tolerance for the non-AUG codons than either type of E. coli ribosome. The presence of a strong or stronger SD sequence increased the efficiency by which E. coli ribosomes could utilize non-AUG codons.(ABSTRACT TRUNCATED AT 250 WORDS)

  8. How Changes in Anti-SD Sequences Would Affect SD Sequences in Escherichia coli and Bacillus subtilis.

    PubMed

    Abolbaghaei, Akram; Silke, Jordan R; Xia, Xuhua

    2017-03-31

    The 3' end of the small ribosomal RNAs (ssu rRNA) in bacteria is directly involved in the selection and binding of mRNA transcripts during translation initiation via well-documented interactions between a Shine-Dalgarno (SD) sequence located upstream of the initiation codon and an anti-SD (aSD) sequence at the 3' end of the ssu rRNA. Consequently, the 3' end of ssu rRNA (3'TAIL) is strongly conserved among bacterial species because a change in the region may impact the translation of many protein-coding genes. Escherichia coli and Bacillus subtilis differ in their 3' ends of ssu rRNA, being GAUCACCUCCUUA3' in E. coli and GAUCACCUCCUUUCU3' or GAUCACCUCCUUUCUA3' in B. subtilis Such differences in 3'TAIL lead to species-specific SDs (designated SDEc for E. coli and SDBs for B. subtilis) that can form strong and well-positioned SD/aSD pairing in one species but not in the other. Selection mediated by the species-specific 3'TAIL is expected to favour SDBs against SDEc in B. subtilis but favour SDEc against SDBs in E. coli Among well-positioned SDs, SDEc is used more in E. coli than in B. subtilis, and SDBs more in B. subtilis than in E. coli Highly expressed genes and genes of high translation efficiency tend to have longer SDs than lowly expressed genes and genes with low translation efficiency in both species, but more so in B. subtilis than in E. coli Both species overuse SDs matching the bolded part of 3'TAIL shown above. The 3'TAIL difference contributes to host-specificity of phages.

  9. Synthetic operon for (R,R)-2,3-butanediol production in Bacillus subtilis and Escherichia coli.

    PubMed

    de Oliveira, Rafael R; Nicholson, Wayne L

    2016-01-01

    To reduce dependence on petroleum, an alternative route to production of the chemical feedstock 2,3-butanediol (2,3-BD) from renewable lignocellulosic sources is desirable. In this communication, the genes encoding the pathway from pyruvate to 2,3-BD (alsS, alsD, and bdhA encoding acetolactate synthase, acetolactate decarboxylase, and butanediol dehydrogenase, respectively) from Bacillus subtilis were engineered into a single tricistronic operon under control of the isopropyl β-D-1-thiogalactopyranoside (IPTG)-inducible Pspac promoter in a shuttle plasmid capable of replication and expression in either B. subtilis or Escherichia coli. We describe the construction and performance of a shuttle plasmid carrying the IPTG-inducible synthetic operon alsSDbdhA coding for 2,3-BD pathway capable of (i) expression in two important representative model microorganisms, the gram-positive B. subtilis and the gram-negative E. coli; (ii) increasing 2,3-BD production in B. subtilis; and (iii) successfully introducing the B. subtilis 2,3-BD pathway into E. coli. The synthetic alsSDbdhA operon constructed using B. subtilis native genes not only increased the 2,3-BD production in its native host but also efficiently expressed the pathway in the heterologous organism E. coli. Construction of an efficient shuttle plasmid will allow investigation of 2,3-BD production performance in related organisms with industrial potential for production of bio-based chemicals.

  10. Bacillus subtilis and Escherichia coli essential genes and minimal cell factories after one decade of genome engineering.

    PubMed

    Juhas, Mario; Reuß, Daniel R; Zhu, Bingyao; Commichau, Fabian M

    2014-11-01

    Investigation of essential genes, besides contributing to understanding the fundamental principles of life, has numerous practical applications. Essential genes can be exploited as building blocks of a tightly controlled cell 'chassis'. Bacillus subtilis and Escherichia coli K-12 are both well-characterized model bacteria used as hosts for a plethora of biotechnological applications. Determination of the essential genes that constitute the B. subtilis and E. coli minimal genomes is therefore of the highest importance. Recent advances have led to the modification of the original B. subtilis and E. coli essential gene sets identified 10 years ago. Furthermore, significant progress has been made in the area of genome minimization of both model bacteria. This review provides an update, with particular emphasis on the current essential gene sets and their comparison with the original gene sets identified 10 years ago. Special attention is focused on the genome reduction analyses in B. subtilis and E. coli and the construction of minimal cell factories for industrial applications.

  11. Cloning and heterologous expression of ectoine biosynthesis genes from Bacillus halodurans in Escherichia coli.

    PubMed

    Anbu Rajan, Lawrance; Joseph, Toms C; Thampuran, Nirmala; James, Roswin; Ashok Kumar, Kesavan; Viswanathan, Chinnusamy; Bansal, Kailash C

    2008-08-01

    The genes involved in the biosynthetic pathway of ectoine (2-methyl-1,4,5,6-tetrahydropyrimidine-4-carboxylic acid) from Bacillus halodurans were cloned as an operon and expressed in E. coli. Analysis of the deduced ectoine biosynthesis cluster amino acid sequence revealed that the ectoine operon contain 2,389 bp, encoded by three genes; ectA, ectB and ectC that encode proteins of 189, 427 and 129 amino acids with deduced molecular masses of 21,048, 47,120 and 14,797 Da respectively. Extracts of induced cells showed two bands at 41 kDa and 17 kDa, possibly corresponding to the products of the later two genes. However the expression of ectA gene could not be ascertained by SDS-PAGE. The activity of the ectA protein was confirmed by an acylation assay. The transgenic E. coli accumulated upto 4.6 mg ectoine/l culture. This is the first report of an engineered E. coli strain carrying the ectoine genes of the alkaliphilic bacterium, B. halodurans.

  12. Toward a bacterial genome technology: integration of the Escherichia coli prophage lambda genome into the Bacillus subtilis 168 chromosome.

    PubMed

    Itaya, M

    1995-07-22

    A novel approach to the cloning large DNAs in the Bacillus subtilis chromosome was examined. An Escherichia coli prophage lambda DNA (48.5 kb) was assembled in the chromosome of B. subtilis. The lambda DNA was first subcloned in four segments, having partially overlapping regions. Assembly of the complete prophage was achieved by successive transformation using three discrete DNA integration modes: overlap-elongation, Campbell-type integration, and gap-filling. In the B. subtilis chromosome, DNA was elongated, using contiguous DNA segments, via overlap-elongation. Jumping from one end of a contiguous DNA stretch to another segment was achieved by Campbell-type integration. The remaining gap was sealed by gap-filling. The incorporated lambda DNA thus assembled was stably replicated as part of the 4188 kb B. subtilis chromosome under non-selective conditions. The present method can be used to accommodate larger DNAs in the B. subtilis chromosome and possible applications of this technique are discussed.

  13. Cloning, overexpression and purification of Bacillus subtilis elongation factor Tu in Escherichia coli.

    PubMed

    Kim, S I; Kim, H Y; Kwak, J H; Kwon, S H; Lee, S Y

    2000-02-29

    To establish the overexpression and one-step purification system of Bacillus subtilis elongation factor-Tu (EF-Tu), the EF-Tu gene was amplified with or without own ribosome binding site (rbs) by PCR and the only PCR product without rbs was subcloned successfully. For the expression of the EF-Tu gene cloned after PCR amplification, a constitutive expression system and inducible expression system with His6 tag at N-terminus or C-terminus, or glutathione-S-transferase (GST) fusion system were examined in E. coli and B. subtilis. Except GST fusion system in E. coli, however, all other trials were unsuccessful at the step of plasmid construction for the EF-Tu expression. The GST/EF-Tu fusion proteins were highly expressed by IPTG induction and obtained as both soluble and insoluble form. From the soluble GST/EF-Tu fusion protein, EF-Tu was obtained to near homogeneity by one-step purification with glutathione-sepharose affinity column chromatography followed by factor Xa treatment. The purified EF-Tu showed high GDP binding activity. These results indicate that the GST/EF-Tu fusion system is favorable to overexpression and purification of B. subtilis EF-Tu.

  14. Initiating nucleotide identity determines efficiency of RNA synthesis from 6S RNA templates in Bacillus subtilis but not Escherichia coli

    PubMed Central

    Cabrera-Ostertag, Ignacio J.; Cavanagh, Amy T.; Wassarman, Karen M.

    2013-01-01

    The 6S RNA is a non-coding small RNA that binds within the active site of housekeeping forms of RNA polymerases (e.g. Eσ70 in Escherichia coli, EσA in Bacillus subtilis) and regulates transcription. Efficient release of RNA polymerase from 6S RNA regulation during outgrowth from stationary phase is dependent on use of 6S RNA as a template to generate a product RNA (pRNA). Interestingly, B. subtilis has two 6S RNAs, 6S-1 and 6S-2, but only 6S-1 RNA appears to be used efficiently as a template for pRNA synthesis during outgrowth. Here, we demonstrate that the identity of the initiating nucleotide is particularly important for the B. subtilis RNA polymerase to use RNA templates. Specifically, initiation with guanosine triphosphate (GTP) is required for efficient pRNA synthesis, providing mechanistic insight into why 6S-2 RNA does not support robust pRNA synthesis as it initiates with adenosine triphosphate (ATP). Intriguingly, E. coli RNA polymerase does not have a strong preference for initiating nucleotide identity. These observations highlight an important difference in biochemical properties of B. subtilis and E. coli RNA polymerases, specifically in their ability to use RNA templates efficiently, which also may reflect the differences in GTP and ATP metabolism in these two organisms. PMID:23761441

  15. [Investigation of ampicillin effect on morphological and mechanical properties of Escherichia coli and Bacillus cereus cells with atomic force microscopy].

    PubMed

    Deriabin, D G; Vasil'chenko, A S; Nikiian, A N

    2011-01-01

    The effect of subbacteriostatic concentrations of ampicillin on morphological and mechanical properties of gramnegative and grampositive cells of Escherichia coli K12 TG1 and Bacillus cereus IP 5832 respectively was studied with atomic force microscopy. Significant heterogeneity of the bacterial populations was shown by the character of the response to the antibiotic effect. The common feature was increase of the cell size likely due to the effect of the inner osmotic pressure on the lowered cell wall strength. In the E. coli population there were besides observed anomalous elongated cells with signs of septation disorder, as well as their structurs, lacking the cytoplasmic liquid fraction. In the B. cereus the inner osmotic pressure mainly enlarged the cell cross section, changing the cell shape from rod to sphere, that was accompanied by significant impairment of the surface structure with liberation of the peptidoglycane fragments to the medium. The particular features of the E. coli K12 TG1 and B. cereus IP 5832 respond to the ampicillin effect were attributed to the differences in the structure of their cell wall, also due to specific properties of the peptidoglycane synthesis and three-dimensional organization.

  16. Heterologous expression of the Bacillus subtilis (natto) alanine dehydrogenase in Escherichia coli and Lactococcus lactis.

    PubMed

    Ye, Wei; Huo, Guicheng; Chen, Junliang; Liu, Fei; Yin, Jingyuan; Yang, Lijie; Ma, Xiaolong

    2010-05-30

    The major objective of the present study is to change the alanine production of Lactic acid bacteria by expression of Bacillus subtilis (natto) alanine dehydrogenase (AlaDH), the gene that is not present in Lactic acid. B. subtilis AlaDH gene (ald) was cloned into a pGEX6p-1 and expressed in E. coli JM109. Its enzyme activity was 48.3U/mg at 30 degrees C and 45.2U/mg at 42 degrees C. This ald gene was then cloned into a vector pNZ8148 to generate a vector pNZ8148/ald. The same ald gene was placed downstream of the ldh promoter from Streptococcus thermophilus to generate pNZ273/ldhp/ald. The pNZ8148/ald and pNZ273/ldhp/ald were introduced separately in Lactococcus lactis NZ9000. As a result of over-expressed ald, the production of alanine detected by HPLC in L. lactis NZ9000 carrying pNZ273/ldhp/ald reached 52mug/ml, an approximately 26-fold increase compared to the parent strain L. lactis NZ9000, but not in L. lactis NZ9000 carrying pNZ8148/ald. This study would help strain improvement to be used in dairy fermentation for developing healthy yogurts with sweet taste or other fermented dairy foods. Copyright 2009 Elsevier GmbH. All rights reserved.

  17. A thermostable phytase from Bacillus sp. MD2: cloning, expression and high-level production in Escherichia coli.

    PubMed

    Tran, Thi Thuy; Mamo, Gashaw; Mattiasson, Bo; Hatti-Kaul, Rajni

    2010-03-01

    Phytase is used as a feed additive for degradation of antinutritional phytate, and the enzyme is desired to be highly thermostable for it to withstand feed formulation conditions. A Bacillus sp. MD2 showing phytase activity was isolated, and the phytase encoding gene was cloned and expressed in Escherichia coli. The recombinant phytase exhibited high stability at temperatures up to 100 degrees C. A higher enzyme activity was obtained when the gene expression was done in the presence of calcium chloride. Production of the enzyme by batch- and fed-batch cultivation in a bioreactor was studied. In batch cultivation, maintaining dissolved oxygen at 20-30% saturation and depleting inorganic phosphate below 1 mM prior to induction by IPTG resulted in over 10 U/ml phytase activity. For fed-batch cultivation, glucose concentration was maintained at 2-3 g/l, and the phytase expression was increased to 327 U/ml. Induction using lactose during fed-batch cultivation showed a lag phase of 4 h prior to an increase in the phytase activity to 71 U/ml during the same period as IPTG-induced production. Up to 90% of the total amount of expressed phytase leaked out from the E. coli cells in both IPTG- and lactose-induced fed-batch cultivations.

  18. High-Level Expression of Bacillus naganoensis Pullulanase from Recombinant Escherichia coli with Auto-Induction: Effect of lac Operator

    PubMed Central

    Xu, Yan; Chen, Wen Bo; Mu, Xiao Qing; Wang, Xinye; Xiao, Rong

    2013-01-01

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

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

    PubMed

    Nie, Yao; Yan, Wei; Xu, Yan; Chen, Wen Bo; Mu, Xiao Qing; Wang, Xinye; Xiao, Rong

    2013-01-01

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

  20. Cloning of Bacillus subtilis leucina A, B and C genes with Escherichia coli plasmids and expression of the leuC gene in E. coli.

    PubMed

    Nagahari, K; Sakaguchi, K

    1978-01-17

    The leucine genes of Bacillus subtilis have been cloned directly from the chromosomal DNA into Escherichia coli leuB cells by selection for the Leu+ phenotype using RSF2124 as a vector plasmid. The hybrid plasmid designated RSF2124-B.leu contained a 4.2 megadalton fragment derived from B. subtilis DNA, including the leu genes. The fragment had one site susceptible to EcoRI* and another site susceptible to BamNI endonuclease. Among the three fragments produced by EcoRI* and BamNI endonucleases, the 1.2 megadalton fragment had the ability to transform B. subtilis leuA, leuB and leuC auxotrophs to leu+. However, B. subtilis ilvB and ilvc auxotrophs were not rescued even by the whole 4.2 megadalton fragment present in the hybrid plasmid. beta-Isopropylmalate dehydrogenase (leuB gene product) activity found in E. coli cells containing the hybrid plasmid was about 60% of that in E. coli wild type cells, despite the high copy number (7.8) of the plasmid per chromosome observed.

  1. Antibacterial action of dispersed single-walled carbon nanotubes on Escherichia coli and Bacillus subtilis investigated by atomic force microscopy.

    PubMed

    Liu, Shaobin; Ng, Andrew Keong; Xu, Rong; Wei, Jun; Tan, Cher Ming; Yang, Yanhui; Chen, Yuan

    2010-12-01

    Single-walled carbon nanotubes (SWCNTs) exhibit strong antibacterial activities. Direct contact between bacterial cells and SWCNTs may likely induce cell damages. Therefore, the understanding of SWCNT-bacteria interactions is essential in order to develop novel SWCNT-based materials for their potential environmental, imaging, therapeutic, and military applications. In this preliminary study, we utilized atomic force microscopy (AFM) to monitor dynamic changes in cell morphology and mechanical properties of two typical bacterial models (gram-negative Escherichia coli and gram-positive Bacillus subtilis) upon incubation with SWCNTs. The results demonstrated that individually dispersed SWCNTs in solution develop nanotube networks on the cell surface, and then destroy the bacterial envelopes with leakage of the intracellular contents. The cell morphology changes observed on air dried samples are accompanied by an increase in cell surface roughness and a decrease in surface spring constant. To mimic the collision between SWCNTs and cells, a sharp AFM tip of 2 nm was chosen to introduce piercings on the cell surface. No clear physical damages were observed if the applied force was below 10 nN. Further analysis also indicates that a single collision between one nanotube and a bacterial cell is unlikely to introduce direct physical damage. Hence, the antibacterial activity of SWCNTs is the accumulation effect of large amount of nanotubes through interactions between SWCNT networks and bacterial cells.

  2. The preventive effect of Bacillus subtilus strain DB9011 against experimental infection with enterotoxcemic Escherichia coli in weaning piglets.

    PubMed

    Tsukahara, Takamitsu; Tsuruta, Takeshi; Nakanishi, Nobuo; Hikita, Chie; Mochizuki, Masami; Nakayama, Keizo

    2013-04-01

    Porcine edema disease (ED) is caused by Shiga toxin 2e-producing Escherichia coli (STEC). Post-weaned piglets often suffer from ED as a result of intestinal infection with STEC, which causes impaired growth performance and high mortality. Antimicrobial therapy is a curative treatment for piglets infected with STEC, but the emergence of antimicrobial-resistant STEC has become a serious problem for Japanese pig farmers. Therefore, an alternative strategy other than antimicrobial therapy is needed for the prevention or treatment of ED. In this study, we evaluated the effect of oral administration of Bacillus subtilis DB9011 (DB9011) to prevent the experimental infection of STEC in weaning piglets. Eight 21-day-old piglets were divided into two groups: STEC challenge with the basal diet, and STEC challenge with DB9011 supplemented diet. The challenge was carried out when the animals were 25, 26 and 27 days old using STEC contained in capsules resistant against gastric digestion. All pigs were euthanized at 36 days of age. DB9011 improved the symptoms of ED and decreased the number of STEC in the ileal digesta and feces. Accordingly, oral administration of DB9011 in weaned piglets prevents ED through the suppression of the growth of STEC in the ileum.

  3. Experimental Research on the Sterilization of Escherichia Coli and Bacillus Subtilis in Drinking Water by Dielectric Barrier Discharge

    NASA Astrophysics Data System (ADS)

    Li, Yang; Yi, Chengwu; Li, Jingjing; Yi, Rongjie; Wang, Huijuan

    2016-02-01

    The bactericidal effect on the representative type of Gram-negative Escherichia coli (E. coli) and Gram-positive Bacillus subtilis in drinking water was investigated in this paper by using dielectric barrier discharge (DBD) advanced oxidation technology. The sterilizing rates under different conditions of reaction time t, input voltage V, pH value, and initial concentration of bacteria C0 were investigated to figure out the optimum sterilization conditions. Our observations and comparisons of cell morphology alteration by scanning electron microscopy and transmission electron microscopy revealed the sterilization mechanisms. The results showed that the sterilizing rate increased obviously with the extension of reaction time t and the rise of input voltage V. The optimal sterilization effect was achieved when the pH value was 7.1. As the initial concentration of bacteria rose, the sterilizing rate decreased. When the input voltage was 2.2 kV and the initial concentration of bacteria was relatively low, the sterilizing rate almost reached 100% after a certain treatment time in neutral aqueous solution. The reasons for the great damage of cell structure and the killing of bacteria are the oxidation of O3, OH and the accumulation of active species produced by DBD. The article provides a certain theoretical and experimental basis for DBD application in water pollution treatment. supported by the Science and Technology Support Project Plan and Social Development of Jiangsu Province, China (No. BE2011732), the Science and Technology Support Project Plan and Social Development of Zhenjiang, Jiangsu Province, China (No. SH2012013)

  4. Biocontrol of the internalization of Salmonella enterica and Enterohaemorrhagic Escherichia coli in mung bean sprouts with an endophytic Bacillus subtilis.

    PubMed

    Shen, Zhenyu; Mustapha, Azlin; Lin, Mengshi; Zheng, Guolu

    2017-06-05

    Internalization of Salmonella enterica and enterohaemorrhagic Escherichia coli (EHEC) in seed sprouts poses a health risk to consumers, and the conventional sanitization methods are not always effective to reduce this risk. This study initiated a biocontrol approach to limit the internalization using endophytic Bacillus subtilis strains, which were isolated from the inner tissue of mung bean seeds or lettuce stems. By using the deferred agar method, 12 strains of B. subtilis out of 94 putative Bacillus isolates displayed inhibitory activity against at least one of the pathogenic indicators, S. enterica Typhimurium ATCC 14028 and E. coli O157:H7 505B. Two B. subtilis isolates (LCA1 and M24) showed a broad inhibitory spectrum against multiple strains of S. enterica and EHEC, Staphylococcus aureus sp., Klebsiella pneumoniae ATCC 700603, and Listeria monocytogenes Scott A, while the laboratory B. subtilis strain 168 was only moderately inhibitory against L. monocytogenes. To facilitate the tracking of the three B. subtilis strains (LCA1, M24, and 168) in the mung bean sprouts, the three strains were genetically engineered to carry the chloramphenicol acetyltransferase (cat), generating the strains LCA1-cat, M24-cat, and 168-cat, respectively. Data of the study using the cat-tagged strains demonstrated that both the two vegetable-associated and the laboratory B. subtilis strains could internalize in mung bean sprouts during the sprouting, but the latter displayed about 1.2 lg CFU/g of seeds lower in internalization. Overall, the presence of the three B. subtilis strains could significantly reduce the internalization of S. enterica or EHEC cocktail in mung bean sprouts during the sprouting. Among them, LCA1 showed the greatest inhibition against the EHEC cocktails with a reduction of about 2.0lg CFU/g of seeds by the end of sprouting (day 5), while 168 had the smallest reduction at about 0.6lg CFU/g of seeds. In addition, the three strains demonstrated a similar

  5. Enhanced extracellular production of recombinant proteins in Escherichia coli by co-expression with Bacillus cereus phospholipase C.

    PubMed

    Su, Lingqia; Jiang, Qi; Yu, Lingang; Wu, Jing

    2017-02-08

    Our laboratory has reported a strategy for improving the extracellular production of recombinant proteins through co-expression with Thermobifida fusca cutinase, which increases membrane permeability via its phospholipid hydrolysis activity. However, the foam generated by the lysophospholipid product makes the fermentation process difficult to control in a fermentor. Phospholipase C (PLC) catalyzes the hydrolysis of phospholipids to produce sn1,2-diacylglycerides and organic phosphate, which do not induce foam formation. Therefore, co-expression with Bacillus cereus PLC was investigated as a method to improve the extracellular production of recombinant proteins. When B. cereus PLC was expressed in Escherichia coli without its signal peptide, 95.3% of the total PLC activity was detected in the culture supernatant. PLC expression enhanced membrane permeability without obvious cell lysis. Then, six test enzymes, three secretory and three cytosolic, were co-expressed with B. cereus PLC. The enhancement of extracellular production correlated strongly with the molecular mass of the test enzyme. Extracellular production of Streptomyces sp. FA1 xylanase (43 kDa), which had the lowest molecular mass among the secretory enzymes, was 4.0-fold that of its individual expression control. Extracellular production of glutamate decarboxylase (51 kDa), which had the lowest molecular mass among the cytosolic enzymes, reached 26.7 U/mL; 88.3% of the total activity produced. This strategy was effectively scaled up using a 3-L fermentor. No obvious foam was generated during this fermentation process. This is the first study to detail the enhanced extracellular production of recombinant proteins through co-expression with PLC. This new strategy, which is especially appropriate for lower molecular mass proteins, allows large-scale protein production in an easily controlled fermentation process.

  6. Secretory expression of nattokinase from Bacillus subtilis YF38 in Escherichia coli.

    PubMed

    Liang, Xiaobo; Jia, Shifang; Sun, Yufang; Chen, Meiling; Chen, Xiuzhu; Zhong, Jin; Huan, Liandong

    2007-11-01

    Nattokinase producing bacterium, B. subtilis YF38, was isolated from douchi, using the fibrin plate method. The gene encoding this enzyme was cloned by polymerase chain reaction (PCR). Cytoplasmic expression of this enzyme in E. coli resulted in inactive inclusion bodies. But with the help of two different signal peptides, the native signal peptide of nattokinase and the signal peptide of PelB, active nattokinase was successfully expressed in E. coli with periplasmic secretion, and the nattokinase in culture medium displayed high fibrinolytic activity. The fibrinolytic activity of the expressed enzyme in the culture was determined to reach 260 urokinase units per micro-liter when the recombinant strain was induced by 0.7 mmol l(-1) isopropyl-beta-D- thiogalactopyranoside (IPTG) at 20 degrees C for 20 h, resulting 49.3 mg active enzyme per liter culture. The characteristic of this recombinant nattokinase is comparable to the native nattokinase from B. subtilis YF38. Secretory expression of nattokinase in E. coli would facilitate the development of this enzyme into a therapeutic product for the control and prevention of thrombosis diseases.

  7. Characterization of the glutamate-specific endopeptidase from Bacillus licheniformis expressed in Escherichia coli.

    PubMed

    Ye, Wei; Wang, Haiying; Ma, Yi; Luo, Xiaochun; Zhang, Weimin; Wang, Jufang; Wang, Xiaoning

    2013-10-10

    Glutamate-specific endopeptidase from Bacillus licheniformis (GSE-BL) is widely used in peptide recovery and synthesis because of its unique substrate specificity. However, the mechanism underlying its specificity is still not thoroughly understood. In this study, the roles of the prosegment and key amino acids involved in the proteolytic activity of GSE-BL were investigated. Loss of the GSE-BL prosegment severely restricted enzymatic activity toward Z-Phe-Leu-Glu-pNA. A homologous model of GSE-BL revealed that it contains the catalytic triad "His47, Asp96 and Ser 167", which was further confirmed by site-directed mutagenesis. In vitro mutagenesis further indicated that Val2, Arg89 and His190 are essential for enzymatic activity toward Z-Phe-Leu-Glu-pNA. Moreover, the catalytic efficiency of Phe57Ala GSE-BL toward Z-Phe-Leu-Glu-pNA was 50% higher than that of the native mature GSE-BL. This is the first study to fully elucidate the key amino acids for proteolytic activity of GSE-BL. Mature GSE-BL could be obtained through self-cleavage alone when Lys at -1 position was replaced by Glu, providing a new strategy for the preparation of mature GSE-BL. This study yielded some valuable insights into the substrate specificity of glutamate-specific endopeptidase, establishing a foundation for broadening the applications of GSE-BL.

  8. Diarrheagenic Escherichia coli

    PubMed Central

    Nataro, James P.; Kaper, James B.

    1998-01-01

    Escherichia coli is the predominant nonpathogenic facultative flora of the human intestine. Some E. coli strains, however, have developed the ability to cause disease of the gastrointestinal, urinary, or central nervous system in even the most robust human hosts. Diarrheagenic strains of E. coli can be divided into at least six different categories with corresponding distinct pathogenic schemes. Taken together, these organisms probably represent the most common cause of pediatric diarrhea worldwide. Several distinct clinical syndromes accompany infection with diarrheagenic E. coli categories, including traveler’s diarrhea (enterotoxigenic E. coli), hemorrhagic colitis and hemolytic-uremic syndrome (enterohemorrhagic E. coli), persistent diarrhea (enteroaggregative E. coli), and watery diarrhea of infants (enteropathogenic E. coli). This review discusses the current level of understanding of the pathogenesis of the diarrheagenic E. coli strains and describes how their pathogenic schemes underlie the clinical manifestations, diagnostic approach, and epidemiologic investigation of these important pathogens. PMID:9457432

  9. Pathogenic Escherichia coli

    USDA-ARS?s Scientific Manuscript database

    Escherichia coli, a member of the Enterobacteriaceae family, is a part of the normal flora of the intestinal tract of humans and a variety of animals. E. coli strains are classified on the basis of antigenic differences in two surface components (serotyping), the somatic antigen (O) of the lipopoly...

  10. PATHOGENIC ESCHERICHIA COLI

    EPA Science Inventory

    Escherichia coli is a bacterial species which inhabits the gastrointestinal tract of man and warm-blooded animals. Because of the ubiquity of this bacterium in the intestinal flora, it serves as an important indicator organism of fecal contamination. E. coli, aside from serving a...

  11. Bacillus subtilis homologs of MviN (MurJ), the putative Escherichia coli lipid II flippase, are not essential for growth.

    PubMed

    Fay, Allison; Dworkin, Jonathan

    2009-10-01

    Although peptidoglycan synthesis is one of the best-studied metabolic pathways in bacteria, the mechanism underlying the membrane translocation of lipid II, the undecaprenyl-disaccharide pentapeptide peptidoglycan precursor, remains mysterious. Recently, it was proposed that the essential Escherichia coli mviN gene encodes the lipid II flippase. Bacillus subtilis contains four proteins that are putatively homologous to MviN, including SpoVB, previously reported to be necessary for spore cortex peptidoglycan synthesis during sporulation. MviN complemented the sporulation defect of a DeltaspoVB mutation, and SpoVB and another of the B. subtilis homologs, YtgP, complemented the growth defect of an E. coli strain depleted for MviN. Thus, these B. subtilis proteins are likely to be MviN homologs. However, B. subtilis strains lacking these four proteins have no defects in growth, indicating that they likely do not serve as lipid II flippases in this organism.

  12. Probing the binding site of curcumin in Escherichia coli and Bacillus subtilis FtsZ--a structural insight to unveil antibacterial activity of curcumin.

    PubMed

    Kaur, Simranjeet; Modi, Niraj H; Panda, Dulal; Roy, Nilanjan

    2010-09-01

    The cytoskeletal protein, FtsZ plays a pivotal role in prokaryotic cell division and is present in majority of the bacterial species. In recent years, inhibitors of FtsZ have been identified that may function as lead compounds for the development of novel antimicrobials. It has been found that curcumin, the main bioactive component of Curcuma longa, inhibits Bacillus subtilis and Escherichia coli growth by inhibiting FtsZ assembly. Though it is experimentally established that curcumin inhibits FtsZ polymerization, the binding site of curcumin in FtsZ is not known. In this study, interaction of curcumin with catalytic core domain of E. coli and B. subtilis FtsZ was investigated using computational docking. 2010 Elsevier Masson SAS. All rights reserved.

  13. Escherichia coli biofilms

    PubMed Central

    Beloin, Christophe; Roux, Agnès; Ghigo, Jean-Marc

    2008-01-01

    Escherichia coli is a predominant species among facultative anaerobic bacteria of the gastrointestinal tract. Both its frequent community lifestyle and the availability of a wide array of genetic tools contributed to establish E. coli as a relevant model organism for the study of surface colonization. Several key factors, including different extracellular appendages, are implicated in E. coli surface colonization and their expression and activity are finely regulated, both in space and time, to ensure productive events leading to mature biofilm formation. This chapter will present known molecular mechanisms underlying biofilm development in both commensal and pathogenic E. coli. PMID:18453280

  14. The aminoglycoside antibiotic kanamycin damages DNA bases in Escherichia coli: caffeine potentiates the DNA-damaging effects of kanamycin while suppressing cell killing by ciprofloxacin in Escherichia coli and Bacillus anthracis.

    PubMed

    Kang, Tina Manzhu; Yuan, Jessica; Nguyen, Angelyn; Becket, Elinne; Yang, Hanjing; Miller, Jeffrey H

    2012-06-01

    The distribution of mutants in the Keio collection of Escherichia coli gene knockout mutants that display increased sensitivity to the aminoglycosides kanamycin and neomycin indicates that damaged bases resulting from antibiotic action can lead to cell death. Strains lacking one of a number of glycosylases (e.g., AlkA, YzaB, Ogt, KsgA) or other specific repair proteins (AlkB, PhrB, SmbC) are more sensitive to these antibiotics. Mutants lacking AlkB display the strongest sensitivity among the glycosylase- or direct lesion removal-deficient strains. This perhaps suggests the involvement of ethenoadenine adducts, resulting from reactive oxygen species and lipid peroxidation, since AlkB removes this lesion. Other sensitivities displayed by mutants lacking UvrA, polymerase V (Pol V), or components of double-strand break repair indicate that kanamycin results in damaged base pairs that need to be removed or replicated past in order to avoid double-strand breaks that saturate the cellular repair capacity. Caffeine enhances the sensitivities of these repair-deficient strains to kanamycin and neomycin. The gene knockout mutants that display increased sensitivity to caffeine (dnaQ, holC, holD, and priA knockout mutants) indicate that caffeine blocks DNA replication, ultimately leading to double-strand breaks that require recombinational repair by functions encoded by recA, recB, and recC, among others. Additionally, caffeine partially protects cells of both Escherichia coli and Bacillus anthracis from killing by the widely used fluoroquinolone antibiotic ciprofloxacin.

  15. Diarrheagenic Escherichia coli.

    PubMed

    Gomes, Tânia A T; Elias, Waldir P; Scaletsky, Isabel C A; Guth, Beatriz E C; Rodrigues, Juliana F; Piazza, Roxane M F; Ferreira, Luís C S; Martinez, Marina B

    2016-12-01

    Most Escherichia coli strains live harmlessly in the intestines and rarely cause disease in healthy individuals. Nonetheless, a number of pathogenic strains can cause diarrhea or extraintestinal diseases both in healthy and immunocompromised individuals. Diarrheal illnesses are a severe public health problem and a major cause of morbidity and mortality in infants and young children, especially in developing countries. E. coli strains that cause diarrhea have evolved by acquiring, through horizontal gene transfer, a particular set of characteristics that have successfully persisted in the host. According to the group of virulence determinants acquired, specific combinations were formed determining the currently known E. coli pathotypes, which are collectively known as diarrheagenic E. coli. In this review, we have gathered information on current definitions, serotypes, lineages, virulence mechanisms, epidemiology, and diagnosis of the major diarrheagenic E. coli pathotypes. Copyright © 2016 Sociedade Brasileira de Microbiologia. Published by Elsevier Editora Ltda. All rights reserved.

  16. The dacA gene of Bacillus stearothermophilus coding for D-alanine carboxypeptidase: cloning, structure and expression in Escherichia coli and Pichia pastoris.

    PubMed

    Despreaux, C W; Manning, R F

    1993-09-06

    The bacterial D-alanine carboxypeptidases (CPases) remove C-terminal D-alanyl residues from sugar-peptide cell wall precursors. The CPases have many characteristics in common with the high-M(r) penicillin-binding proteins (PBPs) whose inhibition by beta-lactam antibiotics is lethal. The CPases are attractive as model PBPs, because of their relatively lower M(r) and higher activity in vitro. We have cloned and sequenced the Bacillus stearothermophilus gene (dacA) coding for a membrane-bound CPase. The nucleotide (nt) sequence of the gene is homologous to that of the Escherichia coli and Bacillus subtilis dacA loci, which also code for membrane-bound CPases. E. coli host cells lysed when expression of B. stearothermophilus dacA was induced. The same coding sequence was expressed in the methylotrophic yeast, Pichia pastoris, using the alcohol oxidase-1 (AOX1) promoter. Over 100 micrograms/ml of CPase was efficiently secreted into the medium after induction by methanol, without adversely affecting this host. The yeast product is indistinguishable from the native enzyme in structure and activity. The ability to secrete large amounts of heterologous protein and the lack of endogenous peptidoglycan metabolism makes P. pastoris an attractive candidate for the production of PBPs.

  17. The 8-amino-7-oxopelargonate synthase from Bacillus sphaericus. Purification and preliminary characterization of the cloned enzyme overproduced in Escherichia coli.

    PubMed Central

    Ploux, O; Marquet, A

    1992-01-01

    The 8-amino-7-oxopelargonate synthase [6-carboxyhexanoyl-CoA:L-alanine carboxyhexanoyltransferase (decarboxylating); EC 2.3.1.47] from Bacillus sphaericus involved in biotin biosynthesis was purified from an Escherichia coli overproducing strain. The purification afforded an electrophoretically homogeneous enzyme with a specific activity of 0.67 unit/mg. The purified enzyme is a monomer of 41 kDa. N-Terminal sequencing of the first 14 amino acid residues showed complete agreement with the predicted sequence from the bioF gene. The pure enzyme showed the characteristic absorption band (425 nm) of pyridoxal 5'-phosphate-dependent enzymes. Furthermore, the holoenzyme was resolved during an affinity step yielding the inactive apoenzyme, which recovered activity and the 425 nm-absorption band on dialysis against pyridoxal 5'-phosphate. Km values for L-alanine and pimeloyl-CoA were respectively 3 mM and 1 microM. Images Fig. 1. PMID:1575677

  18. Isolation and characterization of Bacillus subtilis genes involved in siderophore biosynthesis: relationship between B. subtilis sfpo and Escherichia coli entD genes.

    PubMed Central

    Grossman, T H; Tuckman, M; Ellestad, S; Osburne, M S

    1993-01-01

    In response to iron deprivation, Bacillus subtilis secretes a catecholic siderophore, 2,3-dihydroxybenzoyl glycine, which is similar to the precursor of the Escherichia coli siderophore enterobactin. We isolated two sets of B. subtilis DNA sequences that complemented the mutations of several E. coli siderophore-deficient (ent) mutants with defective enterobactin biosynthesis enzymes. One set contained DNA sequences that complemented only an entD mutation. The second set contained DNA sequences that complemented various combinations of entB, entE, entC, and entA mutations. The two sets of DNA sequences did not appear to overlap. AB. subtilis mutant containing an insertion in the region of the entD homolog grew much more poorly in low-iron medium and with markedly different kinetics. These data indicate that (i) at least five of the siderophore biosynthesis genes of B. subtilis can function in E. coli, (ii) the genetic organization of these siderophore genes in B. subtilis is similar to that in E. coli, and (iii) the B. subtilis entD homolog is required for efficient growth in low-iron medium. The nucleotide sequence of the B. subtilis DNA contained in plasmid pENTA22, a clone expressing the B. subtilis entD homolog, revealed the presence of at least two genes. One gene was identified as sfpo, a previously reported gene involved in the production of surfactin in B. subtilis and which is highly homologous to the E. coli entD gene. We present evidence that the E. coli entD and B. subtilis sfpo genes are interchangeable and that their products are members of a new family of proteins which function in the secretion of peptide molecules. Images PMID:8407792

  19. Expression of the bifunctional Bacillus subtilis TatAd protein in Escherichia coli reveals distinct TatA/B-family and TatB-specific domains.

    PubMed

    Barnett, James P; Lawrence, Janna; Mendel, Sharon; Robinson, Colin

    2011-08-01

    In the Tat protein export pathway of Gram-negative bacteria, TatA and TatB are homologous proteins that carry out distinct and essential functions in separate sub-complexes. In contrast, Gram-positive Tat systems usually lack TatB and the TatA protein is bifunctional. We have used a mutagenesis approach to delineate TatA/B-type domains in the bifunctional TatAd protein from Bacillus subtilis. This involved expression of mutated TatAd variants in Escherichia coli and tests to determine whether the variants could function as TatA or TatB by complementing E. coli tatA and/or tatB mutants. We show that mutations in the C-terminal half of the transmembrane span and the subsequent FGP 'hinge' motif are critical for TatAd function with its partner TatCd subunit, and the same determinants are required for complementation of either tatA or tatB mutants in Escherichia coli. This is thus a critical domain in both TatA and TatB proteins. In contrast, substitution of a series of residues at the N-terminus specifically blocks the ability of TatAd to substitute for E. coli TatB. The results point to the presence of a universally conserved domain in the TatA/B-family, together with a separate N-terminal domain that is linked to the TatB-type function in Gram-negative bacteria.

  20. Delta1-pyrroline-5-carboxylic acid formed by proline dehydrogenase from the Bacillus subtilis ssp. natto expressed in Escherichia coli as a precursor for 2-acetyl-1-pyrroline.

    PubMed

    Huang, Tzou-Chi; Huang, Yi-Wen; Hung, Hui-Ju; Ho, Chi-Tang; Wu, Mei-Li

    2007-06-27

    Proline dehydrogenase (PRODH) catalyzes the biosynthesis of Delta1-pyrroline-5-carboxylic acid (P5C). The Bacillus subtilis subsp. natto gene for the proline dehydrogenase (BnPRODH) was cloned and expressed in Escherichia coli. Nucleotide sequence analysis of the clone revealed an open-reading frame that encodes 302 amino acid polypeptide with a calculated molecular mass of 34.5 kDa. The deduced amino acid sequence showed sequence similarity to bacterial PRODH and PutA of E. coli. The BnPRODH gene was cloned into pET21b and was expressed at a high level in E. coli BL21(DE3). The expressed protein was purified by using nickel ion affinity column chromatography to homogeneity before characterization. The purified recombinant BnPRODH was used to produce P5C. Model system composed of P5C and methylglyoxal was set up to study the formation of 2-acetyl-1-pyrroline. Our data showed that P5C, derived from the conversion of l-proline by the purified recombinant PRODH, might react directly with methylglyoxal to form 2-AP. P5C/methylglyoxal pathway represents the first report of a biological mechanism by which 2-AP may be synthesized in vitro by PRODH.

  1. Recurrent Escherichia coli bacteremia.

    PubMed Central

    Maslow, J N; Mulligan, M E; Arbeit, R D

    1994-01-01

    Escherichia coli is the most common gram-negative organism associated with bacteremia. While recurrent E. coli urinary tract infections are well-described, recurrent E. coli bacteremia appears to be uncommon, with no episodes noted in multiple series of patients with gram-negative bacteremias. We report on 5 patients with recurrent bloodstream infections identified from a series of 163 patients with E. coli bacteremia. For each patient, the isolates from each episode were analyzed by pulsed-field gel electrophoresis (PFGE) and ribotyping and for the presence of E. coli virulence factors. For each of four patients, the index and recurrent episodes of bacteremia represented the same strain as defined by PFGE, and the strains were found to carry one or more virulence factors. The remaining patient, with two episodes of bloodstream infection separated by a 4-year interval, was infected with two isolates that did not carry any virulence factors and that were clonally related by ribotype analysis but differed by PFGE. All five patients had either a local host defense defect (three patients) or impaired systemic defenses (one patient) or both (one patient). Thus, recurrent E. coli bacteremia is likely to represent a multifactorial process that occurs in patients with impaired host defenses who are infected with virulent isolates. Images PMID:7910828

  2. Purification and characterization of an extracellular alpha-L-arabinosidase from a novel isolate Bacillus pumilus ARA and its over-expression in Escherichia coli.

    PubMed

    Pei, Jianjun; Shao, Weilan

    2008-02-01

    The alpha-L-arabinosidase, AraB, was induced when Bacillus pumilus ARA was grown at 50 degrees C in a minimal medium containing xylan. A 56-kDa protein with alpha-L-arabinosidase activity was purified from culture supernatant to gel electrophoretic homogeneity. The optimal activity was at pH 6.4 and 60 degrees C over a 10-min assay. The purified enzyme was stable over a pH range of 5.2-7.6 and had a 1-h half life at 70 degrees C. The enzyme released arabinose from oat spelt xylan. Kinetic experiments at 60 degrees C with p-nitrophenyl alpha-L-arabinofuranoside as substrate gave a K (m), and V (max) of 1.05 mM and 240 U per mg of protein. The NH(2)-terminal amino acid sequence of the enzyme was determined, and its gene araB was subsequently cloned, sequenced, and over-expressed in Escherichia coli. The open reading frame of araB consists of a 1,479-bp fragment encoding a protein of 472 amino acids, which belonged to family 51 of the glycoside hydrolases with an identity of 67% to the protein encoded by abfB of Bacillus subtilis 168.

  3. Proteome-wide identification of lysine propionylation in thermophilic and mesophilic bacteria: Geobacillus kaustophilus, Thermus thermophilus, Escherichia coli, Bacillus subtilis, and Rhodothermus marinus.

    PubMed

    Okanishi, Hiroki; Kim, Kwang; Masui, Ryoji; Kuramitsu, Seiki

    2017-03-01

    Recent studies have revealed the physiological significance of post-translational lysine acylations such as acetylation in the regulation of various cellular processes. Here, we characterized lysine propionylation, a recently discovered post-translational acylation, in five representative bacteria: Geobacillus kaustophilus, Thermus thermophilus, Escherichia coli, Bacillus subtilis, and Rhodothermus marinus. Using antibody-based propionyl peptide enrichment followed by identification with nano-liquid chromatography tandem mass spectrometry, we showed that proteins were subject to lysine propionylation in all five bacterial species analyzed. Notably, many propionylations were identified in the Bacillus-related, thermophilic eubacterium G. kaustophilus, but fewer in the mesophilic eubacterium B. subtilis, suggesting that propionylation event abundance is independent of phylogenetic relationship. We further found propionylation sites in the thermophilic eubacterium T. thermophilus, but the thermophilic eubacterium R. marinus showed the fewest number of sites, indicating that growth temperature is not a determinant of propionylation state. In silico analyses demonstrated that lysine propionylation is related to metabolic pathways, particularly those controlled by acyl-CoA synthetases, similar to lysine acetylation. We also detected dozens of propionylation sites at positions important for protein functions across bacteria, demonstrating the regulatory mechanisms affected by lysine propionylations. Our proteome-wide analyses across bacteria thus provide insights into the general functions of lysine propionylation.

  4. Effect of a 20-kilodalton protein from Bacillus thuringiensis subsp. israelensis on production of the CytA protein by Escherichia coli.

    PubMed Central

    Visick, J E; Whiteley, H R

    1991-01-01

    CytA, a 27-kDa cytolytic crystal protein of Bacillus thuringiensis subsp. israelensis, is produced only at very low levels by recombinant Escherichia coli cells unless a 20-kDa B. thuringiensis subsp. israelensis protein is also present (K. M. McLean and H. R. Whiteley, J. Bacteriol. 169:1017-1023, 1987; L. F. Adams, J. E. Visick, and H. R. Whiteley, J. Bacteriol. 171:521-530, 1989). However, the data reported here demonstrate that the 20-kDa protein is not required for high-level CytA production in E. coli strains carrying mutations in rpoH, groEL, or dnaK, all of which affect the proteolytic ability of the cells. The 20-kDa protein also increases the amount of CryIVD (another B. thuringiensis subsp. israelensis crystal protein) and LacZX90 (a mutant of beta-galactosidase) made by E. coli. The latter phenomenon is attributable to an increase in the half-life of LacZX90, suggesting that the 20-kDa protein may stabilize this protein. The effect of the 20-kDa protein was also examined in vitro and in a T7 RNA polymerase expression system, and the possible significance of these results for the timing of proteolysis and of 20-kDa protein activity is discussed. Finally, the ability of a single antibody to coimmunoprecipitate CytA and the 20-kDa protein from E. coli extracts provides evidence for a protein-protein interaction that may be related to the mechanism of action of the 20-kDa protein. Images PMID:1900280

  5. Cloning, expression and purification of binding domains of lethal factor and protective antigen of Bacillus anthracis in Escherichia coli and evaluation of their related murine antibody.

    PubMed

    Rezaee, Mehdi; Honari, Hossein; Kooshk, Mohammad Reza Ashrafi

    2014-01-01

    Anthrax is common disease between human and animals caused by Bacillus anthracis. The cell binding domain of protective antigen (PAD4) and the binding domain of lethal factor (LFD1) have high immunogenicity potential and always were considered as a vaccine candidate against anthrax. The aims of this study are cloning and expressing of PAD4 and LFD1 in Escherichia coli, purification of the recombinant proteins and determination of their immunogenicity through evaluating of the relative produced polyclonal antibodies in mice. PAD4 and LFD1 genes were cloned in pET28a(+) vector and expressed in E. coli Bl21(DE3)PlysS. Expression and purification of the two recombinant proteins were confirmed by SDS-PAGE and Western blotting techniques. The PAD4 and LFD1 were purified using Ni(+)-NTA affinity chromatography (95-98 %), yielding 37.5 and 45 mg/l of culture, respectively. The antigens were injected three times into mice and production of relative antibodies was evaluated by ELISA test. The results showed that both PAD4 and LFD1 are immunogenic, but LFD1 has higher potential to stimulate Murine immune system. With regard to the high level of LFD1 and PAD4 expression and also significant increment in produced polyclonal antibodies, these recombinant proteins can be considered as a recombinant vaccine candidate against anthrax.

  6. Toward functional genomics in bacteria: Analysis of gene expression in Escherichia coli from a bacterial artificial chromosome library of Bacillus cereus

    PubMed Central

    Rondon, Michelle R.; Raffel, Sandra J.; Goodman, Robert M.; Handelsman, Jo

    1999-01-01

    As the study of microbes moves into the era of functional genomics, there is an increasing need for molecular tools for analysis of a wide diversity of microorganisms. Currently, biological study of many prokaryotes of agricultural, medical, and fundamental scientific interest is limited by the lack of adequate genetic tools. We report the application of the bacterial artificial chromosome (BAC) vector to prokaryotic biology as a powerful approach to address this need. We constructed a BAC library in Escherichia coli from genomic DNA of the Gram-positive bacterium Bacillus cereus. This library provides 5.75-fold coverage of the B. cereus genome, with an average insert size of 98 kb. To determine the extent of heterologous expression of B. cereus genes in the library, we screened it for expression of several B. cereus activities in the E. coli host. Clones expressing 6 of 10 activities tested were identified in the library, namely, ampicillin resistance, zwittermicin A resistance, esculin hydrolysis, hemolysis, orange pigment production, and lecithinase activity. We analyzed selected BAC clones genetically to identify rapidly specific B. cereus loci. These results suggest that BAC libraries will provide a powerful approach for studying gene expression from diverse prokaryotes. PMID:10339608

  7. Development of Pgrac100-based expression vectors allowing high protein production levels in Bacillus subtilis and relatively low basal expression in Escherichia coli.

    PubMed

    Phan, Trang Thi Phuong; Tran, Linh Thuoc; Schumann, Wolfgang; Nguyen, Hoang Duc

    2015-05-21

    In general, fusion of recombinant genes to strong inducible promoters allowing intracellular expression in Bacillus subtilis is a two-step process. The ligation products are transformed into Escherichia coli, followed by identification of the correct plasmid, and this plasmid is subsequently transformed into B. subtilis. This raises the problem that basal level of expression of the recombinant gene could be harmful for E. coli cells. Based on the Pgrac promoter, we optimized the UP element, the -35, 15, -10 and the +1 region to enhance the promoter activity in B. subtilis after induction. However, detailed investigations for a promoter to develop expression vectors that allows high protein production levels in B. subtilis and a relatively low basal expression levels in E. coli has not been studied yet. We screened the previously constructed library of E. coli - B. subtilis shuttle vectors for high level expression in B. subtilis and low basal level in E. coli. Promoter Pgrac100 turned out to meet these criteria, in which ß-galactosidase expression level of Pgrac100-bgaB is about 9.2 times higher than Pgrac01-bgaB in B. subtilis and the ratio of those in induced B. subtilis over un-induced E. coli from Pgrac100-bgaB is 1.3 times higher than Pgrac01-bgaB. Similarly, GFP expression level of Pgrac100-gfp is about 27 times higher than that of Pgrac01-gfp and the ratio from Pgrac100-gfp is 35.5 times higher than Pgrac01-gfp. This promoter was used as a basis for the construction of three novel vectors, pHT253 (His-tag-MCS), pHT254 (MCS-His-tag) and pHT255 (MCS-Strep-tag). Expression of the reporter proteins BgaB and GFP using these expression vectors in B. subtilis at a low IPTG concentration were measured and the fusion proteins could be purified easily in a single step by using Strep-Tactin or IMAC-Ni columns. This paper describes the construction and analysis of an IPTG-inducible expression vector termed Pgrac100 for the high level production of intracellular

  8. Enterotoxigenic Escherichia coli

    PubMed Central

    Fleckenstein, James M; Munson, George M; Rasko, David A

    2013-01-01

    The enterotoxigenic Escherichia coli are a pervasive cause of serious diarrheal illness in developing countries. Presently, there is no vaccine to prevent these infections, and many features of the basic pathogenesis of these organisms remain poorly understood. Until very recently most pathogenesis studies had focused almost exclusively on a small subset of known “classical” virulence genes, namely fimbrial colonization factors and the heat-labile (LT) and heat stable (ST) enterotoxins. However, recent investigations of pathogen-host interactions reveal a surprisingly complex and intricately orchestrated engagement involving the interplay of classical and “novel” virulence genes, as well as participation of genes highly conserved in the E. coli species. These studies may inform further rational approaches to vaccine development for these important pathogens. PMID:23892244

  9. Evidence of the importance of the Met115 for Bacillus thuringiensis subsp. israelensis Cyt1Aa protein cytolytic activity in Escherichia coli.

    PubMed

    Zghal, Raida Zribi; Trigui, Hana; Ben Ali, Mamdouh; Jaoua, Samir

    2008-02-01

    Cyt1Aa is a cytolytic toxin, found together with the delta-endotoxins in Bacillus thuringiensis subsp. israelensis parasporal insecticidal crystals. The latter are used as an environmental friendly insecticide against mosquitoes and black flies. Contrary to Cry delta-endotoxin, the mode of action of Cyt1Aa is not completely understood. In the absence of direct structural data, a novel mutated cyt1Aa gene was used to obtain indirect informations on Cyt1Aa conformation changes in the lipid membrane environment. A mutated cyt1Aa gene named cyt1A97 has been isolated from a B. thuringiensis israelensis strain named BUPM97. The nucleotide sequence predicted a protein of 249 amino acids residues with a calculated molecular mass of 27 kDa. Both nucleotide and amino acid sequences similarity analysis revealed that cyt1A97 presents one amino acid different from the native cyt1Aa gene. This mutation was located in the helix alpha C corresponding to a substitution of Met(115) by a Thr. The heterologous expression of the cyt1A97 and another cyt1Aa-type gene called cyt1A98, not affected by such mutation used as control, was performed in Escherichia coli. It revealed that the mutated Cyt1A97 protein was over produced as inclusion bodies showing a very weak toxicity to E. coli contrarily to Cyt1A98 that stopped E. coli growth. Hence, hydrophobic residue Met at position 115 of Cyt1Aa should play a very important role for the maintenance of the structure and cytolytic functions of Cyt1Aa.

  10. The Escherichia coli TatABC system and a Bacillus subtilis TatAC-type system recognise three distinct targeting determinants in twin-arginine signal peptides.

    PubMed

    Mendel, Sharon; McCarthy, Andrew; Barnett, James P; Eijlander, Robyn T; Nenninger, Anja; Kuipers, Oscar P; Robinson, Colin

    2008-01-18

    The Tat system transports folded proteins across bacterial and thylakoid membranes. In Gram-negative organisms, it is encoded by tatABC genes and the system recognizes substrates bearing signal peptides with a conserved twin-arginine motif. Most Gram-positive organisms lack a tatB gene, indicating major differences in organisation and/or mechanism. Here, we have characterized the essential targeting determinants that are recognized by a Bacillus subtilis TatAC-type system, TatAdCd. Substitution by lysine of either of the twin-arginine residues in the TorA signal peptide can be tolerated, but the presence of twin-lysine residues blocks export completely. We show that additional determinants can be as important as the twin-arginine motif. Replacement of the -1 serine by alanine, in either the TorA or DmsA signal peptide, almost blocks export by either the B. subtilis TatAdCd or Escherichia coli TatABC systems, firmly establishing the importance of this -1 residue in these signal peptides. Surprisingly, the +2 leucine in the DmsA signal peptide (sequence SRRGLV) appears to play an equally important role and substitution by alanine or phenylalanine blocks export by both the B. subtilis and E. coli systems. These data identify three distinct determinants, whose importance varies depending on the signal peptide in question. The data also show that the B. subtilis TatAdCd and E. coli TatABC systems recognize very similar determinants within their target peptides, and exhibit surprisingly similar responses to mutations within these determinants.

  11. Stereospecific synthesis of (R)-2-hydroxy carboxylic acids using recombinant E. coli BL21 overexpressing YiaE from Escherichia coli K12 and glucose dehydrogenase from Bacillus subtilis.

    PubMed

    Yun, Hyungdon; Choi, Hyeon-Lok; Fadnavis, Nitin W; Kim, Byung-Gee

    2005-01-01

    The yiaE gene from Escherichia coli K12 was functionally expressed in E. coli BL21 using an IPTG inducible pET expression system (2.1 U/mg), and YiaE was purified to a specific activity of 18 U/mg. The purified enzyme catalyzes reduction of various aromatic and aliphatic 2-oxo carboxylic acids to the corresponding (R)-2-hydoxy carboxylic acids using NADPH. For practical applications, the problem of NADPH recycle was effectively solved by using recombinant E. coli overexpressing YiaE and glucose dehydrogenase from Bacillus subtilis in the same cell. The recombinant E. coli was used to prepare (R)-phenyllactic acid and (R)-2-hydroxy-4-phenylbutanoic acid from the corresponding 2-oxo carboxylic acids (98% ee) while the alpha-carbonyl group of 2,4-dioxo-4-phenylbutyric acid was reduced regio- and stereospecifically to give (R)-2-hydroxy-4-oxo-4-phenylbutyric acid (97% ee) in quantitative yields. The cells could be recycled for 3 days at room temperature in 100 mM phosphate buffer (pH 7.0) without loss of activity, which reduced to 70% after 1 week.

  12. Tetracycline/H+ antiport and Na+/H+ antiport catalyzed by the Bacillus subtilis TetA(L) transporter expressed in Escherichia coli.

    PubMed

    Guffanti, A A; Krulwich, T A

    1995-08-01

    The properties of TetA(L)-dependent tetracycline/proton and Na+/proton antiport were studied in energized everted vesicles of Escherichia coli transformed with a cloned tetA(L) gene (pJTA1) from Bacillus subtilis. Inhibition patterns by valinomycin and nigericin indicated that both antiports were electrogenic, in contrast to the tetracycline/proton antiport encoded by gram-negative plasmid tet genes. Tetracycline uptake in the everted system was dependent upon a divalent cation, with cobalt being the preferred one. The apparent Km for tetracycline was markedly increased at pH 8.5 versus pH 7.5, whereas the Vmax was unchanged. The much higher apparent Km for Na+ decreased at pH 8.5 relative to that at pH 7.5, as did the Vmax. Na+ did not affect tetracycline uptake, nor did Co2+ and/or tetracycline affect Na+ uptake; complex patterns of inhibition by amiloride and analogs thereof were observed.

  13. Tetracycline/H+ antiport and Na+/H+ antiport catalyzed by the Bacillus subtilis TetA(L) transporter expressed in Escherichia coli.

    PubMed Central

    Guffanti, A A; Krulwich, T A

    1995-01-01

    The properties of TetA(L)-dependent tetracycline/proton and Na+/proton antiport were studied in energized everted vesicles of Escherichia coli transformed with a cloned tetA(L) gene (pJTA1) from Bacillus subtilis. Inhibition patterns by valinomycin and nigericin indicated that both antiports were electrogenic, in contrast to the tetracycline/proton antiport encoded by gram-negative plasmid tet genes. Tetracycline uptake in the everted system was dependent upon a divalent cation, with cobalt being the preferred one. The apparent Km for tetracycline was markedly increased at pH 8.5 versus pH 7.5, whereas the Vmax was unchanged. The much higher apparent Km for Na+ decreased at pH 8.5 relative to that at pH 7.5, as did the Vmax. Na+ did not affect tetracycline uptake, nor did Co2+ and/or tetracycline affect Na+ uptake; complex patterns of inhibition by amiloride and analogs thereof were observed. PMID:7635843

  14. Simultaneous Detection of Listeria monocytogenes, Escherichia coli O157:H7, Bacillus cereus, Salmonella spp., and Staphylococcus aureus in Low-fatted Milk by Multiplex PCR

    PubMed Central

    Kim, Ji-Hyun; Rhim, Seong-Ryul; Kim, Kee-Tae; Paik, Hyun-Dong

    2014-01-01

    A rapid and specific PCR assay for the simultaneous detection of Listeria monocytogenes, Escherichia coli O157:H7, Bacillus cereus, Salmonella spp., and Staphylococcus aureus in foods was developed to reduce the detection time and to increase sensitivity. Multiplex PCR developed in this study produced only actA, fliC, hbl, invA, ileS amplicons, but did not produce any non-specific amplicon. The primer sets successfully amplified the target genes in the multiplex PCR without any non-specific or additional bands on the other strains. The multiplex PCR assays also amplified some target genes from five pathogens, and multiplex amplification was obtained from as little as 1 pg of DNA. According to the results from the sensitivity evaluation, the multiplex PCR developed in this study detected 10 cells/mL of the pathogens inoculated in milk samples, respectively. The results suggested that multiplex PCR was an effective assay demonstrating high specificity for the simultaneous detection of five target pathogens in food system. PMID:26761507

  15. Purification and characterization of two alkaline, thermotolerant alpha-amylases from Bacillus halodurans 38C-2-1 and expression of the cloned gene in Escherichia coli.

    PubMed

    Murakami, Shuichiro; Nishimoto, Haruka; Toyama, Yosuke; Shimamoto, Etsuko; Takenaka, Shinji; Kaulpiboon, Jarunee; Prousoontorn, Manchumas; Limpaseni, Tipaporn; Pongsawasdi, Piamsook; Aoki, Kenji

    2007-10-01

    A newly isolated strain, 38C-2-1, produced alkaline and thermotolerant alpha-amylases and was identified as Bacillus halodurans. The enzymes were purified to homogeneity and named alpha-amylase I and II. These showed molecular masses of 105 and 75 kDa respectively and showed maximal activities at 50-60 degrees C and pH 10-11, and 42 and 38% relative activities at 30 degrees C. These results indicate that the enzymes are thermotolerant. The enzyme activity was not inhibited by a surfactant or a bleaching reagent used in detergents. A gene encoding alpha-amylase I was cloned and named amyI. Production of AmyI with a signal peptide repressed the growth of an Escherichia coli transformant. When enzyme production was induced by the addition of isopropyl beta-D(-)-thiogalactopyranoside in the late exponential growth phase, the highest enzyme yield was observed. It was 45-fold that of the parent strain 38C-2-1.

  16. Differential antimicrobial activity of silver nanoparticles to bacteria Bacillus subtilis and Escherichia coli, and toxicity to crop plant Zea mays and beneficial B. subtilis-inoculated Z. mays

    NASA Astrophysics Data System (ADS)

    Doody, Michael A.; Wang, Dengjun; Bais, Harsh P.; Jin, Yan

    2016-10-01

    As silver nanoparticles (AgNPs) have become increasingly used in commercial antimicrobial agents and industrial and military products, concerns are increasing over their broad environmental and health impacts and risks because they are finding their way to the environment. This study was designed to quantify the antimicrobial activity of citrate-coated AgNPs (c-AgNPs; transmission electron microscope size of 44.9 ± 7.2 nm) to two species of bacteria, i.e., Gram-positive Bacillus subtilis and Gram-negative Escherichia coli, and toxicity to a major crop plant Zea mays and beneficial bacteria-inoculated plant (i.e., B. subtilis-inoculated Z. mays symbiont). Our results reveal that the exposure of c-AgNPs significantly inhibited bacteria growth and altered their growth kinetics. Z. mays experienced significant sublethal effects including reduced root length and biomass, and hyper-accumulation of Ag in roots. The beneficial interactions between B. subtilis and Z. mays were weakened as well because both species suffered sublethal effects. Potential mechanisms leading to the antimicrobial activity and toxicity of c-AgNPs to the bacteria, plant, and plant-bacteria symbiont examined in this study were discussed. Taken together, our findings advance the current knowledge of AgNPs antimicrobial property or toxicity to bacteria, crop plant, and beneficial plant-bacteria symbiotic interaction, which is a critical component for NPs environmental impact and risk assessment.

  17. Antibody responses elicited in mice immunized with Bacillus subtilis vaccine strains expressing Stx2B subunit of enterohaemorragic Escherichia coli O157:H7

    PubMed Central

    Gomes, P.A.D.P.; Bentancor, L.V.; Paccez, J.D.; Sbrogio-Almeida, M.E.; Palermo, M.S.; Ferreira, R.C.C.; Ferreira, L.C.S.

    2009-01-01

    No effective vaccine or immunotherapy is presently available for patients with the hemolytic uremic syndrome (HUS) induced by Shiga-like toxin (Stx) produced by enterohaemorragic Escherichia coli (EHEC) strains, such as those belonging to the O157:H7 serotype. In this work we evaluated the performance of Bacillus subtilis strains, a harmless spore former gram-positive bacterium species, as a vaccine vehicle for the expression of Stx2B subunit (Stx2B). A recombinant B. subtilis vaccine strain expressing Stx2B under the control of a stress inducible promoter was delivered to BALB/c mice via oral, nasal or subcutaneous routes using both vegetative cells and spores. Mice immunized with vegetative cells by the oral route developed low but specific anti-Stx2B serum IgG and fecal IgA responses while mice immunized with recombinant spores developed anti-Stx2B responses only after administration via the parenteral route. Nonetheless, serum anti-Stx2B antibodies raised in mice immunized with the recombinant B. subtilis strain did not inhibit the toxic effects of the native toxin, both under in vitro and in vivo conditions, suggesting that either the quantity or the quality of the induced immune response did not support an effective neutralization of Stx2 produced by EHEC strains. PMID:24031368

  18. A Bacillus subtilis dnaG mutant harbours a mutation in a gene homologous to the dnaN gene of Escherichia coli.

    PubMed

    Ogasawara, N; Moriya, S; Mazza, G; Yoshikawa, H

    1986-01-01

    A dnaG mutation of Bacillus subtilis, dnaG5, was found to be linked closely to recF. We have reported previously that two putative dna genes, 'dnaA' and 'dnaN', highly homologous to Escherichia coli's dnaA and dnaN, respectively, were located adjacent to recF [Ogasawara et al., EMBO J., 4 (1985) 3345-3350]. Transformation by various fragments cloned from the 'dnaA'-recF region of the wild-type cell revealed that a 532-bp AluI fragment containing 5'-portion of the 'dnaN' gene could transform the dnaG5 mutation. The nucleotide (nt) sequence of the same fragment cloned from the mutant cell shows a single nt change in the ORF of 'dnaN' which in turn causes a single amino acid alteration from Gly to Arg. The 'dnaN' gene is now proven to be a dna gene, mutations in which result in instant arrest of chromosomal replication.

  19. Effect of chemical fixatives on accurate preservation of Escherichia coli and Bacillus subtilis structure in cells prepared by freeze-substitution

    SciTech Connect

    Graham, L.L.; Beveridge, T.J. )

    1990-04-01

    Five chemical fixatives were evaluated for their ability to accurately preserve bacterial ultrastructure during freeze-substitution of select Escherichia coli and Bacillus subtilis strains. Radioisotopes were specifically incorporated into the peptidoglycan, lipopolysaccharide, and nucleic acids of E. coli SFK11 and W7 and into the peptidoglycan and RNA of B. subtilis 168 and W23. The ease of extraction of radiolabels, as assessed by liquid scintillation counting during all stages of processing for freeze-substitution, was used as an indicator of cell structural integrity and retention of cellular chemical composition. Subsequent visual examination by electron microscopy was used to confirm ultrastructural conformation. The fixatives used were: 2% (wt/vol) osmium tetroxide and 2% (wt/vol) uranyl acetate; 2% (vol/vol) glutaraldehyde and 2% (wt/vol) uranyl acetate; 2% (vol/vol) acrolein and 2% (wt/vol) uranyl acetate; 2% (wt/vol) gallic acid; and 2% (wt/vol) uranyl acetate. All fixatives were prepared in a substitution solvent of anhydrous acetone. Extraction of cellular constituents depended on the chemical fixative used. A combination of 2% osmium tetroxide-2% uranyl acetate or 2% gallic acid alone resulted in optimum fixation as ascertained by least extraction of radiolabels. In both gram-positive and gram-negative organisms, high levels of radiolabel were detected in the processing fluids in which 2% acrolein-2% uranyl acetate, 2% glutaraldehyde-2% uranyl acetate, or 2% uranyl acetate alone were used as fixatives. Ultrastructural variations were observed in cells freeze-substituted in the presence of different chemical fixatives. We recommend the use of osmium tetroxide and uranyl acetate in acetone for routine freeze-substitution of eubacteria, while gallic acid is recommended for use when microanalytical processing necessitates the omission of osmium.

  20. Inactivation of Escherichia coli, Bacteriophage MS2, and Bacillus Spores under UV/H2O2 and UV/Peroxydisulfate Advanced Disinfection Conditions.

    PubMed

    Sun, Peizhe; Tyree, Corey; Huang, Ching-Hua

    2016-04-19

    Ultraviolet light (UV) combined with peroxy chemicals, such as H2O2 and peroxydisulfate (PDS), have been considered potentially highly effective disinfection processes. This study investigated the inactivation of Escherichia coli, bacteriophage MS2, and Bacillus subtilis spores as surrogates for pathogens under UV/H2O2 and UV/PDS conditions, with the aim to provide further understanding of UV-based advanced disinfection processes (ADPs). Results showed that one additional log of inactivation of E. coli was achieved with 0.3 mM H2O2 or PDS at 5.2 × 10(-5) Einstein·L(-1) photo fluence (at 254 nm) compared with UV irradiation alone. Addition of H2O2 and PDS greatly enhanced the inactivation rate of MS2 by around 15 folds and 3 folds, respectively, whereas the inactivation of B. subtilis spores was slightly enhanced. Reactive species responsible for the inactivation were identified to be •OH, SO4(·-), and CO3(·-) based on manipulation of solution conditions. The CT value of each reactive species was calculated with respect to each microbial surrogate, which showed that the disinfection efficacy ranked as •OH > SO4(·-) > CO3(·-) ≫ O2(·-)/HO2(·). A comprehensive dynamic model was developed and successfully predicted the inactivation of the microbial surrogates in surface water and wastewater matrices. The concepts of UV-efficiency and EE/O were employed to provide a cost-effective evaluation for UV-based ADPs. Overall, the present study suggests that it will be beneficial to upgrade UV disinfection to UV/H2O2 ADP for the inactivation of viral pathogens.

  1. A 20-kilodalton protein is required for efficient production of the Bacillus thuringiensis subsp. israelensis 27-kilodalton crystal protein in Escherichia coli.

    PubMed Central

    Adams, L F; Visick, J E; Whiteley, H R

    1989-01-01

    The 27-kilodalton (kDa) mosquitocidal protein gene from Bacillus thuringiensis subsp. israelensis has been cloned as a 10-kilobase (kb) HindIII fragment from plasmid DNA; efficient expression in Escherichia coli KM1 depends on a region of DNA located approximately 4 kb upstream (K. McLean and H. R. Whiteley, J. Bacteriol. 169:1017-1023, 1987). We have cloned the upstream DNA region and show that it contains a complete open reading frame (ORF) encoding a protein with a molecular mass of 19,584 Da. Sequencing of adjacent stretches of DNA revealed two partial ORFs: one has 55.2% identity in an overlap of 319 amino acids to the putative transposase of IS231 of B. thuringiensis subsp. thuringiensis, and the other, a 78-codon partial ORF, may be the carboxyl terminus of the 67-kDa protein previously observed in maxicells of strain KM1. A 0.8-kb fragment containing only the 20-kDa protein gene greatly enhanced the expression of the 27-kDa protein in E. coli. The introduction of nonsense codons into the 20-kDa protein gene ORF abolished this effect, indicating that the gene product, not the mRNA or DNA, is required for the enhancement. The effect of the 20-kDa protein gene on various fusions of lacZ to the 27-kDa protein gene suggests that the 20-kDa protein acts after the initiation of translation of the 27-kDa protein gene. Images PMID:2644205

  2. ANIMAL ENTEROTOXIGENIC ESCHERICHIA COLI

    PubMed Central

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

    2016-01-01

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

  3. Cytoplasmic pH response to acid stress in individual cells of Escherichia coli and Bacillus subtilis observed by fluorescence ratio imaging microscopy.

    PubMed

    Martinez, Keith A; Kitko, Ryan D; Mershon, J Patrick; Adcox, Haley E; Malek, Kotiba A; Berkmen, Melanie B; Slonczewski, Joan L

    2012-05-01

    The ability of Escherichia coli and Bacillus subtilis to regulate their cytoplasmic pH is well studied in cell suspensions but is poorly understood in individual adherent cells and biofilms. We observed the cytoplasmic pH of individual cells using ratiometric pHluorin. A standard curve equating the fluorescence ratio with pH was obtained by perfusion at a range of external pH 5.0 to 9.0, with uncouplers that collapse the transmembrane pH difference. Adherent cells were acid stressed by switching the perfusion medium from pH 7.5 to pH 5.5. The E. coli cytoplasmic pH fell to a value that varied among individual cells (range of pH 6.2 to 6.8), but a majority of cells recovered (to pH 7.0 to 7.5) within 2 min. In an E. coli biofilm, cells shifted from pH 7.5 to pH 5.5 failed to recover cytoplasmic pH. Following a smaller shift (from pH 7.5 to pH 6.0), most biofilm cells recovered fully, although the pH decreased further than that of isolated adherent cells, and recovery took longer (7 min or longer). Some biofilm cells began to recover pH and then failed, a response not seen in isolated cells. B. subtilis cells were acid shifted from pH 7.5 to pH 6.0. In B. subtilis, unlike the case with E. coli, cytoplasmic pH showed no "overshoot" but fell to a level that was maintained. This level of cytoplasmic pH post-acid shift varied among individual B. subtilis cells (range of pH, 7.0 to 7.7). Overall, the cytoplasmic pHs of individual bacteria show important variation in the acid stress response, including novel responses in biofilms.

  4. Survival of foodborne pathogenic bacteria (Bacillus cereus, Escherichia coli O157:H7, Salmonella enterica serovar Typhimurium, Staphylococcus aureus, and Listeria monocytogenes) and Bacillus cereus spores in fermented alcoholic beverages (beer and refined rice wine).

    PubMed

    Kim, S A; Kim, N H; Lee, S H; Hwang, I G; Rhee, M S

    2014-03-01

    Only limited information is available on the microbiological safety of fermented alcoholic beverages because it is still a common belief that such beverages do not provide a favorable environment for bacterial growth and survival. Thus, in this study, we examined the survival of major foodborne pathogens and spores in fermented alcoholic beverages. Foodborne pathogens (Bacillus cereus, Escherichia coli O157:H7, Listeria monocytogenes, Salmonella enterica serovar Typhimurium, and Staphylococcus aureus) and B. cereus spores (initial population, 3 to 4 log CFU/ml) were inoculated separately into three types of beer and refined rice wine, which were then stored at 5 and 22°C. Bacterial counts were assayed periodically for up to 28 days. Vegetative B. cereus counts decreased rapidly, whereas B. cereus spore counts remained constant (P > 0.05) for a long period of time in all beverages. Vegetative B. cereus cells formed spores in beer at 5 and 22°C, and the spores survived for long periods. Among vegetative cells, E. coli O157:H7 had the highest survival (only 1.49 to 1.56 log reduction during 28 days in beer at 5°C). Beer and refined rice wine supported microbial survival from several days to several weeks. Our results appear to contradict the common belief that pathogens cannot survive in alcoholic beverages. Long-term survival of pathogens (especially B. cereus and E. coli O157:H7) in beer and refined rice wine should be taken into consideration by the manufacturers of these beverages. This study provides basic information that should help further research into microbial survival in alcoholic beverages and increase the microbiological safety regulation of fermented alcoholic beverages.

  5. Gravity sensing by Escherichia coli.

    PubMed

    Shimoshige, Hirokazu; Kobayashi, Hideki; Shimamura, Shigeru; Usami, Ron

    2010-01-01

    We investigated the growth and protein profile of Escherichia coli under various gravity strengths to determine the effects of hypergravity on biochemical reactions. E. coli grows at less than 7,500 g without inhibition. Hypergravity induced OmpW and Antigen 43. Changes in gravity strength altered the expression levels of these proteins. This suggests that hypergravity regulates gene expression in bacteria.

  6. Prokaryotic regulatory systems biology: Common principles governing the functional architectures of Bacillus subtilis and Escherichia coli unveiled by the natural decomposition approach.

    PubMed

    Freyre-González, Julio A; Treviño-Quintanilla, Luis G; Valtierra-Gutiérrez, Ilse A; Gutiérrez-Ríos, Rosa María; Alonso-Pavón, José A

    2012-10-31

    Escherichia coli and Bacillus subtilis are two of the best-studied prokaryotic model organisms. Previous analyses of their transcriptional regulatory networks have shown that they exhibit high plasticity during evolution and suggested that both converge to scale-free-like structures. Nevertheless, beyond this suggestion, no analyses have been carried out to identify the common systems-level components and principles governing these organisms. Here we show that these two phylogenetically distant organisms follow a set of common novel biologically consistent systems principles revealed by the mathematically and biologically founded natural decomposition approach. The discovered common functional architecture is a diamond-shaped, matryoshka-like, three-layer (coordination, processing, and integration) hierarchy exhibiting feedback, which is shaped by four systems-level components: global transcription factors (global TFs), locally autonomous modules, basal machinery and intermodular genes. The first mathematical criterion to identify global TFs, the κ-value, was reassessed on B. subtilis and confirmed its high predictive power by identifying all the previously reported, plus three potential, master regulators and eight sigma factors. The functionally conserved cores of modules, basal cell machinery, and a set of non-orthologous common physiological global responses were identified via both orthologous genes and non-orthologous conserved functions. This study reveals novel common systems principles maintained between two phylogenetically distant organisms and provides a comparison of their lifestyle adaptations. Our results shed new light on the systems-level principles and the fundamental functions required by bacteria to sustain life. Copyright © 2012 Elsevier B.V. All rights reserved.

  7. Physicochemical interaction of Escherichia coli cell envelopes and Bacillus subtilis cell walls with two clays and ability of the composite to immobilize heavy metals from solution.

    PubMed Central

    Walker, S G; Flemming, C A; Ferris, F G; Beveridge, T J; Bailey, G W

    1989-01-01

    Isolated Escherichia coli K-12 cell envelopes or Bacillus subtilis 168 cell walls were reacted with smectite or kaolinite clay in distilled deionized water (pH 6.0); unbound envelopes or walls were separated by sucrose density gradient centrifugation, and the extent of adsorption was calculated. At saturation, both clays adsorbed approximately 1.0 mg (dry weight) of envelopes or walls per mg (dry weight) of clay. Clays showed a preference for edge-on orientation with both walls and envelopes, which was indicative of an aluminum polynuclear bridging mechanism between the wall or envelope surface and the clay edge. The addition of heavy metals increased the incidence of planar surface orientations, which suggested that multivalent metal cation bridging was coming into play and was of increasing importance. The metal-binding capacity of isolated envelopes, walls, clays, and envelope-clay or wall-clay mixtures was determined by atomic absorption spectroscopy after exposure to aqueous 5.0 mM Ag+, Cu2+, Cd2+, Ni2+, Pb2+, Zn2+, and Cr3+ nitrate salt solutions at pHs determined by the buffering capacity of wall, envelope, clay, or composite system. The order of metal uptake was walls greater than envelopes greater than smectite clay greater than kaolinite clay for the individual components, and walls plus smectite greater than walls plus kaolinite greater than envelopes plus smectite greater than envelopes plus kaolinite for the mixtures. On a dry-weight basis, the envelope-clay and wall-clay mixtures bound 20 to 90% less metal than equal amounts of the individual components did.(ABSTRACT TRUNCATED AT 250 WORDS) Images PMID:2516433

  8. Non-recombinant display of the B subunit of the heat labile toxin of Escherichia coli on wild type and mutant spores of Bacillus subtilis

    PubMed Central

    2013-01-01

    Background Mucosal infections are a major global health problem and it is generally accepted that mucosal vaccination strategies, able to block infection at their entry site, would be preferable with respect to other prevention approaches. However, there are still relatively few mucosal vaccines available, mainly because of the lack of efficient delivery systems and of mucosal adjuvants. Recombinant bacterial spores displaying a heterologous antigen have been shown to induce protective immune responses and, therefore, proposed as a mucosal delivery system. A non-recombinant approach has been recently developed and tested to display antigens and enzymes. Results We report that the binding subunit of the heat-labile toxin (LTB) of Escherichia coli efficiently adsorbed on the surface of Bacillus subtilis spores. When nasally administered to groups of mice, spore-adsorbed LTB was able to induce a specific immune response with the production of serum IgG, fecal sIgA and of IFN-γ in spleen and mesenteric lymph nodes (MLN) of the immunized animals. Dot blotting experiments showed that the non-recombinant approach was more efficient than the recombinant system in displaying LTB and that the efficiency of display could be further increased by using mutant spores with an altered surface. In addition, immunofluorescence microscopy experiments showed that only when displayed on the spore surface by the non-recombinant approach LTB was found in its native, pentameric form. Conclusion Our results indicate that non-recombinant spores displaying LTB pentamers can be administered by the nasal route to induce a Th1-biased, specific immune response. Mutant spores with an altered coat are more efficient than wild type spores in adsorbing the antigen, allowing the use of a reduced number of spores in immunization procedures. Efficiency of display, ability to display the native form of the antigen and to induce a specific immune response propose this non-recombinant delivery system as

  9. Non-recombinant display of the B subunit of the heat labile toxin of Escherichia coli on wild type and mutant spores of Bacillus subtilis.

    PubMed

    Isticato, Rachele; Sirec, Teja; Treppiccione, Lucia; Maurano, Francesco; De Felice, Maurilio; Rossi, Mauro; Ricca, Ezio

    2013-10-29

    Mucosal infections are a major global health problem and it is generally accepted that mucosal vaccination strategies, able to block infection at their entry site, would be preferable with respect to other prevention approaches. However, there are still relatively few mucosal vaccines available, mainly because of the lack of efficient delivery systems and of mucosal adjuvants. Recombinant bacterial spores displaying a heterologous antigen have been shown to induce protective immune responses and, therefore, proposed as a mucosal delivery system. A non-recombinant approach has been recently developed and tested to display antigens and enzymes. We report that the binding subunit of the heat-labile toxin (LTB) of Escherichia coli efficiently adsorbed on the surface of Bacillus subtilis spores. When nasally administered to groups of mice, spore-adsorbed LTB was able to induce a specific immune response with the production of serum IgG, fecal sIgA and of IFN-γ in spleen and mesenteric lymph nodes (MLN) of the immunized animals. Dot blotting experiments showed that the non-recombinant approach was more efficient than the recombinant system in displaying LTB and that the efficiency of display could be further increased by using mutant spores with an altered surface. In addition, immunofluorescence microscopy experiments showed that only when displayed on the spore surface by the non-recombinant approach LTB was found in its native, pentameric form. Our results indicate that non-recombinant spores displaying LTB pentamers can be administered by the nasal route to induce a Th1-biased, specific immune response. Mutant spores with an altered coat are more efficient than wild type spores in adsorbing the antigen, allowing the use of a reduced number of spores in immunization procedures. Efficiency of display, ability to display the native form of the antigen and to induce a specific immune response propose this non-recombinant delivery system as a powerful mucosal vaccine

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

    PubMed

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

    2014-03-01

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

  11. Molecular cloning and expression of a novel mesophilic alkaline protease from Bacillus sp. L010 in Escherichia coli.

    PubMed

    Li, Dan; Huang, Fei; Xia, Mengyun; Jiang, Yan; Yang, Yi

    2013-11-04

    Bacteria producing proteases were isolated and selected from the environment and the alkaline proteases with superior performance for commercial exploitation were screened. The strain producing extracellular proteases was isolated by a casein plate and was identified by biochemical and morphological tests and by 16S rDNA sequence analysis. To acquire the open reading frame (ORF) of the protease, degenerate primers designing and genome walking method were used. The precursor and mature peptide of the protease were recombinant expressed in BL21 (DE3). After purification of the active protease, the characteristics and the catalytic ability were detected using synthetic peptide succinyl-Ala-Ala-Pro-Phe-p-nitroanilide as the substrates. Strain L010 isolated was named as Bacillus sp. L010 after identification. The ORF of the protease was 1149-bp long and encoded a protein of 382 amino acids comprised with a 30-residual signal peptide, a 77-residual propeptide, and a 275-residual mature protein, and the encoded protein was one of subtilisins-a member of serine proteases and designated as SprD. The precursor of SprD (pro-SprD) autoprocessed into active SprD mediated by the propeptide when pro-SprD was recombinant expressed in BL21 (DE3). The enzyme exhibited high catalytic efficiency (K(cat)/K(m)) towards synthesis substrates with optimal activity at 70 degrees C and pH 9 - 10. SprD was stable over a range of pH 7.0 to 10.0 and was thermal stable at 25 degrees C - 60 degrees C. The high stability of SprD towards alkaline conditions (pH 7 - 10) and under temperature 25 degrees C - 60 degrees C and the high catalytic efficiency suggested that the protease would find research value and potential applications.

  12. Efficacy of Bacillus subtilis and bacitracin methylene disalicylate on growth performance, digestibility, blood metabolites, immunity, and intestinal microbiota after intramuscular inoculation with Escherichia coli in broilers.

    PubMed

    Manafi, M; Khalaji, S; Hedayati, M; Pirany, N

    2017-05-01

    This experiment was conducted to evaluate the effect of Bacillus subtilis (BS) on broiler performance and health after intramuscular inoculation with E. coli and compare its effect with a growth promoter antibiotic. In a completely randomized design manner, 360 male Ross 308 chicks were divided into 6 treatments and 5 replicates of 12 chicks in each replicate. Experimental treatments included control diet, control + E. coli (0.5 mL of culture containing 108 CFU of E. coli/ml), control + 0.1% BS, control + 0.05% bacitracin methylene disalicylate (BMD), control + E. coli and BS, and control + E. coli and BMD in a factorial arrangement (3 × 2). Addition of BMD or BS to the control diet significantly (P < 0.01) increased body weight and decreased FCR, but E. coli challenge adversely reduced (P < 0.01) body weight and increased FCR, so that the addition of BMD or BS did not compensate growth reduction. E. coli challenged chicks had the lowest vaccine titers for ND, IB, AI, and IBD and the highest were observed in chicks fed BS. The E. coli challenge significantly (P < 0.01) increased albumin, globulin, cholesterol, triglyceride, LDL, ALT, and ALP indices. Addition of BMD and BS decreased albumin and globulin in challenged chick's plasma but had no effect on plasma lipid profile concentration. The E. coli challenge decreased villus height and increased crypt depth and goblet cell numbers significantly (P < 0.01). In birds subjected to BMD or BS, crypt depth decreased and villus height increased (P < 0.01), compared with the control diet. Challenge of E. coli significantly (P < 0.01) increased the bacterial population of E. coli, coliforms, and Salmonella in cecal parts of broilers' intestines. In challenged birds receiving BMD or BS, E. coli, coliform, and Salmonella populations of ceca showed a significant (P < 0.01) reduction. Both BMD and BS increased the digestibility of nutrients significantly (P < 0.01), but a reduction was observed in E. coli challenged groups

  13. Engineering Escherichia coli for methanol conversion.

    PubMed

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

    2015-03-01

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

  14. Enhanced production of carboxymethylcellulase of a marine microorganism, Bacillus subtilis subsp. subtilis A-53 in a pilot-scaled bioreactor by a recombinant Escherichia coli JM109/A-53 from rice bran.

    PubMed

    Lee, Eun-Jung; Lee, Bo-Hwa; Kim, Bo-Kyung; Lee, Jin-Woo

    2013-05-01

    A gene encoding the carboxymethylcellulase (CMCase) of a marine bacterium, Bacillus subtilis subsp. subtilis A-53, was cloned in Escherichia coli JMB109 and the recombinant strain was named as E. coli JMB109/A-53. The optimal conditions of rice bran, ammonium chloride, and initial pH of the medium for cell growth, extracted by Design Expert Software based on response surface methodology, were 100.0 g/l, 7.5 g/l, and 7.0, respectively, whereas those for production of CMCase were 100.0 g/l, 7.5 g/l, and 8.0. The optimal temperatures for cell growth and the production of CMCase by E. coli JM109/A-53 were found to be and 40 and 35 °C, respectively. The optimal agitation speed and aeration rate of a 7 l bioreactor for cell growth were 400 rpm and 1.5 vvm, whereas those for production of CMCase were 400 rpm and 0.5 vvm. The optimal inner pressure for cell growth was 0.06 MPa, which was the same as that for production of CMCase. The production of CMCase by E. coli JM109/A-53 under optimized conditions was 880.2 U/ml, which was 2.9 times higher than that before optimization. In this study, rice bran and ammonium chloride were developed as carbon and nitrogen source for production of CMCase by a recombinant E. coli JM109/A-53 and the productivity of E. coli JM109/A-53 was 5.9 times higher than that of B. subtilis subp. subtilis A-53.

  15. Exonuclease IX of Escherichia coli.

    PubMed Central

    Shafritz, K M; Sandigursky, M; Franklin, W A

    1998-01-01

    The bacteria Escherichia coli contains several exonucleases acting on both double- and single-stranded DNA and in both a 5'-->3' and 3'-->5' direction. These enzymes are involved in replicative, repair and recombination functions. We have identified a new exonuclease found in E.coli, termed exonuclease IX, that acts preferentially on single-stranded DNA as a 3'-->5' exonuclease and also functions as a 3'-phosphodiesterase on DNA containing 3'-incised apurinic/apyrimidinic (AP) sites to remove the product trans -4-hydroxy-2-pentenal 5-phosphate. The enzyme showed essentially no activity as a deoxyribophosphodiesterase acting on 5'-incised AP sites. The activity was isolated as a glutathione S-transferase fusion protein from a sequence of the E.coli genome that was 60% identical to a 260 bp region of the small fragment of the DNA polymerase I gene. The protein has a molecular weight of 28 kDa and is free of AP endonuclease and phosphatase activities. Exonuclease IX is expressed in E.coli , as demonstrated by reverse transcription-PCR, and it may function in the DNA base excision repair and other pathways. PMID:9592142

  16. Robust growth of Escherichia coli.

    PubMed

    Wang, Ping; Robert, Lydia; Pelletier, James; Dang, Wei Lien; Taddei, Francois; Wright, Andrew; Jun, Suckjoon

    2010-06-22

    The quantitative study of the cell growth has led to many fundamental insights in our understanding of a wide range of subjects, from the cell cycle to senescence. Of particular importance is the growth rate, whose constancy represents a physiological steady state of an organism. Recent studies, however, suggest that the rate of elongation during exponential growth of bacterial cells decreases cumulatively with replicative age for both asymmetrically and symmetrically dividing organisms, implying that a "steady-state" population consists of individual cells that are never in a steady state of growth. To resolve this seeming paradoxical observation, we studied the long-term growth and division patterns of Escherichia coli cells by employing a microfluidic device designed to follow steady-state growth and division of a large number of cells at a defined reproductive age. Our analysis of approximately 10(5) individual cells reveals a remarkable stability of growth whereby the mother cell inherits the same pole for hundreds of generations. We further show that death of E. coli is not purely stochastic but is the result of accumulating damages. We conclude that E. coli, unlike all other aging model systems studied to date, has a robust mechanism of growth that is decoupled from cell death.

  17. Peptidoglycan Hydrolases of Escherichia coli

    PubMed Central

    van Heijenoort, Jean

    2011-01-01

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

  18. [Virulence mechanisms of enteropathogenic Escherichia coli].

    PubMed

    Farfán-García, Ana Elvira; Ariza-Rojas, Sandra Catherine; Vargas-Cárdenas, Fabiola Andrea; Vargas-Remolina, Lizeth Viviana

    2016-08-01

    Acute diarrheal disease (ADD) is a global public health problem, especially in developing countries and is one of the causes of mortality in children under five. ADD etiologic agents include viruses, bacteria and parasites in that order. Escherichia coli bacteria it is classified as a major diarrheagenic agent and transmitted by consuming contaminated water or undercooked foods. This review compiled updates on information virulence factors and pathogenic mechanisms involved in adhesion and colonization of seven pathotypes of E. coli called enteropathogenic E. coli (EPEC), enterotoxigenic E. coli (ETEC), enteroinvasive E. coli (EIEC), shigatoxigenic E. coli (STEC), enteroaggregative E. coli (EAEC) and diffusely-adherent E. coli (DAEC). A final pathotype, adherent-invasive E. coli (AIEC) associated with Crohn's disease was also reviewed. The diarrheagenic pathotypes of E. coli affect different population groups and knowledge of the molecular mechanisms involved in the interaction with the human is important to guide research towards the development of vaccines and new tools for diagnosis and control.

  19. Generation of recombinant bacillus Calmette-Guérin and Mycobacterium smegmatis expressing BfpA and intimin as vaccine vectors against enteropathogenic Escherichia coli.

    PubMed

    Vasconcellos, Halyka Luzorio Franzotti; Scaramuzzi, Karina; Nascimento, Ivan Pereira; Da Costa Ferreira, Jorge M; Abe, Cecilia M; Piazza, Roxane M F; Kipnis, Andre; Dias da Silva, Wilmar

    2012-09-07

    Enteropathogenic Escherichia coli (EPEC) is an important cause of diarrhea in children. EPEC adheres to the intestinal epithelium and causes attaching and effacing (A/E) lesions. Recombinant Mycobacterium smegmatis (Smeg) and Mycobacterium bovis BCG strains were constructed to express either BfpA or intimin. The entire bfpA gene and a portion of the intimin gene were amplified by PCR from EPEC genomic DNA and inserted into the pMIP12 vector at the BamHI/KpnI sites. The pMIP_bfpA and pMIP_intimin vectors were introduced separately into Smeg and BCG. Recombinant clones were selected based on kanamycin resistance and designated rSmeg_pMIP_(bfpA or intimin) and rBCG_pMIP_(bfpA or intimin). The expression of bfpA and intimin was detected by Immunoblotting using polyclonal anti-BfpA and anti-intimin antibodies. The immunogenicity of these proteins was assessed in C57BL/6 mice by assaying the feces and serum for the presence of anti-BfpA and anti-intimin IgA and IgG antibodies. TNF-α and INF-γ were produced in vitro by spleen cells from mice immunized with recombinant BfpA, whereas TNF-γ was produced in mice immunized with recombinant intimin. The adhesion of EPEC (E2348/69) to HEp-2 target cells was blocked by IgA or IgG antibodies from mice immunized with recombinant BfpA or intimin but not by antibodies from non-immunized mice. Immunogenic non-infectious vectors containing relevant EPEC virulence genes may be promising vaccine candidates.

  20. Structure of Escherichia coli tryptophanase.

    PubMed

    Ku, Shao Yang; Yip, Patrick; Howell, P Lynne

    2006-07-01

    Pyridoxal 5'-phosphate (PLP) dependent tryptophanase has been isolated from Escherichia coli and its crystal structure has been determined. The structure shares the same fold with and has similar quaternary structure to Proteus vulgaris tryptophanase and tyrosine-phenol lyase, but is found in a closed conformation when compared with these two enzymes. The tryptophanase structure, solved in its apo form, does not have covalent PLP bound in the active site, but two sulfate ions. The sulfate ions occupy the phosphoryl-binding site of PLP and the binding site of the alpha-carboxyl of the natural substrate tryptophan. One of the sulfate ions makes extensive interactions with both the transferase and PLP-binding domains of the protein and appears to be responsible for holding the enzyme in its closed conformation. Based on the sulfate density and the structure of the P. vulgaris enzyme, PLP and the substrate tryptophan were modeled into the active site. The resulting model is consistent with the roles of Arg419 in orienting the substrate to PLP and acidifying the alpha-proton of the substrate for beta-elimination, Lys269 in the formation and decomposition of the PLP quinonoid intermediate, Arg230 in orienting the substrate-PLP intermediates in the optimal conformation for catalysis, and His463 and Tyr74 in determining substrate specificity and suggests that the closed conformation observed in the structure could be induced by substrate binding and that significant conformational changes occur during catalysis. A catalytic mechanism for tryptophanase is proposed. Since E. coli tryptophanase has resisted forming diffraction-quality crystals for many years, the molecular surface of tryptophanase has been analyzed in various crystal forms and it was rationalized that strong crystal contacts occur on the flat surface of the protein and that the size of crystal contact surface seems to correlate with the diffraction quality of the crystal.

  1. Structure of Escherichia Coli Tryptophanase

    SciTech Connect

    Ku,S.; Yip, P.; Howell, P.

    2006-01-01

    Pyridoxal 5'-phosphate (PLP) dependent tryptophanase has been isolated from Escherichia coli and its crystal structure has been determined. The structure shares the same fold with and has similar quaternary structure to Proteus vulgaris tryptophanase and tyrosine-phenol lyase, but is found in a closed conformation when compared with these two enzymes. The tryptophanase structure, solved in its apo form, does not have covalent PLP bound in the active site, but two sulfate ions. The sulfate ions occupy the phosphoryl-binding site of PLP and the binding site of the {alpha}-carboxyl of the natural substrate tryptophan. One of the sulfate ions makes extensive interactions with both the transferase and PLP-binding domains of the protein and appears to be responsible for holding the enzyme in its closed conformation. Based on the sulfate density and the structure of the P. vulgaris enzyme, PLP and the substrate tryptophan were modeled into the active site. The resulting model is consistent with the roles of Arg419 in orienting the substrate to PLP and acidifying the {alpha}-proton of the substrate for {beta}-elimination, Lys269 in the formation and decomposition of the PLP quinonoid intermediate, Arg230 in orienting the substrate-PLP intermediates in the optimal conformation for catalysis, and His463 and Tyr74 in determining substrate specificity and suggests that the closed conformation observed in the structure could be induced by substrate binding and that significant conformational changes occur during catalysis. A catalytic mechanism for tryptophanase is proposed. Since E. coli tryptophanase has resisted forming diffraction-quality crystals for many years, the molecular surface of tryptophanase has been analyzed in various crystal forms and it was rationalized that strong crystal contacts occur on the flat surface of the protein and that the size of crystal contact surface seems to correlate with the diffraction quality of the crystal.

  2. Secondary structure features of ribosomal RNA species within intact ribosomal subunits and efficiency of RNA-protein interactions in thermoacidophilic (Caldariella acidophila, Bacillus acidocaldarius) and mesophilic (Escherichia coli) bacteria.

    PubMed

    Cammarano, P; Mazzei, F; Londei, P; Teichner, A; de Rosa, M; Gambacorta, A

    1983-08-02

    Ribosomal subunits of Caldariella acidophila (max.growth temp., 90 degrees C) have been compared to subunits of Bacillus acidocaldarius (max. growth temp., 70 degrees C) and Escherichia coli (max. growth temp., 47 degrees C) with respect to (a) bihelical content of rRNA; (b) G . C content of bihelical domains and (c) tightness of rRNA-protein interactions. The principal results are as follows. Subunits of C. acidophilia ribosomes (Tm = 90-93 degrees C) exhibit considerable thermal tolerance over their B. acidocaldarius (Tm = 77 degrees C) and E. coli counterparts (Tm = 72 degrees C). Based on the "melting' hyperchromicities of the intact ribosomal subunits a 51-55% fraction of the nucleotides appears to participate in hydrogen-bonded base pairing regardless of ribosome source, whereas a larger fraction, 67-70%, appears to be involved in hydrogen bonding in the naked rRNA species. The G . C content of bihelical domains of both free and ribosome-bound rRNA increases with increasing thermophily; based on hyperchromicity dispersion spectra of intact subunits and free rRNA, the bihelical parts of C. acidophila rRNA are estimated to contain 63-64% G . C, compared to 58.5% G . C for B. acidocaldarius and 55% G . C for E. coli. The increment of ribosome Tm values with increasing thermophily is greater than the increase in Tm for the free rRNA, indicating that within ribosomes bihelical domains of the thermophile rRNA species are stabilized more efficiently than their mesophile counterparts by proteins or/ and other component(s). The efficiency of the rRNA-protein interactions in the mesophile and thermophile ribosomes has been probed by comparing the releases, with LiCl-urea, of the rRNA species from the corresponding ribosomal subunits stuck to a Celite column through their protein moiety; it has been established that the release of C. acidophila rRNA from the Celite-bound ribosomes occurs at salt-urea concentrations about 4-fold higher than those required to release r

  3. Succinate production in Escherichia coli

    PubMed Central

    Thakker, Chandresh; Martínez, Irene; San, Ka-Yiu; Bennett, George N.

    2012-01-01

    Succinate has been recognized as an important platform chemical that can be produced from biomass. While a number of organisms are capable of succinate production naturally, this review focuses on the engineering of Escherichia coli for production of the four-carbon dicarboxylic acid. Important features of a succinate production system are to achieve optimal balance of reducing equivalents generated by consumption of the feedstock, while maximizing the amount of carbon that is channeled to the product. Aerobic and anaerobic production strains have been developed and applied to production from glucose as well as other abundant carbon sources. Metabolic engineering methods and strain evolution have been used and supplemented by the recent application of systems biology and in silico modeling tools to construct optimal production strains. The metabolic capacity of the production strain, as well as the requirement for efficient recovery of succinate and the reliability of the performance under scale-up are important in the overall process. The costs of the overall biorefinery compatible process will determine the economical commercialization of succinate and its impact in larger chemical markets. PMID:21932253

  4. Escherichia coli survival in waters: Temperature dependence

    EPA Science Inventory

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

  5. Strategies for Protein Overproduction in Escherichia coli.

    ERIC Educational Resources Information Center

    Mott, John E.

    1984-01-01

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

  6. Strategies for Protein Overproduction in Escherichia coli.

    ERIC Educational Resources Information Center

    Mott, John E.

    1984-01-01

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

  7. Escherichia coli survival in waters: Temperature dependence

    EPA Science Inventory

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

  8. A comparative study of β-1, 4-endoglucanase (possessing β-1, 4-exoglucanase activity) from Bacillus subtilis LH expressed in Pichia pastoris GS115 and Escherichia coli Rosetta (DE3).

    PubMed

    Zhao, Xi-Hua; Wang, Wei; Wang, Feng-Qing; Wei, Dong-Zhi

    2012-04-01

    β-1, 4-Endoglucanase (EG) from Bacillus subtilis LH was expressed in Escherichia coli Rosetta (DE3) and Pichia pastoris GS115, respectively. The CMCase activity of EG (EGE) from the cell lysate of DE3 reached 20,010U/ml, and that of EG (EGP) from the supernatant of GS115 was only 2008U/ml. EGE and EGP were bifunctional cellulases excluding β-1, 4-glucosidase (BGL). The CMCases of them, optimally active at 65°C and pH 6.8, exhibited more than 80% residual activity at pH 5-10 and 60% activity at 40-70°C and pH 5-9. EGE (EGP) mixed with BGL had more than 1.5-fold higher CMCase and filter paperase activities compared to EGE (EGP). N-glycosylation protected EGP from immobilized-papain attack and accounted for 30kDa and a higher thermostability, whereas EGE was decomposed into a 33kDa active truncated EG (EGT) and two 18kDa fragments. EGE and EGP performed much better than EGT in denim biostoning. Copyright © 2011 Elsevier Ltd. All rights reserved.

  9. Expression in Escherichia coli of a sub-gene encoding the lipoyl and peripheral subunit-binding domains of the dihydrolipoamide acetyltransferase component of the pyruvate dehydrogenase complex of Bacillus stearothermophilus.

    PubMed

    Hipps, D S; Perham, R N

    1992-05-01

    A sub-gene encoding the N-terminal 170 residues of the dihydrolipoamide acetyltransferase chain of the pyruvate dehydrogenase multienzyme complex of Bacillus stearothermophilus was over-expressed in Escherichia coli. The expressed polypeptide consists of the lipoyl domain, inter-domain linker and peripheral subunit-binding domain; these were found to have folded into their native functional conformations as judged by reductive acetylation of the lipoyl domain, limited proteolysis of the linker region and ability to bind the dihydrolipoamide dehydrogenase dimer. The di-domain was largely (80%) unlipoylated; a small proportion (4%) was correctly modified with lipoic acid and the remainder (16%) was aberrantly modified with octanoic acid. A polyclonal antiserum was raised that recognized both the di-domain and the individual component domains. The 400 MHz 1H-n.m.r. spectrum of the di-domain showed resonances corresponding to those seen in spectra of the lipoyl domain, plus others characteristic of amino acid residues in the flexible linker region. Further, as yet unidentified, resonances are likely to be derived from the peripheral subunit-binding domain. The existence and independent folding of the peripheral subunit-binding domain is thus confirmed and its purification in large-scale amounts for detailed structural analysis is now possible.

  10. Expression in Escherichia coli of a sub-gene encoding the lipoyl and peripheral subunit-binding domains of the dihydrolipoamide acetyltransferase component of the pyruvate dehydrogenase complex of Bacillus stearothermophilus.

    PubMed Central

    Hipps, D S; Perham, R N

    1992-01-01

    A sub-gene encoding the N-terminal 170 residues of the dihydrolipoamide acetyltransferase chain of the pyruvate dehydrogenase multienzyme complex of Bacillus stearothermophilus was over-expressed in Escherichia coli. The expressed polypeptide consists of the lipoyl domain, inter-domain linker and peripheral subunit-binding domain; these were found to have folded into their native functional conformations as judged by reductive acetylation of the lipoyl domain, limited proteolysis of the linker region and ability to bind the dihydrolipoamide dehydrogenase dimer. The di-domain was largely (80%) unlipoylated; a small proportion (4%) was correctly modified with lipoic acid and the remainder (16%) was aberrantly modified with octanoic acid. A polyclonal antiserum was raised that recognized both the di-domain and the individual component domains. The 400 MHz 1H-n.m.r. spectrum of the di-domain showed resonances corresponding to those seen in spectra of the lipoyl domain, plus others characteristic of amino acid residues in the flexible linker region. Further, as yet unidentified, resonances are likely to be derived from the peripheral subunit-binding domain. The existence and independent folding of the peripheral subunit-binding domain is thus confirmed and its purification in large-scale amounts for detailed structural analysis is now possible. Images Fig. 1. Fig. 2. Fig. 4. Fig. 5. Fig. 6. PMID:1590756

  11. Toxigenic Escherichia Coli and Childhood Diarrhea

    PubMed Central

    Mundell, Dave H.; Anselmo, Carl R.; Thrupp, Lauri D.; Wishnow, Rodney M.

    1976-01-01

    Stool specimens were examined from 40 children with diarrhea who were under three years of age to determine the incidence of enterotoxigenic Escherichia coli in endemic diarrhea. Heat-labile E. coli enterotoxin was assayed in the very sensitive and reproducible cultured adrenal tumor cell system. Toxigenic E. coli were isolated from only one stool specimen and in this case infection with Shigella dysenteriae was also present. None of the eight classic enteropathogenic E. coli isolates were positive in the adrenal assay. This study suggests that heat-labile enterotoxin-producing E. coli are not an important cause of endemic childhood diarrhea in Southern California. PMID:775792

  12. The effect of lemon, orange and bergamot essential oils and their components on the survival of Campylobacter jejuni, Escherichia coli O157, Listeria monocytogenes, Bacillus cereus and Staphylococcus aureus in vitro and in food systems.

    PubMed

    Fisher, K; Phillips, C A

    2006-12-01

    To investigate the effectiveness of oils and vapours of lemon (Citrus limon), sweet orange (Citrus sinensis) and bergamot (Citrus bergamia) and their components against a number of common foodborne pathogens. The disc diffusion method was used to screen the oils and vapours against Listeria monocytogenes, Staphylococcus aureus, Bacillus cereus, Escherichia coli O157 and Campylobacter jejuni. The survival of each species, demonstrated to be susceptible in the in vitro studies, was tested on cabbage leaf for 60 s by direct contact and on chicken skin for 10 min by direct contact and 24 h by vapour. The results indicate that bergamot was the most inhibitory essential oil (EO) and citral and linalool mimicked its effect (P > 0.001). Citral and linalool vapours produced 6 log reductions in L. monocytogenes, Staph. aureus and B. cereus populations on cabbage leaf after 8-10 h exposure but bergamot vapour exposure, while producing a similar reduction in L. monocytogenes and B. cereus populations, had no effect on Staph. aureus. Bergamot was the most effective of the oils tested and linalool the most effective anti-bacterial component. Gram-positive bacteria were more susceptible than Gram-negative bacteria in vitro, although Camp. jejuni and E. coli O157 were inhibited by bergamot and linalool oils and by linalool vapour. All bacteria tested were less susceptible in food systems than in vitro. Of the Gram-positive bacteria tested Staph. aureus was the least susceptible to both the oils and the components tested. Results suggest the possibility that citrus EOs, particularly bergamot, could be used as a way of combating the growth of common causes of food poisoning.

  13. Na+/H+ antiport activity conferred by Bacillus subtilis tetA(L), a 5' truncation product of tetA(L), and related plasmid genes upon Escherichia coli.

    PubMed Central

    Cheng, J; Baldwin, K; Guffanti, A A; Krulwich, T A

    1996-01-01

    An Escherichia coli transformant expressing the Bacillus subtilis tetA(L) gene from a weak promoter was challenged by growth on medium with low, increasing tetracycline concentrations. Changes in the substrate preference ratios of the TetA(L)-mediated resistances and antiports were examined in view of recent findings suggesting that TetA(L) catalyzes efflux of Na+ in exchange for protons in addition to having the ability to catalyze metal-tetracycline/H+ antiport. After growth of the transformant on 1 microgram or more of tetracycline per ml for 12 to 15 h, the tetA(L) gene in the plasmid was found to be disrupted by an IS10 element 50 bp from the 5' end of the coding sequence. This disrupted recombinant plasmid, pKB1, conferred greater tetracycline resistance and higher levels of membrane metal-tetracycline/proton antiport than the original plasmid, pJTA1, but conferred lower NA+ resistance and Na+/H+ antiport levels than the original plasmid. The results indicate that the 5' end of the gene is necessary for optimal Na+/H+ antiport but that some such activity as well as robust tetracycline/H+ antiport persists in its absence. Two plasmid genes, tet(K) and qacA, were compared with tetA(L) vis-à-vis their abilities to enhance the Na+/H+ antiporter activity of everted vesicles from E. coli transformants. tet(K), which is more closely related to tetA(L), catalyzed 22Na+ uptake by energized vesicles, whereas the less closely related qacA gene did not. PMID:8849239

  14. 21 CFR 866.3255 - Escherichia coli serological reagents.

    Code of Federal Regulations, 2012 CFR

    2012-04-01

    ... 21 Food and Drugs 8 2012-04-01 2012-04-01 false Escherichia coli serological reagents. 866.3255... coli serological reagents. (a) Identification. Escherichia coli serological reagents are devices that consist of antigens and antisera used in serological tests to identify Escherichia coli from cultured...

  15. Clinical Implications of Enteroadherent Escherichia coli

    PubMed Central

    Arenas-Hernández, Margarita M.P.; Martínez-Laguna, Ygnacio; Torres, Alfredo G.

    2012-01-01

    Pathogenic Escherichia coli that colonize the small intestine primarily cause gastrointestinal illness in infants and travelers. The main categories of pathogenic E. coli that colonize the epithelial lining of the small intestine are enterotoxigenic E. coli enteropathogenic E. coli and enteroaggregative E. coli. These organisms accomplish their pathogenic process by a complex, coordinated multistage strategy, including non-intimate adherence mediated by various adhesins. These so called “enteroadherent E. coli ” categories subsequently produced toxins or effector proteins that are either secreted to the milieu or injected to the host cell. Finally, destruction of the intestinal microvilli results from the intimate adherence or the toxic effect exerted over the epithelia, resulting in water secretion and diarrhea. In this review, we summarize the current state of knowledge regarding these enteroadherent E. coli strains and the present clinical understanding of how these organisms colonize the human intestine and cause disease. PMID:22798032

  16. In-stream Escherichia coli Modeling

    NASA Astrophysics Data System (ADS)

    Pandey, P.; Soupir, M.

    2013-12-01

    Elevated levels of pathogenic bacteria indicators such as Escherichia coli (E. coli) in streams are a serious concern. Controlling E. coli levels in streams requires improving our existing understanding of fate and transport of E. coli at watershed scale. In-stream E. coli concentrations are potentially linked to non-point pollution sources (i.e., agricultural land). Water of a natural stream can receive E. coli by either through overland flow (via runoff from cropland) or resuspension from the streambed to the water column. Calculating in-stream total E. coli loads requires estimation of particle attached bacteria as well free floating E. coli transport. Currently water quality models commonly used for predicting E. coli levels in stream water have limited capability for predicting E. coli levels in the water column as well as in the streambed sediment. The challenges in calculating in-stream E. coli levels include difficulties in modeling the complex interactions between sediment particles and E. coli. Here we have developed a watershed scale model (integrated with Soil and Water Assessment Tool (SWAT)), which involves calculation of particle attached E. coli, to predict in-stream E. coli concentrations. The proposed model predicts E. coli levels in streambed bed sediment as well as in the water column. An extensive in-stream E. coli monitoring was carried out to verify the model predictions, and results indicate that the model performed well. The study proposed here will improve understanding on in-stream bacterial contamination, and help improving existing water quality models for predicting pathogenic bacteria levels in ambient water bodies.

  17. Native valve Escherichia coli endocarditis following urosepsis.

    PubMed

    Rangarajan, D; Ramakrishnan, S; Patro, K C; Devaraj, S; Krishnamurthy, V; Kothari, Y; Satyaki, N

    2013-05-01

    Gram-negative organisms are a rare cause of infective endocarditis. Escherichia coli, the most common cause of urinary tract infection and gram-negative septicemia involves endocardium rarely. In this case report, we describe infection of native mitral valve by E. coli following septicemia of urinary tract origin in a diabetic male; subsequently, he required prosthetic tissue valve replacement indicated by persistent sepsis and congestive cardiac failure.

  18. Oral Administration of a Select Mixture of Bacillus Probiotics Affects the Gut Microbiota and Goblet Cell Function following Escherichia coli Challenge in Newly Weaned Pigs of Genotype MUC4 That Are Supposed To Be Enterotoxigenic E. coli F4ab/ac Receptor Negative

    PubMed Central

    Zhang, Wei; Zhou, Dong; Wu, Qiong; Song, Dan; Dicksved, Johan; Wang, Jiu-Feng

    2016-01-01

    ABSTRACT Structural disruption of the gut microbiota and impaired goblet cell function are collateral etiologic factors in enteric diseases. Low, moderate, or high doses of a Bacillus licheniformis-B. subtilis mixture (BLS mix) were orally administered to piglets of genotype MUC4 that are supposed to be F4-expressing enterotoxigenic Escherichia coli strain (F4+ ETEC) F4ab/ac receptor negative (i.e., MUC4-resistant piglets) for 1 week before F4+ ETEC challenge. The luminal contents were collected from the mucosa of the colon on day 8 after F4+ ETEC challenge. The BLS mix attenuated E. coli-induced expansion of Bacteroides uniformis, Eubacterium eligens, Acetanaerobacterium, and Sporobacter populations. Clostridium and Turicibacter populations increased following F4+ ETEC challenge in pigs pretreated with low-dose BLS mix. Lactobacillus gasseri and Lactobacillus salivarius populations increased after administration of BLS mix during E. coli infection. The beneficial effects of BLS mix were due in part to the expansion of certain Clostridium, Lactobacillus, and Turicibacter populations, with a corresponding increase in the number of goblet cells in the ileum via upregulated Atoh1 expression, in turn increasing MUC2 production and thus preserving the mucus barrier and enhancing host defenses against enteropathogenic bacteria. However, excessive BLS mix consumption may increase the risk for enteritis, partly through disruption of colonic microbial ecology, characterized by expansion of Proteobacteria and impaired goblet cell function in the ileum. Our findings suggest that oral administration of BLS mix reprograms the gut microbiota and enhances goblet cell function to ameliorate enteritis. IMPORTANCE The present study is important for improving our understanding of the protective role of probiotics against Escherichia coli infection in piglets. Structural disruption of the gut microbiota and impaired goblet cell function are collateral etiologic factors in enteric

  19. Extraintestinal pathogenic Escherichia coli: "the other bad E coli".

    PubMed

    Johnson, James R; Russo, Thomas A

    2002-03-01

    Extraintestinal pathogenic Escherichia coli (ExPEC), the specialized strains of E coli that cause most extraintestinal E coli infections, represent a major but little-appreciated health threat. Although the reasons for their evolution remain mysterious, by virtue of their numerous virulence traits ExPEC clearly possess a unique ability to cause disease outside the host intestinal tract. Broader appreciation of the existence and importance of ExPEC and better understandings of their distinctive virulence mechanisms, reservoirs, and transmission pathways may lead to effective preventive interventions against the morbid and costly infections ExPEC cause.

  20. Synthesis and processing in Escherichia coli of human leucocyte interferon fused with the signal sequence of Bacillus amyloliquefaciens a-amylase

    SciTech Connect

    Sorokin, A.V.; Avakov, A.S.; Bogush, V.G.; Kostrov, S.V.; Gaiga, G.Z.; Iomantas, Yu.V.; Abalakina, E.G.; Stepanov, A.I.; Strongin, A.Ya.; Kozlov, Yu.I.; Debabov, V.G.

    1985-11-01

    Earlier, the authors reported cloning of the alpha-amylase gene of B. amyloliquefaciens in B. subtilis and E. coli. Currently, the authors report results on the expression of the hybrid gene consisting of the DNA fragment coding for the leader part of B. amyloliquefaciens alpha-amylase and the structural part of the human interferon alpha-2 in E. coli cells. This gene contains an additional methionine codon at the 5'-terminal, which codes for the interferon structure (without its own signal peptide). The interferon gene was inserted into plasmid /sub p/TG 278 at the cleavage site of EcoRI. The structure of the plasmid thus obtained the signal peptide of amylase, five amino acids (Val-Gly-Glu-Phe-Met), and the structural part of the interferon. The E. coli C600 cells carrying plasmid pTGA6 were used to study interferon secretions. The interferon activity was determined radioimmunologically with the use of monoclonal anti-bodies NK2.

  1. Escherichia Coli--Key to Modern Genetics.

    ERIC Educational Resources Information Center

    Bregegere, Francois

    1982-01-01

    Mid-nineteenth century work by Mendel on plant hybrids and by Pasteur on fermentation gave birth by way of bacterial genetics to modern-day molecular biology. The bacterium Escherichia Coli has occupied a key position in genetic studies leading from early gene identification with DNA to current genetic engineering using recombinant DNA technology.…

  2. Detection of O antigens in Escherichia coli

    USDA-ARS?s Scientific Manuscript database

    Lipopolysaccharide on the surface of Escherichia coli constitute the O antigens, which are important virulence factors that are targets of both the innate and adaptive immune system and play a major role in host-pathogen interactions. O antigens that are responsible for antigenic specificity of the ...

  3. Widespread Antisense Transcription in Escherichia coli

    PubMed Central

    Dornenburg, James E.; DeVita, Anne M.; Palumbo, Michael J.; Wade, Joseph T.

    2010-01-01

    ABSTRACT The vast majority of annotated transcripts in bacteria are mRNAs. Here we identify ~1,000 antisense transcripts in the model bacterium Escherichia coli. We propose that these transcripts are generated by promiscuous transcription initiation within genes and that many of them regulate expression of the overlapping gene. PMID:20689751

  4. Escherichia Coli--Key to Modern Genetics.

    ERIC Educational Resources Information Center

    Bregegere, Francois

    1982-01-01

    Mid-nineteenth century work by Mendel on plant hybrids and by Pasteur on fermentation gave birth by way of bacterial genetics to modern-day molecular biology. The bacterium Escherichia Coli has occupied a key position in genetic studies leading from early gene identification with DNA to current genetic engineering using recombinant DNA technology.…

  5. Escherichia coli and Sudden Infant Death Syndrome

    PubMed Central

    Bettelheim, Karl A.; Goldwater, Paul N.

    2015-01-01

    This review examines the association of strains of Escherichia coli with sudden infant death syndrome (SIDS) and the possible role these bacteria play in this enigmatic condition. The review addresses evidence for E. coli in SIDS infants, potential sources of E. coli in the environment, colonization by commensal and pathogenic strains, the variety of currently accepted pathotypes, and how these pathotypes could compromise intestinal integrity and induce inflammation. Both intestinal and extraintestinal pathotypes are compared in relation to the apparent liability in which virulence traits can be gained or lost by strains of E. coli. The way in which E. coli infections fit with current views on infant sleeping position and other SIDS risk factors is highlighted. PMID:26191064

  6. [Changes in fine morphological structures of Escherichia coli, Staphylococcus aureus and spores of Bacillus anthracis vaccine strain STI under the action of the disinfectant "Veltolen"].

    PubMed

    Ivanova, E B; Kurilov, V Ia; Andrus, V N

    2004-01-01

    Changes in fine morphological structures of E. coli, S. aureus and spores of B. anthracis vaccine strain STI under the action of the disinfectant "Veltolen" manufactured by the Closed Joint-Stock Company "Velt", were evaluated. When used at concentrations of 0.0025-0.025%, the preparation induced the loosening of the cell wall in all microorganisms under study, the intensive formation of bubbles on the cell wall surface with their subsequent separation from the cell wall and the formation of "rosettes". In case of more prolonged exposure (up to 60 minutes) and higher concentrations of the preparation these phenomena became more intensive and finally led to the destruction of bacterial cells.

  7. The flexible feedstock concept in Industrial Biotechnology: Metabolic engineering of Escherichia coli, Corynebacterium glutamicum, Pseudomonas, Bacillus and yeast strains for access to alternative carbon sources.

    PubMed

    Wendisch, Volker F; Brito, Luciana Fernandes; Gil Lopez, Marina; Hennig, Guido; Pfeifenschneider, Johannes; Sgobba, Elvira; Veldmann, Kareen H

    2016-09-20

    Most biotechnological processes are based on glucose that is either present in molasses or generated from starch by enzymatic hydrolysis. At the very high, million-ton scale production volumes, for instance for fermentative production of the biofuel ethanol or of commodity chemicals such as organic acids and amino acids, competing uses of carbon sources e.g. in human and animal nutrition have to be taken into account. Thus, the biotechnological production hosts E. coli, C. glutamicum, pseudomonads, bacilli and Baker's yeast used in these large scale processes have been engineered for efficient utilization of alternative carbon sources. This flexible feedstock concept is central to the use of non-glucose second and third generation feedstocks in the emerging bioeconomy. The metabolic engineering efforts to broaden the substrate scope of E. coli, C. glutamicum, pseudomonads, B. subtilis and yeasts to include non-native carbon sources will be reviewed. Strategies to enable simultaneous consumption of mixtures of native and non-native carbon sources present in biomass hydrolysates will be summarized and a perspective on how to further increase feedstock flexibility for the realization of biorefinery processes will be given. Copyright © 2016 Elsevier B.V. All rights reserved.

  8. Electrophoretic Mobilities of Escherichia coli O157:H7 and Wild-Type Escherichia coli Strains

    PubMed Central

    Lytle, Darren A.; Rice, Eugene W.; Johnson, Clifford H.; Fox, Kim R.

    1999-01-01

    The electrophoretic mobilities (EPMs) of a number of Escherichia coli O157:H7 and wild-type E. coli strains were measured. The effects of pH and ionic strength on the EPMs were investigated. The EPMs of E. coli O157:H7 strains differed from those of wild-type strains. As the suspension pH decreased, the EPMs of both types of strains increased. PMID:10388724

  9. Hydrogen production by recombinant Escherichia coli strains

    PubMed Central

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

    2012-01-01

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

  10. Escherichia coli field contamination of pecan nuts.

    PubMed

    Marcus, K A; Amling, H J

    1973-09-01

    More pecan samples collected from grazed orchards were contaminated with Escherichia coli than were samples from nongrazed orchards. No differences in frequency of contamination between mechanically and manually harvested nuts occurred. Nutmeats from whole uncracked pecans that were soaked for 24 h in a lactose broth solution containing E. coli did not become contaminated. Twentyfour percent of the whole pecans soaked in water for 48 h to simulate standing in a rain puddle developed openings along shell suture lines which did not completely close when the nuts were redried.

  11. Uropathogenic Escherichia coli-Associated Exotoxins.

    PubMed

    Welch, Rodney A

    2016-06-01

    Escherichia coli are a common cause of infectious disease outside of the gastrointestinal tract. Several independently evolved E. coli clades are common causes of urinary tract and bloodstream infections. There is ample epidemiological and in vitro evidence that several different protein toxins common to many, but not all, of these strains are likely to aid the colonization and immune-evasion ability of these bacteria. This review discusses our current knowledge and areas of ignorance concerning the contribution of the hemolysin; cytotoxic-necrotizing factor-1; and the autotransporters, Sat, Pic, and Vat, to extraintestinal human disease.

  12. The evolution of the Escherichia coli phylogeny.

    PubMed

    Chaudhuri, Roy R; Henderson, Ian R

    2012-03-01

    Escherichia coli is familiar to biologists as a classical model system, ubiquitous in molecular biology laboratories around the world. Outside of the laboratory, E. coli strains exist as an almost universal component of the lower-gut flora of humans and animals. Although usually a commensal, E. coli has an alter ego as a pathogen, and is associated with diarrhoeal disease and extra-intestinal infections. The study of E. coli diversity predates the availability of molecular data, with strains initially distinguished by serotyping and metabolic profiling, and genomic diversity illustrated by DNA hybridisation. The quantitative study of E. coli diversity began with the application of multi-locus enzyme electrophoresis (MLEE), and has progressed with the accumulation of nucleotide sequence data, from single genes through multi-locus sequence typing (MLST) to whole genome sequencing. Phylogenetic methods have shed light on the processes of genomic evolution in this extraordinarily diverse species, and revealed the origins of pathogenic E. coli strains, including members of the phylogenetically indistinguishable "genus"Shigella. In May and June 2011, an outbreak of haemorrhagic uraemic syndrome in Germany was linked to a strain of enterohaemorrhagic E. coli (EHEC) O104:H4. Application of high-throughput sequencing technologies allowed the genome and origins of the outbreak strain to be characterised in real time as the outbreak was in progress.

  13. Automatic tracking of Escherichia coli bacteria.

    PubMed

    Xie, Jun; Khan, Shahid; Shah, Mubarak

    2008-01-01

    In this paper, we present an automatic method for estimating the trajectories of Escherichia coli bacteria from in vivo phase-contrast microscopy videos. To address the low-contrast boundaries in cellular images, an adaptive kernel-based technique is applied to detect cells in sequence of frames. Then a novel matching gain measure is introduced to cope with the challenges such as dramatic changes of cells' appearance and serious overlapping and occlusion. For multiple cell tracking, an optimal matching strategy is proposed to improve the handling of cell collision and broken trajectories. The results of successful tracking of Escherichia coli from various phase-contrast sequences are reported and compared with manually-determined trajectories, as well as those obtained from existing tracking methods. The stability of the algorithm with different parameter values is also analyzed and discussed.

  14. ELECTRON MICROSCOPY OF PLASMOLYSIS IN ESCHERICHIA COLI.

    PubMed

    COTA-ROBLES, E H

    1963-03-01

    Cota-Robles, Eugene H. (University of California, Riverside). Electron microscopy of plasmolysis in Escherichia coli. J. Bacteriol. 85:499-503. 1963.-Escherichia coli cells plasmolyzed in 0.35 m sucrose reveal plasmolysis at one tip of a cell or in the center of dividing cells in which protoplast partition has been complete. Central plasmolysis reveals that protoplast separation can be completed before the invagination of the cell wall is complete. These studies support the concept that these cells divide by constriction. The strength of the union between cell wall and cytoplasm is not uniform around the entire cell. It is strongest along the sides of these rod-shaped cells and weakest at one tip of the single cell. Thus, a single cell generally forms one cup-shaped vacuole in which the cytoplasm has collapsed away from one tip of the cell.

  15. ELECTRON MICROSCOPY OF PLASMOLYSIS IN ESCHERICHIA COLI

    PubMed Central

    Cota-Robles, Eugene H.

    1963-01-01

    Cota-Robles, Eugene H. (University of California, Riverside). Electron microscopy of plasmolysis in Escherichia coli. J. Bacteriol. 85:499–503. 1963.—Escherichia coli cells plasmolyzed in 0.35 m sucrose reveal plasmolysis at one tip of a cell or in the center of dividing cells in which protoplast partition has been complete. Central plasmolysis reveals that protoplast separation can be completed before the invagination of the cell wall is complete. These studies support the concept that these cells divide by constriction. The strength of the union between cell wall and cytoplasm is not uniform around the entire cell. It is strongest along the sides of these rod-shaped cells and weakest at one tip of the single cell. Thus, a single cell generally forms one cup-shaped vacuole in which the cytoplasm has collapsed away from one tip of the cell. Images PMID:14042923

  16. Topical Application of Escherichia coli-Vectored Vaccine as a Simple Method for Eliciting Protective Immunity

    PubMed Central

    Zhang, Jianfeng; Shi, Zhongkai; Kong, Fan-kun; Jex, Edward; Huang, Zhigang; Watt, James M.; Van Kampen, Kent R.; Tang, De-chu C.

    2006-01-01

    We report here that animals can be protected against lethal infection by Clostridium tetani cells and Bacillus anthracis spores following topical application of intact particles of live or γ-irradiated Escherichia coli vectors overproducing tetanus and anthrax antigens, respectively. Cutaneous γδT cells were rapidly recruited to the administration site. Live E. coli cells were not found in nonskin tissues after topical application, although fragments of E. coli DNA were disseminated transiently. Evidence suggested that intact E. coli particles in the outer layer of skin may be disrupted by a γδT-cell-mediated innate defense mechanism, followed by the presentation of E. coli ligand-adjuvanted intravector antigens to the immune system and rapid degradation of E. coli components. The nonreplicating E. coli vector overproducing an exogenous immunogen may foster the development of a new generation of vaccines that can be manufactured rapidly and administered noninvasively in a wide variety of disease settings. PMID:16714593

  17. Phage therapy: the Escherichia coli experience.

    PubMed

    Brüssow, Harald

    2005-07-01

    Phages have been proposed as natural antimicrobial agents to fight bacterial infections in humans, in animals or in crops of agricultural importance. Phages have also been discussed as hygiene measures in food production facilities and hospitals. These proposals have a long history, but are currently going through a kind of renaissance as documented by a spate of recent reviews. This review discusses the potential of phage therapy with a specific example, namely Escherichia coli.

  18. Oral Administration of a Select Mixture of Bacillus Probiotics Affects the Gut Microbiota and Goblet Cell Function following Escherichia coli Challenge in Newly Weaned Pigs of Genotype MUC4 That Are Supposed To Be Enterotoxigenic E. coli F4ab/ac Receptor Negative.

    PubMed

    Zhang, Wei; Zhu, Yao-Hong; Zhou, Dong; Wu, Qiong; Song, Dan; Dicksved, Johan; Wang, Jiu-Feng

    2017-02-01

    Structural disruption of the gut microbiota and impaired goblet cell function are collateral etiologic factors in enteric diseases. Low, moderate, or high doses of a Bacillus licheniformis-B. subtilis mixture (BLS mix) were orally administered to piglets of genotype MUC4 that are supposed to be F4-expressing enterotoxigenic Escherichia coli strain (F4(+) ETEC) F4ab/ac receptor negative (i.e., MUC4-resistant piglets) for 1 week before F4(+) ETEC challenge. The luminal contents were collected from the mucosa of the colon on day 8 after F4(+) ETEC challenge. The BLS mix attenuated E. coli-induced expansion of Bacteroides uniformis, Eubacterium eligens, Acetanaerobacterium, and Sporobacter populations. Clostridium and Turicibacter populations increased following F4(+) ETEC challenge in pigs pretreated with low-dose BLS mix. Lactobacillus gasseri and Lactobacillus salivarius populations increased after administration of BLS mix during E. coli infection. The beneficial effects of BLS mix were due in part to the expansion of certain Clostridium, Lactobacillus, and Turicibacter populations, with a corresponding increase in the number of goblet cells in the ileum via upregulated Atoh1 expression, in turn increasing MUC2 production and thus preserving the mucus barrier and enhancing host defenses against enteropathogenic bacteria. However, excessive BLS mix consumption may increase the risk for enteritis, partly through disruption of colonic microbial ecology, characterized by expansion of Proteobacteria and impaired goblet cell function in the ileum. Our findings suggest that oral administration of BLS mix reprograms the gut microbiota and enhances goblet cell function to ameliorate enteritis.

  19. 77 FR 9888 - Shiga Toxin-Producing Escherichia coli

    Federal Register 2010, 2011, 2012, 2013, 2014

    2012-02-21

    ... program for the six non-O157 STEC, as it already does for E. coli O157:H7. The Agency intended to begin... Food Safety and Inspection Service Shiga Toxin-Producing Escherichia coli in Certain Raw Beef Products... manufacturing trimmings for six non-O157 Shiga toxin-producing Escherichia coli (STEC) serogroups (O26, O45...

  20. Systems Metabolic Engineering of Escherichia coli.

    PubMed

    Choi, Kyeong Rok; Shin, Jae Ho; Cho, Jae Sung; Yang, Dongsoo; Lee, Sang Yup

    2017-03-01

    Systems metabolic engineering, which recently emerged as metabolic engineering integrated with systems biology, synthetic biology, and evolutionary engineering, allows engineering of microorganisms on a systemic level for the production of valuable chemicals far beyond its native capabilities. Here, we review the strategies for systems metabolic engineering and particularly its applications in Escherichia coli. First, we cover the various tools developed for genetic manipulation in E. coli to increase the production titers of desired chemicals. Next, we detail the strategies for systems metabolic engineering in E. coli, covering the engineering of the native metabolism, the expansion of metabolism with synthetic pathways, and the process engineering aspects undertaken to achieve higher production titers of desired chemicals. Finally, we examine a couple of notable products as case studies produced in E. coli strains developed by systems metabolic engineering. The large portfolio of chemical products successfully produced by engineered E. coli listed here demonstrates the sheer capacity of what can be envisioned and achieved with respect to microbial production of chemicals. Systems metabolic engineering is no longer in its infancy; it is now widely employed and is also positioned to further embrace next-generation interdisciplinary principles and innovation for its upgrade. Systems metabolic engineering will play increasingly important roles in developing industrial strains including E. coli that are capable of efficiently producing natural and nonnatural chemicals and materials from renewable nonfood biomass.

  1. Systems Metabolic Engineering of Escherichia coli.

    PubMed

    Choi, Kyeong Rok; Shin, Jae Ho; Cho, Jae Sung; Yang, Dongsoo; Lee, Sang Yup

    2016-05-01

    Systems metabolic engineering, which recently emerged as metabolic engineering integrated with systems biology, synthetic biology, and evolutionary engineering, allows engineering of microorganisms on a systemic level for the production of valuable chemicals far beyond its native capabilities. Here, we review the strategies for systems metabolic engineering and particularly its applications in Escherichia coli. First, we cover the various tools developed for genetic manipulation in E. coli to increase the production titers of desired chemicals. Next, we detail the strategies for systems metabolic engineering in E. coli, covering the engineering of the native metabolism, the expansion of metabolism with synthetic pathways, and the process engineering aspects undertaken to achieve higher production titers of desired chemicals. Finally, we examine a couple of notable products as case studies produced in E. coli strains developed by systems metabolic engineering. The large portfolio of chemical products successfully produced by engineered E. coli listed here demonstrates the sheer capacity of what can be envisioned and achieved with respect to microbial production of chemicals. Systems metabolic engineering is no longer in its infancy; it is now widely employed and is also positioned to further embrace next-generation interdisciplinary principles and innovation for its upgrade. Systems metabolic engineering will play increasingly important roles in developing industrial strains including E. coli that are capable of efficiently producing natural and nonnatural chemicals and materials from renewable nonfood biomass.

  2. DNA-damaging activity of patulin in Escherichia coli.

    PubMed Central

    Lee, K S; Röschenthaler, R J

    1986-01-01

    At a concentration of 10 micrograms/ml, patulin caused single-strand DNA breaks in living cells of Escherichia coli. At 50 micrograms/ml, double-strand breaks were observed also. Single-strand breaks were repaired in the presence of 10 micrograms of patulin per ml within 90 min when the cells were incubated at 37 degrees C in M9-salts solution without a carbon source. The same concentration also induced temperature-sensitive lambda prophage and a prophage of Bacillus megaterium. When an in vitro system with permeabilized Escherichia coli cells was used, patulin at 10 micrograms/ml induced DNA repair synthesis and inhibited DNA replication. The in vivo occurrence of DNA strand breaks and DNA repair correlated with the in vitro induction of repair synthesis. In vitro the RNA synthesis was less affected, and overall protein synthesis was not inhibited at 10 micrograms/ml. Only at higher concentrations (250 to 500 micrograms/ml) was inhibition of in vitro protein synthesis observed. Thus, patulin must be regarded as a mycotoxin with selective DNA-damaging activity. PMID:2431653

  3. Diversity of CRISPR loci in Escherichia coli.

    PubMed

    Díez-Villaseñor, C; Almendros, C; García-Martínez, J; Mojica, F J M

    2010-05-01

    CRISPR (clustered regularly interspaced short palindromic repeats) and CAS (CRISPR-associated sequence) proteins are constituents of a novel genetic barrier that limits horizontal gene transfer in prokaryotes by means of an uncharacterized mechanism. The fundamental discovery of small RNAs as the guides of the defence apparatus arose as a result of Escherichia coli studies. However, a survey of the system diversity in this species in order to further contribute to the understanding of the CRISPR mode of action has not yet been performed. Here we describe two CRISPR/CAS systems found in E. coli, following the analysis of 100 strains representative of the species' diversity. Our results substantiate different levels of activity between loci of both CRISPR types, as well as different target preferences and CRISPR relevances for particular groups of strains. Interestingly, the data suggest that the degeneration of one CRISPR/CAS system in E. coli ancestors could have been brought about by self-interference.

  4. Thymineless death in Escherichia coli: strain specificity.

    PubMed

    Cummings, D J; Mondale, L

    1967-06-01

    Thymineless death of various ultraviolet (UV)-sensitive strains of Escherichia coli B and K-12 was investigated. It was found that E. coli B, B(s-12), K-12 rec-21, and possibly K-12 Lon(-), all sensitive to UV, were also sensitive to thymine starvation. However, other UV-sensitive strains of E. coli were found to display the typical resistant-type kinetics of thymineless death. The correlation of these results with various other cellular processes suggested that the filament-forming ability of the bacteria might be involved in the mechanism of thymineless death. It was apparent from the present results that capacity for host-cell reactivation, recombination ability, thymine dimer excision, and probably induction of a defective prophage had little to do with determining sensitivity to thymine deprivation.

  5. Thymineless Death in Escherichia coli: Strain Specificity

    PubMed Central

    Cummings, Donald J.; Mondale, Lee

    1967-01-01

    Thymineless death of various ultraviolet (UV)-sensitive strains of Escherichia coli B and K-12 was investigated. It was found that E. coli B, Bs−12, K-12 rec-21, and possibly K-12 Lon−, all sensitive to UV, were also sensitive to thymine starvation. However, other UV-sensitive strains of E. coli were found to display the typical resistant-type kinetics of thymineless death. The correlation of these results with various other cellular processes suggested that the filament-forming ability of the bacteria might be involved in the mechanism of thymineless death. It was apparent from the present results that capacity for host-cell reactivation, recombination ability, thymine dimer excision, and probably induction of a defective prophage had little to do with determining sensitivity to thymine deprivation. Images PMID:5337772

  6. Interaction between Escherichia coli and lunar fines

    NASA Technical Reports Server (NTRS)

    Johansson, K. R.

    1983-01-01

    A sample of mature lunar fines (10084.151) was solubilized to a high degree (about 17 percent) by the chelating agent salicylic acid (0.01. M). The neutralized (pH adjusted to 7.0) leachate was found to inhibit the growth of Escherichia coli (ATCC 259922) in a minimial mineral salts glucose medium; however, the inhibition was somewhat less than that caused by neutralized salicylic acid alone. The presence of lunar fines in the minimal medium was highly stimulatory to growth of E. coli following an early inhibitory response. The bacterium survived less well in the lunar leachate than in distilled water, no doubt because of the salicylate. It was concluded that the sample of lunar soil tested has nutritional value to E. coli and that certain products of fermentation helped to solubilize the lunar soil.

  7. Escherichia coli O 27 in adult diarrhoea.

    PubMed Central

    Hobbs, B. C.; Rowe, B.; Kendall, M.; Turnbull, P. C.; Ghosh, A. C.

    1976-01-01

    Escherichia coli O 27 H 7 was found in 16 stool samples submitted during a Caribbean cruise (Cruise Z) by 29 patients reporting with diarrhoea. A retrospective search revealed E. coli O 27 H 7 in 11 of 20 and 2 of 14 stool cultures from patients on two previous cruises (Y and X respectively) and in a culture from fresh cream (Cruise Y). The repeated occurrence of E. coli O 27 H 7 in the absence of any other apparent cause suggested that this serotype may have been responsible for the diarrhoea. The results of pathogenicity tests suggested that this strain elaborated heat-stable (ST) enterotoxin. The possibility that food may have been the vector is discussed. PMID:794406

  8. Frequency-Dependent Escherichia coli Chemotaxis Behavior

    NASA Astrophysics Data System (ADS)

    Zhu, Xuejun; Si, Guangwei; Deng, Nianpei; Ouyang, Qi; Wu, Tailin; He, Zhuoran; Jiang, Lili; Luo, Chunxiong; Tu, Yuhai

    2012-03-01

    We study Escherichia coli chemotaxis behavior in environments with spatially and temporally varying attractant sources by developing a unique microfluidic system. Our measurements reveal a frequency-dependent chemotaxis behavior. At low frequency, the E. coli population oscillates in synchrony with the attractant. In contrast, in fast-changing environments, the population response becomes smaller and out of phase with the attractant waveform. These observations are inconsistent with the well-known Keller-Segel chemotaxis equation. A new continuum model is proposed to describe the population level behavior of E. coli chemotaxis based on the underlying pathway dynamics. With the inclusion of a finite adaptation time and an attractant consumption rate, our model successfully explains the microfluidic experiments at different stimulus frequencies.

  9. Production of curcuminoids in engineered Escherichia coli.

    PubMed

    Kim, Eun Ji; Cha, Mi Na; Kim, Bog-Gyu; Ahn, Joong-Hoon

    2017-03-09

    Curcumin, a hydrophobic polyphenol derived from the rhizome of the herb Curcuma longa, possesses diverse pharmacological properties including anti-inflammatory, antioxidant, antiproliferative and antiangiogenic activity. Two curcuminoids (dicinnamoylmethane and bisdemethoxycurcumin) were synthesized from glucose in Escherichia coli. PAL (phenylalanine ammonia lyase) or TAL (tyrosine ammonia lyase), along with Os4CL (p-coumaroyl-CoA ligase) and CUS (curcumin synthase), were introduced in to E. coli, and each strain produced dicinnamoylmethane or bisdemethoxycurcumin, respectively. In order to increase the production of curcuminoids in E. coli, the shikimic acid biosynthesis pathway which increases the substrates for curcuminoid biosynthesis, was engineered. Using engineered strains, the production of bisdemethoxycurcumin increased from 0.32 to 4.63 mg/L, and that of dicinnamoylmethane from 1.24 mg/L and 6.95 mg/L.

  10. Prodigiosin - A Multifaceted Escherichia coli Antimicrobial Agent

    PubMed Central

    Zorec, Maša; Stopar, David

    2016-01-01

    Despite a considerable interest in prodigiosin, the mechanism of its antibacterial activity is still poorly understood. In this work, Escherichia coli cells were treated with prodigiosin to determine its antimicrobial effect on bacterial physiology. The effect of prodigiosin was concentration dependent. In prodigiosin treated cells above MIC value no significant DNA damage or cytoplasmic membrane disintegration was observed. The outer membrane, however, becomes leaky. Cells had severely decreased respiration activity. In prodigiosin treated cells protein and RNA synthesis were inhibited, cells were elongated but could not divide. Pre-treatment with prodigiosin improved E. coli survival rate in media containing ampicillin, kanamycin and erythromycin but not phleomycin. The results suggest that prodigiosin acts as a bacteriostatic agent in E. coli cells. If prodigiosin was diluted, cells resumed growth. The results indicate that prodigiosin has distinct mode of antibacterial action in different bacteria. PMID:27612193

  11. A new Escherichia coli cell division gene, ftsK.

    PubMed Central

    Begg, K J; Dewar, S J; Donachie, W D

    1995-01-01

    A mutation in a newly discovered Escherichia coli cell division gene, ftsK, causes a temperature-sensitive late-stage block in division but does not affect chromosome replication or segregation. This defect is specifically suppressed by deletion of dacA, coding for the peptidoglycan DD-carboxypeptidase, PBP 5. FtsK is a large polypeptide (147 kDa) consisting of an N-terminal domain with several predicted membrane-spanning regions, a proline-glutamine-rich domain, and a C-terminal domain with a nucleotide-binding consensus sequence. FtsK has extensive sequence identity with a family of proteins from a wide variety of prokaryotes and plasmids. The plasmid proteins are required for intercellular DNA transfer, and one of the bacterial proteins (the SpoIIIE protein of Bacillus subtilis) has also been implicated in intracellular chromosomal DNA transfer. PMID:7592387

  12. Inactivation of Escherichia coli by citral.

    PubMed

    Somolinos, M; García, D; Condón, S; Mackey, B; Pagán, R

    2010-06-01

    The aim was to evaluate (i) the resistance of Escherichia coli BJ4 to citral in a buffer system as a function of citral concentration, treatment medium pH, storage time and initial inoculum size, (ii) the role of the sigma factor RpoS on citral resistance of E. coli, (iii) the role of the cell envelope damage in the mechanism of microbial inactivation by citral and (iiii) possible synergistic effects of mild heat treatment and pulsed electric fields (PEF) treatment combined with citral. The initial inoculum size greatly affected the efficacy of citral against E. coli cells. Exposure to 200 microl l(-1) of citral at pH 4.0 for 24 h at 20 degrees C caused the inactivation of more than 5 log(10) cycles of cells starting at an inoculum size of 10(6) or 10(7) CFU ml(-1), whereas increasing the cell concentration to 10(9) CFU ml(-1) caused <1 log(10) cycle of inactivation. Escherichia coli showed higher resistance to citral at pH 4.0 than pH 7.0. The rpoS null mutant strain E. coli BJ4L1 was less resistant to citral than the wild-type strain. Occurrence of sublethal injury to both the cytoplasmic and outer membranes was demonstrated by adding sodium chloride or bile salts to the recovery media. The majority of sublethally injured cells by citral required energy and lipid synthesis for repair. A strongly synergistic lethal effect was shown by mild heat treatment combined with citral but the presence of citral during the application of a PEF treatment did not show any advantage. This work confirms that cell envelope damage is an important event in citral inactivation of bacteria, and it describes the key factors on the inactivation of E. coli cells by citral. Knowledge about the mechanism of microbial inactivation by citral helps establish successful combined preservation treatments.

  13. Cation Transport in Escherichia coli

    PubMed Central

    Schultz, Stanley G.; Solomon, A. K.

    1961-01-01

    Methods have been developed to study the intracellular Na and K concentrations in E. coli, strain K-12. These intracellular cation concentrations have been shown to be functions of the extracellular cation concentrations and the age of the bacterial culture. During the early logarithmic phase of growth, the intracellular K concentration greatly exceeds that of the external medium, whereas the intracellular Na concentration is lower than that of the growth medium. As the age of the culture increases, the intracellular K concentration falls and the intracellular Na concentration rises, changes which are related to the fall in the pH of the medium and to the accumulation of the products of bacterial metabolism. When stationary phase cells, which are rich in Na and poor in K, are resuspended in fresh growth medium, there is a rapid reaccumulation of K and extrusion of Na. These processes represent oppositely directed net ion movements against concentration gradients, and have been shown to be dependent upon the presence of an intact metabolic energy supply. PMID:13909521

  14. Cation Transport in Escherichia coli

    PubMed Central

    Schultz, Stanley G.; Epstein, Wolfgang; Solomon, A. K.

    1963-01-01

    The resuspension of K-poor, Na-rich stationary phase E. coli in fresh medium at pH 7.0 results in a rapid uptake of K and extrusion of Na by the cells. In all experiments net K uptake exceeded net Na extrusion. An investigation of the uptake of glucose, PO4, and Mg and the secretion of H by these cells indicates that the excess K uptake is not balanced by the simultaneous uptake of anions but must be accompanied by the extrusion of cations from the cell. The kinetics of net K uptake are consistent with the existence of two parallel influx processes. The first is rapid, of brief duration, and accounts for approximately 60 per cent of the total net K uptake. This process is a function of the extracellular K concentration, is inhibited in acid media, and appears to be a 1 for 1 exchange of extracellular K for intracellular H. The second influx process has a half-time of approximately 12 minutes, and is not affected by acid media. This process is a function of the intracellular Na concentration, is dependent upon the presence of K in the medium, and may be ascribed to a 1 for 1 exchange of extracellular K for intracellular Na. PMID:14080819

  15. Biodegradation of aromatic compounds by Escherichia coli.

    PubMed

    Díaz, E; Ferrández, A; Prieto, M A; García, J L

    2001-12-01

    Although Escherichia coli has long been recognized as the best-understood living organism, little was known about its abilities to use aromatic compounds as sole carbon and energy sources. This review gives an extensive overview of the current knowledge of the catabolism of aromatic compounds by E. coli. After giving a general overview of the aromatic compounds that E. coli strains encounter and mineralize in the different habitats that they colonize, we provide an up-to-date status report on the genes and proteins involved in the catabolism of such compounds, namely, several aromatic acids (phenylacetic acid, 3- and 4-hydroxyphenylacetic acid, phenylpropionic acid, 3-hydroxyphenylpropionic acid, and 3-hydroxycinnamic acid) and amines (phenylethylamine, tyramine, and dopamine). Other enzymatic activities acting on aromatic compounds in E. coli are also reviewed and evaluated. The review also reflects the present impact of genomic research and how the analysis of the whole E. coli genome reveals novel aromatic catabolic functions. Moreover, evolutionary considerations derived from sequence comparisons between the aromatic catabolic clusters of E. coli and homologous clusters from an increasing number of bacteria are also discussed. The recent progress in the understanding of the fundamentals that govern the degradation of aromatic compounds in E. coli makes this bacterium a very useful model system to decipher biochemical, genetic, evolutionary, and ecological aspects of the catabolism of such compounds. In the last part of the review, we discuss strategies and concepts to metabolically engineer E. coli to suit specific needs for biodegradation and biotransformation of aromatics and we provide several examples based on selected studies. Finally, conclusions derived from this review may serve as a lead for future research and applications.

  16. Biodegradation of Aromatic Compounds by Escherichia coli

    PubMed Central

    Díaz, Eduardo; Ferrández, Abel; Prieto, María A.; García, José L.

    2001-01-01

    Although Escherichia coli has long been recognized as the best-understood living organism, little was known about its abilities to use aromatic compounds as sole carbon and energy sources. This review gives an extensive overview of the current knowledge of the catabolism of aromatic compounds by E. coli. After giving a general overview of the aromatic compounds that E. coli strains encounter and mineralize in the different habitats that they colonize, we provide an up-to-date status report on the genes and proteins involved in the catabolism of such compounds, namely, several aromatic acids (phenylacetic acid, 3- and 4-hydroxyphenylacetic acid, phenylpropionic acid, 3-hydroxyphenylpropionic acid, and 3-hydroxycinnamic acid) and amines (phenylethylamine, tyramine, and dopamine). Other enzymatic activities acting on aromatic compounds in E. coli are also reviewed and evaluated. The review also reflects the present impact of genomic research and how the analysis of the whole E. coli genome reveals novel aromatic catabolic functions. Moreover, evolutionary considerations derived from sequence comparisons between the aromatic catabolic clusters of E. coli and homologous clusters from an increasing number of bacteria are also discussed. The recent progress in the understanding of the fundamentals that govern the degradation of aromatic compounds in E. coli makes this bacterium a very useful model system to decipher biochemical, genetic, evolutionary, and ecological aspects of the catabolism of such compounds. In the last part of the review, we discuss strategies and concepts to metabolically engineer E. coli to suit specific needs for biodegradation and biotransformation of aromatics and we provide several examples based on selected studies. Finally, conclusions derived from this review may serve as a lead for future research and applications. PMID:11729263

  17. Profiling of Escherichia coli Chromosome database.

    PubMed

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

    2008-01-01

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

  18. Current Interventions for Controlling Pathogenic Escherichia coli.

    PubMed

    Kim, Nam Hee; Cho, Tae Jin; Rhee, Min Suk

    2017-01-01

    This review examined scientific reports and articles published from 2007 to 2016 regarding the major environmental sources of pathogenic Escherichia coli and the routes by which they enter the human gastrointestinal tract. The literature describes novel techniques used to combat pathogenic E. coli transmitted to humans from livestock and agricultural products, food-contact surfaces in processing environments, and food products themselves. Although prevention before contamination is always the best "intervention," many studies aim to identify novel chemical, physical, and biological techniques that inactivate or eliminate pathogenic E. coli cells from breeding livestock, growing crops, and manufactured food products. Such intervention strategies target each stage of the food chain from the perspective of "Farm to Table food safety" and aim to manage major reservoirs of pathogenic E. coli throughout the entire process. Issues related to, and recent trends in, food production must address not only the safety of the food itself but also the safety of those who consume it. Thus, research aims to discover new "natural" antimicrobial agents and to develop "multiple hurdle technology" or other novel technologies that preserve food quality. In addition, this review examines the practical application of recent technologies from the perspective of product quality and safety. It provides comprehensive insight into intervention measures used to ensure food safety, specifically those aimed at pathogenic E. coli. Copyright © 2017 Elsevier Inc. All rights reserved.

  19. The 503nm pigment of Escherichia coli

    PubMed Central

    Kamitakahara, Joyce R.; Polglase, W. J.

    1970-01-01

    The yield of cell protein was one-third less for streptomycin-dependent Escherichia coli B than for the wild-type parent strain when both were grown aerobically on a medium with limiting glucose, but anaerobically the yield of protein was similar for both strains. The transient pigment absorbing at 503nm that is known to be present in E. coli and other organisms was not detectable in streptomycin-dependent mutants nor in a non-dependent (energy-deficient) revertant. When wild-type E. coli B was grown on limiting glucose–salts medium containing 2,4 dinitrophenol, the yield of cell protein was decreased and formation of the 503nm pigment was inhibited. Fumarase, aconitase and glucose 6-phosphate dehydrogenase were de-repressed in E. coli B cells grown with excess of glucose in a medium containing 2,4-dinitrophenol. In air-oxidized, wild-type E. coli B cells, the 503nm pigment appeared before reduced cytochromes when gluconate was the substrate but failed to appear when succinate was the substrate. The results provide evidence for a role of the 503nm pigment in aerobic energy metabolism, possibly as an electron acceptor from NADPH. PMID:4395501

  20. Intramammary challenge with Escherichia coli following immunization with a curli-producing Escherichia coli.

    PubMed

    Todhunter, D A; Smith, K L; Hogan, J S; Nelson, L

    1991-03-01

    Holstein and Jersey cattle were immunized with a curli-producing strain of Escherichia coli (pCRL65/A012) or a noncurli-producing strain (pUC18/HB101) to determine differences in resistance to establishment of experimental intramammary infection. Cows (n = 6 per group) were immunized at 14 d prior to drying off, 7 d of involution, and at calving with 3 x 10(10) E. coli in Freund's Incomplete Adjuvant. At 30 d of lactation, one mammary quarter of each cow was infused with a wild strain of E. coli (727). Escherichia coli 727 was isolated from a naturally occurring intramammary infection and produced curli. All challenged quarters became infected, and all cows developed acute clinical mastitis. Geometric mean duration of intramammary infections was 6 d for both immunization groups. All infections were spontaneously eliminated within 10 d. No differences occurred between immunization groups in blood selenium and glutathione peroxidase activity, plasma selenium, number of E. coli 727 isolated from secretion after challenge, rectal temperature and SCC response, clinical status of mammary quarters, or DMI. Reduction in milk production after challenge was greater for cows immunized with E. coli pCRL65/A012. Immunization of dairy cattle with a curli-producing strain of E. coli did not protect against experimental intramammary challenge during lactation.

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

    EPA Science Inventory

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

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

    EPA Science Inventory

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

  3. Virulence factors of uropathogenic Escherichia coli.

    PubMed

    Emody, L; Kerényi, M; Nagy, G

    2003-10-01

    Virulence factors of Escherichia coli are of two main types; those produced on the surface of the cell and those produced within the cell and then exported to the site of action. Those on the surface include different sorts of fimbriae that have a role in adhesion to the surface of host cells but may also have additional roles such as tissue invasion, biofilm formation or cytokine induction. The activities of cell wall components are discussed and several exported virulence factors are described that have anti host cell activities. Others virulence factors enable the bacteria to grow in an environment of iron restriction.

  4. Acid tolerance of enterohemorrhagic Escherichia coli.

    PubMed Central

    Benjamin, M M; Datta, A R

    1995-01-01

    Enterohemorrhagic Escherichia coli (EHEC) strains were tested for their ability to survive in acid pH at 37 degrees C. No loss of viability was observed in an O157:H7 EHEC strain (ATCC 43895) at pH levels of 3.0 and 2.5 for at least 5 h. The level of acid tolerance of most EHEC isolates was very high, similar to that of Shigella flexneri strains. The acid tolerance was dependent on the growth phase and pH of the growth medium. PMID:7747983

  5. Detection of Escherichia coli enterotoxins in stools.

    PubMed Central

    Merson, M H; Yolken, R H; Sack, R B; Froehlich, J L; Greenberg, H B; Huq, I; Black, R W

    1980-01-01

    We determined whether enterotoxigenic Escherichia coli diarrhea could be diagnosed by direct examination of stools for heat-labile (LT) and heat-stable (ST) enterotoxins. The Y-1 adrenal cell and an enzyme-linked immunosorbent assay (ELISA) detected LT in 85 and 93%, respectively, of stool specimens obtained from adults with acute diarrhea from whom an LT- and ST-producing organism had been isolated. Furthermore, the ELISA assay detected LT in 8 of 35 stool specimens from which no LT-producing E. coli had been isolated. The infant mouse assay was utilized to detect ST in these stool specimens and was found to be an insensitive method, showing positive results in only 36% of the specimens from which an ST-producing organism was isolated. Further studies are warranted to determine the diagnostic value of direct detection of LT in stools, especially by the ELISA method. PMID:6995331

  6. Production of recombinant avidin in Escherichia coli.

    PubMed

    Airenne, K J; Sarkkinen, P; Punnonen, E L; Kulomaa, M S

    1994-06-24

    A recombinant avidin (re-Avd), containing amino acids (aa) 1-123 of the native chicken egg-white Avd, was produced in Escherichia coli. When cells were grown at 37 degrees C production was over 1 microgram/ml, due to altering the codon preference of the first ten codons. The re-Avd was recovered as a soluble protein from cells grown at 25 or 30 degrees C, whereas at 37 degrees C it was mostly insoluble in inclusion bodies. Our results indicated that, despite the potentially harmful biotin-binding activity of Avd, it is possible to produce biologically active Avd in E. coli which then can easily be purified by affinity chromatography on a biotin column in a single step.

  7. Engineering ethanologenic Escherichia coli for levoglucosan utilization.

    PubMed

    Layton, Donovan S; Ajjarapu, Avanthi; Choi, Dong Won; Jarboe, Laura R

    2011-09-01

    Levoglucosan is a major product of biomass pyrolysis. While this pyrolyzed biomass, also known as bio-oil, contains sugars that are an attractive fermentation substrate, commonly-used biocatalysts, such as Escherichia coli, lack the ability to metabolize this anhydrosugar. It has previously been shown that recombinant expression of the levoglucosan kinase enzyme enables use of levoglucosan as carbon and energy source. Here, ethanologenic E. coli KO11 was engineered for levoglucosan utilization by recombinant expression of levoglucosan kinase from Lipomyces starkeyi. Our engineering strategy uses a codon-optimized gene that has been chromosomally integrated within the pyruvate to ethanol (PET) operon and does not require additional antibiotics or inducers. Not only does this engineered strain use levoglucosan as sole carbon source, but it also ferments levoglucosan to ethanol. This work demonstrates that existing biocatalysts can be easily modified for levoglucosan utilization. Copyright © 2011 Elsevier Ltd. All rights reserved.

  8. Designed phosphoprotein recognition in Escherichia coli.

    PubMed

    Sawyer, Nicholas; Gassaway, Brandon M; Haimovich, Adrian D; Isaacs, Farren J; Rinehart, Jesse; Regan, Lynne

    2014-11-21

    Protein phosphorylation is a central biological mechanism for cellular adaptation to environmental changes. Dysregulation of phosphorylation signaling is implicated in a wide variety of diseases. Thus, the ability to detect and quantify protein phosphorylation is highly desirable for both diagnostic and research applications. Here we present a general strategy for detecting phosphopeptide-protein interactions in Escherichia coli. We first redesign a model tetratricopeptide repeat (TPR) protein to recognize phosphoserine in a sequence-specific fashion and characterize the interaction with its target phosphopeptide in vitro. We then combine in vivo site-specific incorporation of phosphoserine with split mCherry assembly to observe the designed phosphopeptide-protein interaction specificity in E. coli. This in vivo strategy for detecting and characterizing phosphopeptide-protein interactions has numerous potential applications for the study of natural interactions and the design of novel ones.

  9. Engineering the Escherichia coli Fermentative Metabolism

    NASA Astrophysics Data System (ADS)

    Orencio-Trejo, M.; Utrilla, J.; Fernández-Sandoval, M. T.; Huerta-Beristain, G.; Gosset, G.; Martinez, A.

    Fermentative metabolism constitutes a fundamental cellular capacity for industrial biocatalysis. Escherichia coli is an important microorganism in the field of metabolic engineering for its well-known molecular characteristics and its rapid growth. It can adapt to different growth conditions and is able to grow in the presence or absence of oxygen. Through the use of metabolic pathway engineering and bioprocessing techniques, it is possible to explore the fundamental cellular properties and to exploit its capacity to be applied as industrial biocatalysts to produce a wide array of chemicals. The objective of this chapter is to review the metabolic engineering efforts carried out with E. coli by manipulating the central carbon metabolism and fermentative pathways to obtain strains that produce metabolites with high titers, such as ethanol, alanine, lactate and succinate.

  10. Escherichia coli growth under modeled reduced gravity

    NASA Technical Reports Server (NTRS)

    Baker, Paul W.; Meyer, Michelle L.; Leff, Laura G.

    2004-01-01

    Bacteria exhibit varying responses to modeled reduced gravity that can be simulated by clino-rotation. When Escherichia coli was subjected to different rotation speeds during clino-rotation, significant differences between modeled reduced gravity and normal gravity controls were observed only at higher speeds (30-50 rpm). There was no apparent affect of removing samples on the results obtained. When E. coli was grown in minimal medium (at 40 rpm), cell size was not affected by modeled reduced gravity and there were few differences in cell numbers. However, in higher nutrient conditions (i.e., dilute nutrient broth), total cell numbers were higher and cells were smaller under reduced gravity compared to normal gravity controls. Overall, the responses to modeled reduced gravity varied with nutrient conditions; larger surface to volume ratios may help compensate for the zone of nutrient depletion around the cells under modeled reduced gravity.

  11. Escherichia coli growth under modeled reduced gravity

    NASA Technical Reports Server (NTRS)

    Baker, Paul W.; Meyer, Michelle L.; Leff, Laura G.

    2004-01-01

    Bacteria exhibit varying responses to modeled reduced gravity that can be simulated by clino-rotation. When Escherichia coli was subjected to different rotation speeds during clino-rotation, significant differences between modeled reduced gravity and normal gravity controls were observed only at higher speeds (30-50 rpm). There was no apparent affect of removing samples on the results obtained. When E. coli was grown in minimal medium (at 40 rpm), cell size was not affected by modeled reduced gravity and there were few differences in cell numbers. However, in higher nutrient conditions (i.e., dilute nutrient broth), total cell numbers were higher and cells were smaller under reduced gravity compared to normal gravity controls. Overall, the responses to modeled reduced gravity varied with nutrient conditions; larger surface to volume ratios may help compensate for the zone of nutrient depletion around the cells under modeled reduced gravity.

  12. Transport proteins promoting Escherichia coli pathogenesis.

    PubMed

    Tang, Fengyi; Saier, Milton H

    2014-01-01

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

  13. Transport proteins promoting Escherichia coli pathogenesis

    PubMed Central

    Tang, Fengyi; Saier, Milton H.

    2014-01-01

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

  14. Extracellular recombinant protein production from Escherichia coli.

    PubMed

    Ni, Ye; Chen, Rachel

    2009-11-01

    Escherichia coli is the most commonly used host for recombinant protein production and metabolic engineering. Extracellular production of enzymes and proteins is advantageous as it could greatly reduce the complexity of a bioprocess and improve product quality. Extracellular production of proteins is necessary for metabolic engineering applications in which substrates are polymers such as lignocelluloses or xenobiotics since adequate uptake of these substrates is often an issue. The dogma that E. coli secretes no protein has been challenged by the recognition of both its natural ability to secrete protein in common laboratory strains and increased ability to secrete proteins in engineered cells. The very existence of this review dedicated to extracellular production is a testimony for outstanding achievements made collectively by the community in this regard. Four strategies have emerged to engineer E. coli cells to secrete recombinant proteins. In some cases, impressive secretion levels, several grams per liter, were reached. This secretion level is on par with other eukaryotic expression systems. Amid the optimism, it is important to recognize that significant challenges remain, especially when considering the success cannot be predicted a priori and involves much trials and errors. This review provides an overview of recent developments in engineering E. coli for extracellular production of recombinant proteins and an analysis of pros and cons of each strategy.

  15. Engineering Escherichia coli to bind to cyanobacteria.

    PubMed

    Zhang, Zijian; Meng, Liuyi; Ni, Congjian; Yao, Lanqiu; Zhang, Fengyu; Jin, Yuji; Mu, Xuelang; Zhu, Shiyu; Lu, Xiaoyu; Liu, Shiyu; Yu, Congyu; Wang, Chenggong; Zheng, Pu; Wu, Jie; Kang, Li; Zhang, Haoqian M; Ouyang, Qi

    2017-03-01

    We engineered Escherichia coli cells to bind to cyanobacteria by heterologously producing and displaying lectins of the target cyanobacteria on their surface. To prove the efficacy of our approach, we tested this design on Microcystis aeruginosa with microvirin (Mvn), the lectin endogenously produced by this cyanobacterium. The coding sequence of Mvn was C-terminally fused to the ice nucleation protein NC (INPNC) gene and expressed in E. coli. Results showed that E. coli cells expressing the INPNC::Mvn fusion protein were able to bind to M. aeruginosa and the average number of E. coli cells bound to each cyanobacterial cell was enhanced 8-fold. Finally, a computational model was developed to simulate the binding reaction and help reconstruct the binding parameters. To our best knowledge, this is the first report on the binding of two organisms in liquid culture mediated by the surface display of lectins and it may serve as a novel approach to mediate microbial adhesion.

  16. Inactivation of Escherichia coli by ultrasonic irradiation.

    PubMed

    Furuta, M; Yamaguchi, M; Tsukamoto, T; Yim, B; Stavarache, C E; Hasiba, K; Maeda, Y

    2004-04-01

    Ultrasonic inactivation of Escherichia coli XL1-Blue has been investigated by high-intensity ultrasonic waves from horn type sonicator (27.5 kHz) utilizing the "squeeze-film effect". The amplitude of the vibration face contacting the sample solution was used as an indication of the ultrasonic power intensity. The inactivation of the E. coli cells by ultrasonic irradiation shows pseudo first-order behavior. The inactivation rate constant gradually increased with increasing amplitude of the vibration face and showed rapid increase above 3 microm (p-p). In contrast, the H2O2 formation was not observed below 3 microm (p-p), indicating that the ultrasonic shock wave might be more important than indirect effect of OH radicals formed by ultrasonic cavitation in this system. The optimal thickness of the squeeze film was determined as 2 mm for the E. coli inactivation. More than 99% of E. coli cells was inactivated within 180-s sonication at the amplitude of 3 microm (p-p) and 2 mm of the thickness of the squeeze film.

  17. 21 CFR 866.3255 - Escherichia coli serological reagents.

    Code of Federal Regulations, 2014 CFR

    2014-04-01

    ... 21 Food and Drugs 8 2014-04-01 2014-04-01 false Escherichia coli serological reagents. 866.3255 Section 866.3255 Food and Drugs FOOD AND DRUG ADMINISTRATION, DEPARTMENT OF HEALTH AND HUMAN SERVICES... antisera conjugated with a fluorescent dye used to identify Escherichia coli directly from...

  18. 21 CFR 866.3255 - Escherichia coli serological reagents.

    Code of Federal Regulations, 2013 CFR

    2013-04-01

    ... 21 Food and Drugs 8 2013-04-01 2013-04-01 false Escherichia coli serological reagents. 866.3255 Section 866.3255 Food and Drugs FOOD AND DRUG ADMINISTRATION, DEPARTMENT OF HEALTH AND HUMAN SERVICES... antisera conjugated with a fluorescent dye used to identify Escherichia coli directly from...

  19. Susceptibilities of Escherichia coli and Staphylococcus aureus to Aloe barbadensis.

    PubMed

    Shilpakala, S R; Prathiba, J; Malathi, R

    2009-01-01

    The in vitro susceptibilities of Escherichia coli and Staphylococcus aureus were evaluated and the two organisms were susceptible to the inner gel of aloe barbadensis, though it was more effective against Staphylococcus aureus than Escherichia coli. The reduction for Aloe Vera (AV) needed to suppress the growth of the gram-positive bacterium was attributed to the structural differences between the two organisms.

  20. Engineering the biological conversion of methanol to specialty chemicals in Escherichia coli.

    PubMed

    Whitaker, W Brian; Jones, J Andrew; Bennett, R Kyle; Gonzalez, Jacqueline E; Vernacchio, Victoria R; Collins, Shannon M; Palmer, Michael A; Schmidt, Samuel; Antoniewicz, Maciek R; Koffas, Mattheos A; Papoutsakis, Eleftherios T

    2017-01-01

    Methanol is an attractive substrate for biological production of chemicals and fuels. Engineering methylotrophic Escherichia coli as a platform organism for converting methanol to metabolites is desirable. Prior efforts to engineer methylotrophic E. coli were limited by methanol dehydrogenases (Mdhs) with unfavorable enzyme kinetics. We engineered E. coli to utilize methanol using a superior NAD-dependent Mdh from Bacillus stearothermophilus and ribulose monophosphate (RuMP) pathway enzymes from B. methanolicus. Using (13)C-labeling, we demonstrate this E. coli strain converts methanol into biomass components. For example, the key TCA cycle intermediates, succinate and malate, exhibit labeling up to 39%, while the lower glycolytic intermediate, 3-phosphoglycerate, up to 53%. Multiple carbons are labeled for each compound, demonstrating a cycling RuMP pathway for methanol assimilation to support growth. By incorporating the pathway to synthesize the flavanone naringenin, we demonstrate the first example of in vivo conversion of methanol into a specialty chemical in E. coli.

  1. Humoral immune responses of amphioxus Branchiostoma belcheri to challenge with Escherichia coli.

    PubMed

    Pang, Qiuxiang; Zhang, Shicui; Liu, Xuemei; Wu, Di

    2006-08-01

    Humoral parameters of amphioxus Branchiostoma belcheri, including lysozyme, antimicrobial activity, microbial agglutinin and haemagglutinins were measured before and after challenge with Escherichia coli. Humoral fluids from unchallenged B. belcheri had lysozyme, antimicrobial, microbial agglutinating and haemagglutinating activities, which may represent part of the baseline level of innate immunity in this organism. After challenge with E. coli, the lysozyme activity, growth-inhibiting activities against E. coli and Vibrio alginolyticus, microbial agglutinating activities against Micrococcus lysodeikticus, Bacillus subtilis and Staphylococus aureus, and haemagglutinating activities against rabbit and human A and O erythrocytes in the humoral fluids were all increased significantly. In contrast, the agglutinating activities against Vibrio harveyi and E. coli and the haemagglutinating activity against human B erythrocytes in the humoral fluids were reduced in response to E. coli challenge. It appears that the humoral fluids of B. belcheri contain components that are able to differentiate different microbes and different human blood cell types.

  2. Effects of Toluene on Escherichia coli

    PubMed Central

    Jackson, Robert W.; DeMoss, J. A.

    1965-01-01

    Jackson, Robert W. (University of California, San Diego, La Jolla), and J. A. DeMoss. Effects of toluene on Escherichia coli. J. Bacteriol. 90:1420–1425. 1965.—When toluene is added at appropriate levels to exponentially growing cultures of Escherichia coli, a time-dependent loss of turbidity is observed which is concurrent with a loss of material to the medium and with unmasking of β-galactosidase. In addition, the galactoside permease system is totally destroyed. Electron micrographs confirm the indications that the cells are not being lysed by toluene, although the cytoplasm collapses to the interior of the cell. Included in the material lost from the cell after toluene treatment is 85% of the total ribonucleic acid (RNA), the principal source of which appears to be the ribosomes. The loss of RNA is temperature-dependent. Protein is also lost to the medium as a function of both temperature and available toluene. Up to 25% of the total protein is found in the medium, the precise amount depending on the level of toluene employed. Zone centrifugation studies of extracts from treated cells indicate that toluene elicits a rapid disaggregation of ribosomes that is terminated, at any stage, by disruption of the cells. The disaggregation is temperature-dependent and does not occur at 4 C. It appears to be distinct from the actual degradation of ribosomal RNA and is accompanied by an accumulation of small particles during the initial phases of treatment at 21 C. Toluene added to crude extracts of normal E. coli cells is unable to cause detectable ribosome destruction. Images PMID:5321488

  3. Pattern Formation of Bacterial Colonies by Escherichia coli

    NASA Astrophysics Data System (ADS)

    Tokita, Rie; Katoh, Takaki; Maeda, Yusuke; Wakita, Jun-ichi; Sano, Masaki; Matsuyama, Tohey; Matsushita, Mitsugu

    2009-07-01

    We have studied the morphological diversity and change in bacterial colonies, using the bacterial species Escherichia coli, as a function of both agar concentration Ca and nutrient concentration Cn. We observed various colony patterns, classified them into four types by pattern characteristics and established a morphological diagram by dividing it into four regions. They are regions A [diffusion-limited aggregation (DLA)-like], B (Eden-like), C (concentric-ring), and D (fluid-spreading). In particular, we have observed a concentric-ring colony growth for E. coli. We focused on the periodic growth in region C and obtained the following results: (i) A colony grows cyclically with the growing front repeating an advance (migration phase) and a momentary rest (consolidation phase) alternately. (ii) The growth width L and the bulge width W in one cycle decrease asymptotically to certain values, when Ca is increased. (iii) L does not depend on Cn, while W is an increasing function of Cn. Plausible mechanisms are proposed to explain the experimental results, by comparing them with those obtained for other bacterial species such as Proteus mirabilis and Bacillus subtilis.

  4. Comparison of 61 Sequenced Escherichia coli Genomes

    PubMed Central

    Lukjancenko, Oksana; Wassenaar, Trudy M.

    2010-01-01

    Escherichia coli is an important component of the biosphere and is an ideal model for studies of processes involved in bacterial genome evolution. Sixty-one publically available E. coli and Shigella spp. sequenced genomes are compared, using basic methods to produce phylogenetic and proteomics trees, and to identify the pan- and core genomes of this set of sequenced strains. A hierarchical clustering of variable genes allowed clear separation of the strains into clusters, including known pathotypes; clinically relevant serotypes can also be resolved in this way. In contrast, when in silico MLST was performed, many of the various strains appear jumbled and less well resolved. The predicted pan-genome comprises 15,741 gene families, and only 993 (6%) of the families are represented in every genome, comprising the core genome. The variable or ‘accessory’ genes thus make up more than 90% of the pan-genome and about 80% of a typical genome; some of these variable genes tend to be co-localized on genomic islands. The diversity within the species E. coli, and the overlap in gene content between this and related species, suggests a continuum rather than sharp species borders in this group of Enterobacteriaceae. PMID:20623278

  5. Role of Escherichia coli in Biofuel Production.

    PubMed

    Koppolu, Veerendra; Vasigala, Veneela Kr

    2016-01-01

    Increased energy consumption coupled with depleting petroleum reserves and increased greenhouse gas emissions have renewed our interest in generating fuels from renewable energy sources via microbial fermentation. Central to this problem is the choice of microorganism that catalyzes the production of fuels at high volumetric productivity and yield from cheap and abundantly available renewable energy sources. Microorganisms that are metabolically engineered to redirect renewable carbon sources into desired fuel products are contemplated as best choices to obtain high volumetric productivity and yield. Considering the availability of vast knowledge in genomic and metabolic fronts, Escherichia coli is regarded as a primary choice for the production of biofuels. Here, we reviewed the microbial production of liquid biofuels that have the potential to be used either alone or in combination with the present-day fuels. We specifically highlighted the metabolic engineering and synthetic biology approaches used to improve the production of biofuels from E. coli over the past few years. We also discussed the challenges that still exist for the biofuel production from E. coli and their possible solutions.

  6. Role of Escherichia coli in Biofuel Production

    PubMed Central

    Koppolu, Veerendra; Vasigala, Veneela KR

    2016-01-01

    Increased energy consumption coupled with depleting petroleum reserves and increased greenhouse gas emissions have renewed our interest in generating fuels from renewable energy sources via microbial fermentation. Central to this problem is the choice of microorganism that catalyzes the production of fuels at high volumetric productivity and yield from cheap and abundantly available renewable energy sources. Microorganisms that are metabolically engineered to redirect renewable carbon sources into desired fuel products are contemplated as best choices to obtain high volumetric productivity and yield. Considering the availability of vast knowledge in genomic and metabolic fronts, Escherichia coli is regarded as a primary choice for the production of biofuels. Here, we reviewed the microbial production of liquid biofuels that have the potential to be used either alone or in combination with the present-day fuels. We specifically highlighted the metabolic engineering and synthetic biology approaches used to improve the production of biofuels from E. coli over the past few years. We also discussed the challenges that still exist for the biofuel production from E. coli and their possible solutions. PMID:27441002

  7. Microbubble assisted polyhydroxybutyrate production in Escherichia coli.

    PubMed

    Inan, Kadriye; Sal, Fulya Ay; Rahman, Asif; Putman, Ryan J; Agblevor, Foster A; Miller, Charles D

    2016-07-09

    One of the potential limitations of large scale aerobic Escherichia coli fermentation is the need for increased dissolved oxygen for culture growth and bioproduct generation. As culture density increases the poor solubility of oxygen in water becomes one of the limiting factors for cell growth and product formation. A potential solution is to use a microbubble dispersion (MBD) generating device to reduce the diameter and increase the surface area of sparged bubbles in the fermentor. In this study, a recombinant E. coli strain was used to produce polyhydroxybutyrate (PHB) under conventional and MBD aerobic fermentation conditions. In conventional fermentation operating at 350 rpm and 0.8 vvm air flow rate, an OD600 of 6.21 and PHB yield of 23 % (dry cell basis) was achieved. MBD fermentation with similar bioreactor operating parameters produced an OD600 of 8.17 and PHB yield of 43 % PHB, which was nearly double that of the conventional fermentation. This study demonstrated that using a MBD generator can increase oxygen mass transfer into the aqueous phase, increasing E. coli growth and bioproduct generation.

  8. Regulation of alcohol fermentation by Escherichia coli

    SciTech Connect

    Clark, D.P.

    1989-01-01

    The purpose of this project is to elucidate the way in which the fermentative synthesis of ethanol is regulated in the facultative anaerobe Escherichia coli. We are also investigating the control of other genes required for fermentation and anaerobic growth. We have isolated both structural and regulatory mutations affecting the expression of alcohol dehydrogenase, the enzyme responsible for the final step in alcohol synthesis. Some of these regulatory mutations also affect other anaerobically induced genes. The adh gene has been cloned and sequenced. The ADH protein is one of the largest highly expressed proteins in E. coli and requires approximately 2700bp of DNA for its cloning sequence. We have also isolated mutations affecting the fermentative lactate dehydrogenase. In consequence it is now possible to construct E. coli strains defective in the production of any one or more of their normal fermentation products (i.e. formate, acetate, lactate, ethanol and succinate). The factors affecting the ratio of fermentation products are being investigated by in vivo NMR spectroscopy.

  9. Regulation of alcohol fermentation by Escherichia coli

    SciTech Connect

    Clark, D.P.

    1990-01-01

    The purpose of this project is to elucidate the way in which the synthesis of ethanol and related fermentation products are regulated in the facultative anaerobe Escherichia coli. We are also investigating the control of other genes required for anaerobic growth. We have isolated both structural and regulatory mutations affecting the expression of alcohol dehydrogenase, the enzyme responsible for the final step in alcohol synthesis. Some of these regulatory mutations also affect other anaerobically induced genes. The adh gene has been cloned and sequenced. The ADH protein is one of the largest highly expressed proteins in E. coli and requires approximately 2700bp of DNA for its coding sequence. We have also isolated mutations affecting the fermentative lactate dehydrogenase and have recently cloned the ldh gene. In consequence it is now possible to construct E. coli strains defective in the production of any one or more of their normal fermentation products (i.e. formate, acetate, lactate, ethanol and succinate). The factors affecting ratio of fermentation products are being investigated by in vivo NMR spectroscopy.

  10. Arabidopsis alternative oxidase sustains Escherichia coli respiration.

    PubMed Central

    Kumar, A M; Söll, D

    1992-01-01

    Glutamyl-tRNA reductase, encoded by the hemA gene, is the first enzyme in porphyrin biosynthesis in many organisms. Hemes, important porphyrin derivatives, are essential components of redox enzymes, such as cytochromes. Thus a hemA Escherichia coli strain (SASX41B) is deficient in cytochrome-mediated aerobic respiration. Upon complementation of this strain with an Arabidopsis thaliana cDNA library, we isolated a clone which permitted the SASX41B strain to grow aerobically. The clone encodes the gene for Arabidopsis alternative oxidase, whose deduced amino acid sequence was found to have 71% identity with that of the enzyme from the voodoo lily, Sauromatum guttatum. The Arabidopsis protein is expressed as a 31-kDa protein in E. coli and confers on this organism cyanide-resistant growth, which in turn is sensitive to salicylhydroxamate. This implies that a single polypeptide is sufficient for alternative oxidase activity. Based on these observations we propose that a cyanide-insensitive respiratory pathway operates in the transformed E. coli hemA strain. Introduction of this pathway now opens the way to genetic/molecular biological investigations of alternative oxidase and its cofactor. Images PMID:1438286

  11. Long term effects of Escherichia coli mastitis.

    PubMed

    Blum, Shlomo E; Heller, Elimelech D; Leitner, Gabriel

    2014-07-01

    Escherichia coli is one of the most frequently diagnosed causes of bovine mastitis, and is typically associated with acute, clinical mastitis. The objective of the present study was to evaluate the long term effects of intramammary infections by E. coli on milk yield and quality, especially milk coagulation. Twenty-four Israeli Holstein cows diagnosed with clinical mastitis due to intramammary infection by E. coli were used in this study. Mean lactation number, days in milk (DIM) and daily milk yield (DMY) at the time of infection was 3.3 ± 1.3, 131.7 days ± 78.6 and 45.7 L ± 8.4, respectively. DMY, milk constituents, somatic cells count (SCC), differential leukocytes count and coagulation parameters were subsequently assessed. Two patterns of inflammation were identified: 'short inflammation', characterized by <15% decrease in DMY and <30 days until return to normal (n = 5), and 'long inflammation', characterized by >15% decrease in DMY and >30 days to reach a new maximum DMY (n = 19). The estimated mean loss of marketable milk during the study was 200 L/cow for 'short inflammation' cases, and 1,500 L/cow for 'long inflammation' ones. Significant differences between 'short' and 'long inflammation' effects were found in almost all parameters studied. Long-term detrimental effects on milk quality were found regardless of clinical or bacteriological cure of affected glands.

  12. WGS accurately predicts antimicrobial resistance in Escherichia coli

    USDA-ARS?s Scientific Manuscript database

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

  13. Oxygen sensitivity of an Escherichia coli mutant.

    PubMed Central

    Adler, H; Mural, R; Suttle, B

    1992-01-01

    Genetic evidence indicates that Oxys-6, an oxygen-sensitive mutant of Escherichia coli AB1157, is defective in the region of the hemB locus. Oxys-6 is capable of growth under aerobic conditions only if cultures are initiated at low-inoculum levels. Aerobic liquid cultures are limited to a cell density of 10(7) cells per ml by the accumulation of a metabolically produced, low-molecular-weight, heat-stable material in complex organic media. Both Oxys-6 and AB1157 cells produce the material, but only aerobic cultures of the mutant are inhibited by it. The material is produced by both intact cells and cell extracts in complex media. This reaction also occurs when the amino acid L-lysine is substituted for complex media. Images PMID:1551829

  14. An overview of atypical enteropathogenic Escherichia coli.

    PubMed

    Hernandes, Rodrigo T; Elias, Waldir P; Vieira, Mônica A M; Gomes, Tânia A T

    2009-08-01

    The enteropathogenic Escherichia coli (EPEC) pathotype is currently divided into two groups, typical EPEC (tEPEC) and atypical EPEC (aEPEC). The property that distinguishes these two groups is the presence of the EPEC adherence factor plasmid, which is only found in tEPEC. aEPEC strains are emerging enteropathogens that have been detected worldwide. Herein, we review the serotypes, virulence properties, genetic relationships, epidemiology, reservoir and diagnosis of aEPEC, including those strains not belonging to the classical EPEC serogroups (nonclassical EPEC serogroups). The large variety of serotypes and genetic virulence properties of aEPEC strains from nonclassical EPEC serogroups makes it difficult to determine which strains are truly pathogenic.

  15. Mechanism of Escherichia coli Resistance to Pyrrhocoricin

    PubMed Central

    Narayanan, Shalini; Modak, Joyanta K.; Ryan, Catherine S.; Garcia-Bustos, Jose; Davies, John K.

    2014-01-01

    Due to their lack of toxicity to mammalian cells and good serum stability, proline-rich antimicrobial peptides (PR-AMPs) have been proposed as promising candidates for the treatment of infections caused by antimicrobial-resistant bacterial pathogens. It has been hypothesized that these peptides act on multiple targets within bacterial cells, and therefore the likelihood of the emergence of resistance was considered to be low. Here, we show that spontaneous Escherichia coli mutants resistant to pyrrhocoricin arise at a frequency of approximately 6 × 10−7. Multiple independently derived mutants all contained a deletion in a nonessential gene that encodes the putative peptide uptake permease SbmA. Sensitivity could be restored to the mutants by complementation with an intact copy of the sbmA gene. These findings question the viability of the development of insect PR-AMPs as antimicrobials. PMID:24590485

  16. Escherichia coli fliAZY operon.

    PubMed Central

    Mytelka, D S; Chamberlin, M J

    1996-01-01

    We have cloned the Escherichia coli fliAZY operon, which contains the fliA gene (the alternative sigma factor sigma F) and two novel genes, fliZ and fliY. Transcriptional mapping of this operon shows two start sites, one of which is preceded by a canonical E sigma F-dependent consensus and is dependent on sigma F for expression in vivo and in vitro. We have overexpressed and purified sigma F and demonstrated that it can direct core polymerase to E sigma F-dependent promoters. FliZ and FliY are not required for motility but may regulate sigma F activity, perhaps in response to a putative cell density signal that may be detected by FliY, a member of the bacterial extracellular solute-binding protein family 3. PMID:8550423

  17. Animal models of enteroaggregative Escherichia coli infection

    PubMed Central

    Philipson, Casandra W.; Bassaganya-Riera, Josep; Hontecillas, Raquel

    2013-01-01

    Enteroaggregative Escherichia coli (EAEC) has been acknowledged as an emerging cause of gastroenteritis worldwide for over two decades. Epidemiologists are revealing the role of EAEC in diarrheal outbreaks as a more common occurrence than ever suggested before. EAEC induced diarrhea is most commonly associated with travelers, children and immunocompromised individuals however its afflictions are not limited to any particular demographic. Many attributes have been discovered and characterized surrounding the capability of EAEC to provoke a potent pro-inflammatory immune response, however cellular and molecular mechanisms underlying initiation, progression and outcomes are largely unknown. This limited understanding can be attributed to heterogeneity in strains and the lack of adequate animal models. This review aims to summarize current knowledge about EAEC etiology, pathogenesis and clinical manifestation. Additionally, current animal models and their limitations will be discussed along with the value of applying systems-wide approaches such as computational modeling to study host-EAEC interactions. PMID:23680797

  18. Recombinational construction in Escherichia coli of infectious adenoviral genomes

    PubMed Central

    Crouzet, Joël; Naudin, Laurent; Orsini, Cécile; Vigne, Emmanuelle; Ferrero, Lucy; Le Roux, Aude; Benoit, Patrick; Latta, Martine; Torrent, Christophe; Branellec, Didier; Denèfle, Patrice; Mayaux, Jean-François; Perricaudet, Michel; Yeh, Patrice

    1997-01-01

    A two-step gene replacement procedure was developed that generates infectious adenoviral genomes through homologous recombination in Escherichia coli. As a prerequisite, a human adenovirus serotype 5 (Ad5)-derived genome was first introduced as a PacI restriction fragment into an incP-derived replicon which, in contrast to ColE1-derivatives (e.g., pBR322 or pUC plasmids), is functional in a polA mutant of E. coli. Any modification can be introduced at will following two consecutive homologous recombinations between the incP/Ad5 replicon and the ColE1 plasmid. The overall procedure requires only the in vitro engineering of the ColE1-derivative by flanking the desired modification with small stretches of identical sequences. In the first step, a cointegrate between the tetracycline-resistant incP/Ad5 replicon and the kanamycin-resistant ColE1-derivative is selected by growing the polA host in the presence of both antibiotics. Resolution of this cointegrate is further selected in sucrose growth conditions due to the loss of a conditional suicide marker (the sacB gene of Bacillus subtilis) present in the ColE1 plasmid, leading to unmodified and modified incP/Ad5 replicons that can be differentiated upon restriction analysis. Consecutive rounds of this two-step cloning procedure allowed the introduction of multiple independent modifications within the virus genome, with no requirement for an intermediate virus. The potential of this procedure is demonstrated by the recovery of several E1E3E4-deleted adenoviruses following transfection of the corresponding E. coli-derived genomes in IGRP2 cells. PMID:9037067

  19. The thermal impulse response of Escherichia coli

    PubMed Central

    Paster, Eli; Ryu, William S.

    2008-01-01

    Swimming Escherichia coli responds to changes in temperature by modifying its motor behavior. Previous studies using populations of cells have shown that E. coli accumulate in spatial thermal gradients, but these experiments did not cleanly separate thermal responses from chemotactic responses. Here we have isolated the thermal response by studying the behavior of single, tethered cells. The motor output of cells grown at 33°C was measured at constant temperature, from 10° to 40°C, and in response to small, impulsive increases in temperature, from 23° to 43°C. The thermal impulse response at temperatures < 31°C is similar to the chemotactic impulse response: Both follow a similar time course, share the same directionality, and show biphasic characteristics. At temperatures > 31°C, some cells show an inverted response, switching from warm- to cold-seeking behavior. The fraction of inverted responses increases nonlinearly with temperature, switching steeply at the preferred temperature of 37°C. PMID:18385380

  20. Characterization of enterotoxigenic bovine Escherichia coli.

    PubMed Central

    Sivaswamy, G; Gyles, C L

    1976-01-01

    Among 300 isolates of bovine Escherichia coli, 56 which had been found enterotoxigenic in calf gut loops were characterized on the basis of O and K antigens, colonial morphology and resistance to seven antimicrobial drugs. The 56 isolates enterotoxigenic in the calf were compared with the nonenterotoxigenic ones. Of the 56 enterotoxigenic E. coli the majority possessed the A type of K antigen and had OK groups, O9:K(PS274) or O101:K(RVC118). Fourteen of these isolates had the K99 antigen. None of 27 isolates found enterotoxigenic in the piglet but not in the calf possessed the K99 antigen or belonged to OK groups O9:K(PS274) or O101:K(RVC118). Comparison of the patterns of resistance to seven antimicrobial drugs showed that all enterotoxigenic and nonenterotoxigenic isolates were susceptible to nitrofurantoin and sulphachlorphyridiazine and that there was no significant difference in the patterns between the two groups. The majority of enterotoxigenic isolates were mucoid, whereas most of the nonenterotoxigenic isolates were nonmucoid. PMID:793694

  1. The crystal structure Escherichia coli Spy

    PubMed Central

    Kwon, Eunju; Kim, Dong Young; Gross, Carol A; Gross, John D; Kim, Kyeong Kyu

    2010-01-01

    Escherichia coli spheroplast protein y (EcSpy) is a small periplasmic protein that is homologous with CpxP, an inhibitor of the extracytoplasmic stress reponse. Stress conditions such as spheroplast formation induce the expression of Spy via the Cpx or the Bae two-component systems in E. coli, though the function of Spy is unknown. Here, we report the crystal structure of EcSpy, which reveals a long kinked hairpin-like structure of four α-helices that form an antiparallel dimer. The dimer contains a curved oval shape with a highly positively charged concave surface that may function as a ligand binding site. Sequence analysis reveals that Spy is highly conserved over the Enterobacteriaceae family. Notably, three conserved regions that contain identical residues and two LTxxQ motifs are placed at the horizontal end of the dimer structure, stablizing the overall fold. CpxP also contains the conserved sequence motifs and has a predicted secondary structure similar to Spy, suggesting that Spy and CpxP likely share the same fold. PMID:20799348

  2. The crystal structure Escherichia coli Spy.

    PubMed

    Kwon, Eunju; Kim, Dong Young; Gross, Carol A; Gross, John D; Kim, Kyeong Kyu

    2010-11-01

    Escherichia coli spheroplast protein y (EcSpy) is a small periplasmic protein that is homologous with CpxP, an inhibitor of the extracytoplasmic stress response. Stress conditions such as spheroplast formation induce the expression of Spy via the Cpx or the Bae two-component systems in E. coli, though the function of Spy is unknown. Here, we report the crystal structure of EcSpy, which reveals a long kinked hairpin-like structure of four α-helices that form an antiparallel dimer. The dimer contains a curved oval shape with a highly positively charged concave surface that may function as a ligand binding site. Sequence analysis reveals that Spy is highly conserved over the Enterobacteriaceae family. Notably, three conserved regions that contain identical residues and two LTxxQ motifs are placed at the horizontal end of the dimer structure, stabilizing the overall fold. CpxP also contains the conserved sequence motifs and has a predicted secondary structure similar to Spy, suggesting that Spy and CpxP likely share the same fold.

  3. Chemotaxis Toward Sugars in Escherichia coli

    PubMed Central

    Adler, Julius; Hazelbauer, Gerald L.; Dahl, M. M.

    1973-01-01

    Using a quantitative assay for measuring chemotaxis, we tested a variety of sugars and sugar derivatives for their ability to attract Escherichia coli bacteria. The most effective attractants, i.e., those that have thresholds near 10−5 M or below, are N-acetyl-d-glucosamine, 6-deoxy-d-glucose, d-fructose, d-fucose, 1-d-glycerol-β-d-galactoside, galactitol, d-galactose, d-glucosamine, d-glucose, α-d-glucose-1-phosphate, lactose, maltose, d-mannitol, d-mannose, methyl-β-d-galactoside, methyl-β-d-glucoside, d-ribose, d-sorbitol, and trehalose. Lactose, and probably d-glucose-1-phosphate, are attractive only after conversion to the free monosaccharide, while the other attractants do not require breakdown for taxis. Nine different chemoreceptors are involved in detecting these various attractants. They are called the N-acetyl-glucosamine, fructose, galactose, glucose, maltose, mannitol, ribose, sorbitol, and trehalose chemoreceptors; the specificity of each was studied. The chemoreceptors, with the exception of the one for d-glucose, are inducible. The galactose-binding protein serves as the recognition component of the galactose chemoreceptor. E. coli also has osmotically shockable binding activities for maltose and d-ribose, and these appear to serve as the recognition components for the corresponding chemoreceptors. PMID:4580570

  4. Expanding ester biosynthesis in Escherichia coli

    PubMed Central

    Rodriguez, Gabriel M; Tashiro, Yohei; Atsumi, Shota

    2015-01-01

    To expand the capabilities of whole-cell biocatalysis, we have engineered Escherichia coli to produce various esters. The alcohol O-acyltransferase (ATF) class of enzyme uses acyl-CoA units for ester formation. The release of free CoA upon esterification with an alcohol provides the free energy to facilitate ester formation. The diversity of CoA molecules found in nature in combination with various alcohol biosynthetic pathways allows for the biosynthesis of a multitude of esters. Small to medium volatile esters have extensive applications in the flavor, fragrance, cosmetic, solvent, paint and coating industries. The present work enables the production of these compounds by designing several ester pathways in E. coli. The engineered pathways generated acetate esters of ethyl, propyl, isobutyl, 2-methyl-1-butyl, 3-methyl-1-butyl and 2-phenylethyl alcohols. In particular, we achieved high-level production of isobutyl acetate from glucose (17.2 g l−1). This strategy was expanded to realize pathways for tetradecyl acetate and several isobutyrate esters. PMID:24609358

  5. [Enteroinvasive Escherichia coli. Pathogenesis and epidemiology].

    PubMed

    Prats, G; Llovet, T

    1995-03-01

    Enteroinvasive Escherichia coli (EIEC) is an intestinal pathogen causing enteritis, with a similar pathogenic mechanism to that of Shigella, which causes an epithelial invasion of the large bowel leading to inflammation and ulceration of the mucosa. The patients often develop the symptoms of bacillary dysentery. The EIEC strains are atypical in their biochemical reactions and may ferment lactose late or not at all, are lysine decarboxilase negative, and non motile. In addition, most EIEC strains express somatic antigens which are either strongly related or identical to Shigella antigens. EIEC invasion is mediated by a large plasmid (140 MDa) coding for the production of several outer membrane proteins involved in invasiveness. These strains have been isolated with some regularity in South America, the Extreme Orient, and Eastern Europe. In Spain the incidence of enteroinvasive E. coli is extraordinarily low (0.2%), the serogroup O124 being the most frequently isolated. EIEC enteritis has been associated to sporadic cases occurring in travellers. Occasional outbreaks related to ingestion of contaminated water or food and person to person have been reported.

  6. Isobutanol production from cellobiose in Escherichia coli.

    PubMed

    Desai, Shuchi H; Rabinovitch-Deere, Christine A; Tashiro, Yohei; Atsumi, Shota

    2014-04-01

    Converting lignocellulosics into biofuels remains a promising route for biofuel production. To facilitate strain development for specificity and productivity of cellulosic biofuel production, a user friendly Escherichia coli host was engineered to produce isobutanol, a drop-in biofuel candidate, from cellobiose. A beta-glucosidase was expressed extracellularly by either excretion into the media, or anchoring to the cell membrane. The excretion system allowed for E. coli to grow with cellobiose as a sole carbon source at rates comparable to those with glucose. The system was then combined with isobutanol production genes in three different configurations to determine whether gene arrangement affected isobutanol production. The most productive strain converted cellobiose to isobutanol in titers of 7.64 ± 0.19 g/L with a productivity of 0.16 g/L/h. These results demonstrate that efficient cellobiose degradation and isobutanol production can be achieved by a single organism, and provide insight for optimization of strains for future use in a consolidated bioprocessing system for renewable production of isobutanol.

  7. gltBDF operon of Escherichia coli.

    PubMed Central

    Castaño, I; Bastarrachea, F; Covarrubias, A A

    1988-01-01

    A 2.0-kilobase DNA fragment carrying antibiotic resistance markers was inserted into the gltB gene of Escherichia coli previously cloned in a multicopy plasmid. Replacement of the chromosomal gltB+ gene by the gltB225::omega mutation led to cells unable to synthesize glutamate synthase, utilize growth rate-limiting nitrogen sources, or derepress their glutamine synthetase. The existence of a gltBDF operon encoding the large (gltB) and small (gltD) subunits of glutamate synthase and a regulatory peptide (gltF) at 69 min of the E. coli linkage map was deduced from complementation analysis. A plasmid carrying the entire gltB+D+F+ operon complemented cells for all three of the mutant phenotypes associated with the polar gltB225::omega mutation in the chromosome. By contrast, plasmids carrying gltB+ only complemented cells for glutamate synthase activity. A major tricistronic mRNA molecule was detected from Northern (RNA blot) DNA-RNA hybridization experiments with DNA probes containing single genes of the operon. A 30,200-dalton polypeptide was identified as the gltF product, the lack of which was responsible for the inability of cells to use nitrogen-limiting sources associated with gltB225::omega. Images PMID:2448295

  8. RESISTANCE OF ESCHERICHIA COLI TO TETRACYCLINES.

    PubMed

    FRANKLIN, T J; GODFREY, A

    1965-01-01

    1. A strain of Escherichia coli highly resistant to chlortetracycline and partially cross-resistant to tetracycline has been isolated. 2. The nitro-reductase system of the resistant cells was inhibited to a smaller extent by chlortetracycline than was the corresponding enzyme of sensitive cells. 3. The incorporation of leucine in vitro into the ribosomal protein of cell-free preparations from sensitive and resistant cells was equally inhibited by chlortetracycline. 4. Resistant cells accumulated much less chlortetracycline and tetracycline than did sensitive cells when both were cultured in the presence of these drugs. 5. The uptake of tetracycline by both sensitive and resistant E. coli was dependent on the presence of glucose in the medium. 6. Fractionation of cells cultured in medium containing [(14)C]chlortetracycline indicated that the largest proportion of radioactivity in sensitive cells was in the fraction consisting mainly of cell-wall material. There was no concentration of radioactivity in any one fraction of the resistant cells. 7. No evidence could be obtained for a specific tetracycline-excretion system in the resistant cells. 8. The significance of these results in relation to current theories of the antibiotic action of and resistance to the tetracycline drugs is discussed.

  9. Tuning Escherichia coli for membrane protein overexpression.

    PubMed

    Wagner, Samuel; Klepsch, Mirjam M; Schlegel, Susan; Appel, Ansgar; Draheim, Roger; Tarry, Michael; Högbom, Martin; van Wijk, Klaas J; Slotboom, Dirk J; Persson, Jan O; de Gier, Jan-Willem

    2008-09-23

    A simple generic method for optimizing membrane protein overexpression in Escherichia coli is still lacking. We have studied the physiological response of the widely used "Walker strains" C41(DE3) and C43(DE3), which are derived from BL21(DE3), to membrane protein overexpression. For unknown reasons, overexpression of many membrane proteins in these strains is hardly toxic, often resulting in high overexpression yields. By using a combination of physiological, proteomic, and genetic techniques we have shown that mutations in the lacUV5 promoter governing expression of T7 RNA polymerase are key to the improved membrane protein overexpression characteristics of the Walker strains. Based on this observation, we have engineered a derivative strain of E. coli BL21(DE3), termed Lemo21(DE3), in which the activity of the T7 RNA polymerase can be precisely controlled by its natural inhibitor T7 lysozyme (T7Lys). Lemo21(DE3) is tunable for membrane protein overexpression and conveniently allows optimizing overexpression of any given membrane protein by using only a single strain rather than a multitude of different strains. The generality and simplicity of our approach make it ideal for high-throughput applications.

  10. Tuning Escherichia coli for membrane protein overexpression

    PubMed Central

    Wagner, Samuel; Klepsch, Mirjam M.; Schlegel, Susan; Appel, Ansgar; Draheim, Roger; Tarry, Michael; Högbom, Martin; van Wijk, Klaas J.; Slotboom, Dirk J.; Persson, Jan O.; de Gier, Jan-Willem

    2008-01-01

    A simple generic method for optimizing membrane protein overexpression in Escherichia coli is still lacking. We have studied the physiological response of the widely used “Walker strains” C41(DE3) and C43(DE3), which are derived from BL21(DE3), to membrane protein overexpression. For unknown reasons, overexpression of many membrane proteins in these strains is hardly toxic, often resulting in high overexpression yields. By using a combination of physiological, proteomic, and genetic techniques we have shown that mutations in the lacUV5 promoter governing expression of T7 RNA polymerase are key to the improved membrane protein overexpression characteristics of the Walker strains. Based on this observation, we have engineered a derivative strain of E. coli BL21(DE3), termed Lemo21(DE3), in which the activity of the T7 RNA polymerase can be precisely controlled by its natural inhibitor T7 lysozyme (T7Lys). Lemo21(DE3) is tunable for membrane protein overexpression and conveniently allows optimizing overexpression of any given membrane protein by using only a single strain rather than a multitude of different strains. The generality and simplicity of our approach make it ideal for high-throughput applications. PMID:18796603

  11. Nucleotide excision repair in Escherichia coli.

    PubMed Central

    Van Houten, B

    1990-01-01

    One of the best-studied DNA repair pathways is nucleotide excision repair, a process consisting of DNA damage recognition, incision, excision, repair resynthesis, and DNA ligation. Escherichia coli has served as a model organism for the study of this process. Recently, many of the proteins that mediate E. coli nucleotide excision have been purified to homogeneity; this had led to a molecular description of this repair pathway. One of the key repair enzymes of this pathway is the UvrABC nuclease complex. The individual subunits of this enzyme cooperate in a complex series of partial reactions to bind to and incise the DNA near a damaged nucleotide. The UvrABC complex displays a remarkable substrate diversity. Defining the structural features of DNA lesions that provide the specificity for damage recognition by the UvrABC complex is of great importance, since it represents a unique form of protein-DNA interaction. Using a number of in vitro assays, researchers have been able to elucidate the action mechanism of the UvrABC nuclease complex. Current research is devoted to understanding how these complex events are mediated within the living cell. PMID:2181258

  12. Expanding ester biosynthesis in Escherichia coli.

    PubMed

    Rodriguez, Gabriel M; Tashiro, Yohei; Atsumi, Shota

    2014-04-01

    To expand the capabilities of whole-cell biocatalysis, we have engineered Escherichia coli to produce various esters. The alcohol O-acyltransferase (ATF) class of enzyme uses acyl-CoA units for ester formation. The release of free CoA upon esterification with an alcohol provides the free energy to facilitate ester formation. The diversity of CoA molecules found in nature in combination with various alcohol biosynthetic pathways allows for the biosynthesis of a multitude of esters. Small to medium volatile esters have extensive applications in the flavor, fragrance, cosmetic, solvent, paint and coating industries. The present work enables the production of these compounds by designing several ester pathways in E. coli. The engineered pathways generated acetate esters of ethyl, propyl, isobutyl, 2-methyl-1-butyl, 3-methyl-1-butyl and 2-phenylethyl alcohols. In particular, we achieved high-level production of isobutyl acetate from glucose (17.2 g l(-1)). This strategy was expanded to realize pathways for tetradecyl acetate and several isobutyrate esters.

  13. Independence of replisomes in Escherichia coli chromosomalreplication

    SciTech Connect

    Breier, Adam M.; Weier, Heinz-Ulrich G.; Cozzarelli, Nicholas R.

    2005-03-13

    In Escherichia coli DNA replication is carried out by the coordinated action of the proteins within a replisome. After replication initiation, the two bidirectionally oriented replisomes from a single origin are colocalized into higher-order structures termed replication factories. The factory model postulated that the two replisomes are also functionally coupled. We tested this hypothesis by using DNA combing and whole-genome microarrays. Nascent DNA surrounding oriC in single, combed chromosomes showed instead that one replisome, usually the leftward one, was significantly ahead of the other 70% of the time. We next used microarrays to follow replication throughout the genome by measuring DNA copy number. We found in multiple E. coli strains that the replisomes are independent, with the leftward replisome ahead of the rightward one. The size of the bias was strain-specific, varying from 50 to 130 kb in the array results. When we artificially blocked one replisome, the other continued unabated, again demonstrating independence. We suggest an improved version of the factory model that retains the advantages of threading DNA through colocalized replisomes at about equal rates, but allows the cell flexibility to overcome obstacles encountered during elongation.

  14. Metabolism of Escherichia coli injured by copper.

    PubMed

    Domek, M J; Robbins, J E; Anderson, M E; McFeters, G A

    1987-01-01

    Escherichia coli injured by copper in carbonate buffer simulating the drinking water environment showed decreased oxygen utilization. Oxygraph measurements revealed that copper-injured bacteria had a rate of oxygen utilization that was less than 25% of that of control cells. Respirometry experiments measured rates over a longer period of time and showed similar trends. Nuclear magnetic resonance spectroscopy (13C nmr) and gas chromatography were used to identify differences in metabolism between healthy and injured populations of E. coli. The rate of glucose utilization by injured cells under anaerobic conditions was 64% of that of healthy cells. The rates of lactate and ethanol accumulation were 88 and 50% of the control, respectively. The 13C nmr studies of oxygenated cultures revealed differences in the accumulation of acetate and glutamine. Aerobic utilization of glucose and succinate by injured cells were 87 and 21% of the rate of the controls, respectively. Additional studies revealed injured cells had a decreased ability to reduce 2-(p-iodophenyl)-3-(p-nitrophenyl)-5-phenyltetrazolium chloride (INT) with a variety of carbohydrate substrates. Injured cells reduced greater quantities of INT than healthy cells when NADH was used as a substrate. A comparison of metabolic end products suggested that injured cells also had considerable differences in carbon flow compared with healthy cells.

  15. Biosynthesis of ethylene glycol in Escherichia coli.

    PubMed

    Liu, Huaiwei; Ramos, Kristine Rose M; Valdehuesa, Kris Niño G; Nisola, Grace M; Lee, Won-Keun; Chung, Wook-Jin

    2013-04-01

    Ethylene glycol (EG) is an important platform chemical with steadily expanding global demand. Its commercial production is currently limited to fossil resources; no biosynthesis route has been delineated. Herein, a biosynthesis route for EG production from D-xylose is reported. This route consists of four steps: D-xylose → D-xylonate → 2-dehydro-3-deoxy-D-pentonate → glycoaldehyde → EG. Respective enzymes, D-xylose dehydrogenase, D-xylonate dehydratase, 2-dehydro-3-deoxy-D-pentonate aldolase, and glycoaldehyde reductase, were assembled. The route was implemented in a metabolically engineered Escherichia coli, in which the D-xylose → D-xylulose reaction was prevented by disrupting the D-xylose isomerase gene. The most efficient construct produced 11.7 g L(-1) of EG from 40.0 g L(-1) of D-xylose. Glycolate is a carbon-competing by-product during EG production in E. coli; blockage of glycoaldehyde → glycolate reaction was also performed by disrupting the gene encoding aldehyde dehydrogenase, but from this approach, EG productivity was not improved but rather led to D-xylonate accumulation. To channel more carbon flux towards EG than the glycolate pathway, further systematic metabolic engineering and fermentation optimization studies are still required to improve EG productivity.

  16. Cyclomodulins in urosepsis strains of Escherichia coli.

    PubMed

    Dubois, Damien; Delmas, Julien; Cady, Anne; Robin, Frédéric; Sivignon, Adeline; Oswald, Eric; Bonnet, Richard

    2010-06-01

    Determinants of urosepsis in Escherichia coli remain incompletely defined. Cyclomodulins (CMs) are a growing functional family of toxins that hijack the eukaryotic cell cycle. Four cyclomodulin types are actually known in E. coli: cytotoxic necrotizing factors (CNFs), cycle-inhibiting factor (Cif), cytolethal distending toxins (CDTs), and the pks-encoded toxin. In the present study, the distribution of CM-encoding genes and the functionality of these toxins were investigated in 197 E. coli strains isolated from patients with community-acquired urosepsis (n = 146) and from uninfected subjects (n = 51). This distribution was analyzed in relation to the phylogenetic background, clinical origin, and antibiotic resistance of the strains. It emerged from this study that strains harboring the pks island and the cnf1 gene (i) were strongly associated with the B2 phylogroup (P, <0.001), (ii) frequently harbored both toxin-encoded genes in phylogroup B2 (33%), and (iii) were predictive of a urosepsis origin (P, <0.001 to 0.005). However, the prevalences of the pks island among phylogroup B2 strains, in contrast to those of the cnf1 gene, were not significantly different between fecal and urosepsis groups, suggesting that the pks island is more important for the colonization process and the cnf1 gene for virulence. pks- or cnf1-harboring strains were significantly associated with susceptibility to antibiotics (amoxicillin, cotrimoxazole, and quinolones [P, <0.001 to 0.043]). Otherwise, only 6% and 1% of all strains harbored the cdtB and cif genes, respectively, with no particular distribution by phylogenetic background, antimicrobial susceptibility, or clinical origin.

  17. The extracellular RNA complement of Escherichia coli.

    PubMed

    Ghosal, Anubrata; Upadhyaya, Bimal Babu; Fritz, Joëlle V; Heintz-Buschart, Anna; Desai, Mahesh S; Yusuf, Dilmurat; Huang, David; Baumuratov, Aidos; Wang, Kai; Galas, David; Wilmes, Paul

    2015-01-21

    The secretion of biomolecules into the extracellular milieu is a common and well-conserved phenomenon in biology. In bacteria, secreted biomolecules are not only involved in intra-species communication but they also play roles in inter-kingdom exchanges and pathogenicity. To date, released products, such as small molecules, DNA, peptides, and proteins, have been well studied in bacteria. However, the bacterial extracellular RNA complement has so far not been comprehensively characterized. Here, we have analyzed, using a combination of physical characterization and high-throughput sequencing, the extracellular RNA complement of both outer membrane vesicle (OMV)-associated and OMV-free RNA of the enteric Gram-negative model bacterium Escherichia coli K-12 substrain MG1655 and have compared it to its intracellular RNA complement. Our results demonstrate that a large part of the extracellular RNA complement is in the size range between 15 and 40 nucleotides and is derived from specific intracellular RNAs. Furthermore, RNA is associated with OMVs and the relative abundances of RNA biotypes in the intracellular, OMV and OMV-free fractions are distinct. Apart from rRNA fragments, a significant portion of the extracellular RNA complement is composed of specific cleavage products of functionally important structural noncoding RNAs, including tRNAs, 4.5S RNA, 6S RNA, and tmRNA. In addition, the extracellular RNA pool includes RNA biotypes from cryptic prophages, intergenic, and coding regions, of which some are so far uncharacterised, for example, transcripts mapping to the fimA-fimL and ves-spy intergenic regions. Our study provides the first detailed characterization of the extracellular RNA complement of the enteric model bacterium E. coli. Analogous to findings in eukaryotes, our results suggest the selective export of specific RNA biotypes by E. coli, which in turn indicates a potential role for extracellular bacterial RNAs in intercellular communication. © 2015 The

  18. The extracellular RNA complement of Escherichia coli

    PubMed Central

    Ghosal, Anubrata; Upadhyaya, Bimal Babu; Fritz, Joëlle V; Heintz-Buschart, Anna; Desai, Mahesh S; Yusuf, Dilmurat; Huang, David; Baumuratov, Aidos; Wang, Kai; Galas, David; Wilmes, Paul

    2015-01-01

    The secretion of biomolecules into the extracellular milieu is a common and well-conserved phenomenon in biology. In bacteria, secreted biomolecules are not only involved in intra-species communication but they also play roles in inter-kingdom exchanges and pathogenicity. To date, released products, such as small molecules, DNA, peptides, and proteins, have been well studied in bacteria. However, the bacterial extracellular RNA complement has so far not been comprehensively characterized. Here, we have analyzed, using a combination of physical characterization and high-throughput sequencing, the extracellular RNA complement of both outer membrane vesicle (OMV)-associated and OMV-free RNA of the enteric Gram-negative model bacterium Escherichia coli K-12 substrain MG1655 and have compared it to its intracellular RNA complement. Our results demonstrate that a large part of the extracellular RNA complement is in the size range between 15 and 40 nucleotides and is derived from specific intracellular RNAs. Furthermore, RNA is associated with OMVs and the relative abundances of RNA biotypes in the intracellular, OMV and OMV-free fractions are distinct. Apart from rRNA fragments, a significant portion of the extracellular RNA complement is composed of specific cleavage products of functionally important structural noncoding RNAs, including tRNAs, 4.5S RNA, 6S RNA, and tmRNA. In addition, the extracellular RNA pool includes RNA biotypes from cryptic prophages, intergenic, and coding regions, of which some are so far uncharacterised, for example, transcripts mapping to the fimA-fimL and ves-spy intergenic regions. Our study provides the first detailed characterization of the extracellular RNA complement of the enteric model bacterium E. coli. Analogous to findings in eukaryotes, our results suggest the selective export of specific RNA biotypes by E. coli, which in turn indicates a potential role for extracellular bacterial RNAs in intercellular communication. PMID:25611733

  19. Identification of pseudouridine methyltransferase in Escherichia coli

    PubMed Central

    Ero, Rya; Peil, Lauri; Liiv, Aivar; Remme, Jaanus

    2008-01-01

    In ribosomal RNA, modified nucleosides are found in functionally important regions, but their function is obscure. Stem–loop 69 of Escherichia coli 23S rRNA contains three modified nucleosides: pseudouridines at positions 1911 and 1917, and N3 methyl-pseudouridine (m3Ψ) at position 1915. The gene for pseudouridine methyltransferase was previously not known. We identified E. coli protein YbeA as the methyltransferase methylating Ψ1915 in 23S rRNA. The E. coli ybeA gene deletion strain lacks the N3 methylation at position 1915 of 23S rRNA as revealed by primer extension and nucleoside analysis by HPLC. Methylation at position 1915 is restored in the ybeA deletion strain when recombinant YbeA protein is expressed from a plasmid. In addition, we show that purified YbeA protein is able to methylate pseudouridine in vitro using 70S ribosomes but not 50S subunits from the ybeA deletion strain as substrate. Pseudouridine is the preferred substrate as revealed by the inability of YbeA to methylate uridine at position 1915. This shows that YbeA is acting at the final stage during ribosome assembly, probably during translation initiation. Hereby, we propose to rename the YbeA protein to RlmH according to uniform nomenclature of RNA methyltransferases. RlmH belongs to the SPOUT superfamily of methyltransferases. RlmH was found to be well conserved in bacteria, and the gene is present in plant and in several archaeal genomes. RlmH is the first pseudouridine specific methyltransferase identified so far and is likely to be the only one existing in bacteria, as m3Ψ1915 is the only methylated pseudouridine in bacteria described to date. PMID:18755836

  20. Identification of pseudouridine methyltransferase in Escherichia coli.

    PubMed

    Ero, Rya; Peil, Lauri; Liiv, Aivar; Remme, Jaanus

    2008-10-01

    In ribosomal RNA, modified nucleosides are found in functionally important regions, but their function is obscure. Stem-loop 69 of Escherichia coli 23S rRNA contains three modified nucleosides: pseudouridines at positions 1911 and 1917, and N3 methyl-pseudouridine (m(3)Psi) at position 1915. The gene for pseudouridine methyltransferase was previously not known. We identified E. coli protein YbeA as the methyltransferase methylating Psi1915 in 23S rRNA. The E. coli ybeA gene deletion strain lacks the N3 methylation at position 1915 of 23S rRNA as revealed by primer extension and nucleoside analysis by HPLC. Methylation at position 1915 is restored in the ybeA deletion strain when recombinant YbeA protein is expressed from a plasmid. In addition, we show that purified YbeA protein is able to methylate pseudouridine in vitro using 70S ribosomes but not 50S subunits from the ybeA deletion strain as substrate. Pseudouridine is the preferred substrate as revealed by the inability of YbeA to methylate uridine at position 1915. This shows that YbeA is acting at the final stage during ribosome assembly, probably during translation initiation. Hereby, we propose to rename the YbeA protein to RlmH according to uniform nomenclature of RNA methyltransferases. RlmH belongs to the SPOUT superfamily of methyltransferases. RlmH was found to be well conserved in bacteria, and the gene is present in plant and in several archaeal genomes. RlmH is the first pseudouridine specific methyltransferase identified so far and is likely to be the only one existing in bacteria, as m(3)Psi1915 is the only methylated pseudouridine in bacteria described to date.

  1. Biochemical properties and yields of diverse bacterial laccase-like multicopper oxidases expressed in Escherichia coli

    PubMed Central

    Ihssen, Julian; Reiss, Renate; Luchsinger, Ronny; Thöny-Meyer, Linda; Richter, Michael

    2015-01-01

    Laccases are multi-copper oxidases that oxidize a broad range of substrates at the expense of molecular oxygen, without any need for co-factor regeneration. These enzymes bear high potential for the sustainable synthesis of fine chemicals and the modification of (bio)polymers. Here we describe cloning and expression of five novel bacterial laccase-like multi copper oxidases (LMCOs) of diverse origin which were identified by homology searches in online databases. Activity yields under different expression conditions and temperature stabilities were compared to three previously described enzymes from Bacillus subtilis, Bacillus pumilus and Bacillus clausii. In almost all cases, a switch to oxygen-limited growth conditions after induction increased volumetric activity considerably. For proteins with predicted signal peptides for secretion, recombinant expression with and without signal sequence was investigated. Bacillus CotA-type LMCOs outperformed enzymes from Streptomyces and Gram-negative bacteria with respect to activity yields in Escherichia coli and application relevant biochemical properties. The novel Bacillus coagulans LMCO combined high activity yields in E. coli with unprecedented activity at strong alkaline pH and high storage stability, making it a promising candidate for further development. PMID:26068013

  2. Biochemical properties and yields of diverse bacterial laccase-like multicopper oxidases expressed in Escherichia coli.

    PubMed

    Ihssen, Julian; Reiss, Renate; Luchsinger, Ronny; Thöny-Meyer, Linda; Richter, Michael

    2015-06-12

    Laccases are multi-copper oxidases that oxidize a broad range of substrates at the expense of molecular oxygen, without any need for co-factor regeneration. These enzymes bear high potential for the sustainable synthesis of fine chemicals and the modification of (bio)polymers. Here we describe cloning and expression of five novel bacterial laccase-like multi copper oxidases (LMCOs) of diverse origin which were identified by homology searches in online databases. Activity yields under different expression conditions and temperature stabilities were compared to three previously described enzymes from Bacillus subtilis, Bacillus pumilus and Bacillus clausii. In almost all cases, a switch to oxygen-limited growth conditions after induction increased volumetric activity considerably. For proteins with predicted signal peptides for secretion, recombinant expression with and without signal sequence was investigated. Bacillus CotA-type LMCOs outperformed enzymes from Streptomyces and Gram-negative bacteria with respect to activity yields in Escherichia coli and application relevant biochemical properties. The novel Bacillus coagulans LMCO combined high activity yields in E. coli with unprecedented activity at strong alkaline pH and high storage stability, making it a promising candidate for further development.

  3. Escherichia coli survival in waters: temperature dependence.

    PubMed

    Blaustein, R A; Pachepsky, Y; Hill, R L; Shelton, D R; Whelan, G

    2013-02-01

    Knowing the survival rates of water-borne Escherichia coli is important in evaluating microbial contamination and making appropriate management decisions. E. coli survival rates are dependent on temperature, a dependency that is routinely expressed using an analogue of the Q₁₀ model. This suggestion was made 34 years ago based on 20 survival curves taken from published literature, but has not been revisited since then. The objective of this study was to re-evaluate the accuracy of the Q₁₀ equation, utilizing data accumulated since 1978. We assembled a database of 450 E. coli survival datasets from 70 peer-reviewed papers. We then focused on the 170 curves taken from experiments that were performed in the laboratory under dark conditions to exclude the effects of sunlight and other field factors that could cause additional variability in results. All datasets were tabulated dependencies "log concentration vs. time." There were three major patterns of inactivation: about half of the datasets had a section of fast log-linear inactivation followed by a section of slow log-linear inactivation; about a quarter of the datasets had a lag period followed by log-linear inactivation; and the remaining quarter were approximately linear throughout. First-order inactivation rate constants were calculated from the linear sections of all survival curves and the data grouped by water sources, including waters of agricultural origin, pristine water sources, groundwater and wells, lakes and reservoirs, rivers and streams, estuaries and seawater, and wastewater. Dependency of E. coli inactivation rates on temperature varied among the water sources. There was a significant difference in inactivation rate values at the reference temperature between rivers and agricultural waters, wastewaters and agricultural waters, rivers and lakes, and wastewater and lakes. At specific sites, the Q₁₀ equation was more accurate in rivers and coastal waters than in lakes making the value of

  4. Polymorphisms in the umuDC region of Escherichia species. [Escherichia coli; Escherichia alkalescens; Escherichia dispar; Escherichia aurescens

    SciTech Connect

    Sedgwick, S.G.; Robson, M.; Malik, F.

    1988-04-01

    The umuDC operon of Escherichia coli encodes mutagenic DNA repair. The umuDC regions of multiple isolates of E. coli, E. alkalescens, and E. dispar and a single stock of E. aurescens were mapped by nucleotide hybridization. umuDC is located at one end of a conserved tract of restriction endonuclease sites either 12.5 or 14 kilobase pairs long. Rearrangements, including possible deletions, were seen in the polymorphic DNA flanking the conserved tract. Restriction site polymorphisms were not found around the DNA repair gene recA or polA. The junctions of the conserved region contain direct repeats of nucleotide sequences resembling the termini of the Tn3 group of transposons. Possible mechanisms for the generation of these variants are discussed.

  5. Genome Sequence of Escherichia coli Tailed Phage Utah

    PubMed Central

    Leavitt, Justin C.; Heitkamp, Alexandra J.; Bhattacharjee, Ananda S.; Gilcrease, Eddie B.

    2017-01-01

    ABSTRACT Escherichia coli bacteriophage Utah is a member of the chi-like tailed phage cluster in the Siphoviridae family. We report here the complete 59,024-bp sequence of the genome of phage Utah. PMID:28360173

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

    EPA Science Inventory

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

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

    EPA Science Inventory

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

  8. Inhibition of Thiamine Transport by Chloroethylthiamine in Escherichia coli

    PubMed Central

    Iwashima, Akio; Nose, Yoshitsugu

    1972-01-01

    Chloroethylthiamine was found to inhibit an entrapment of thiamine as thiamine monophosphate by blocking thiamine monophosphokinase in the cytoplasm after thiamine was taken up by the cells of Escherichia coli. PMID:4565550

  9. Overexpression of vsr in Escherichia coli is mutagenic.

    PubMed

    Doiron, K M; Viau, S; Koutroumanis, M; Cupples, C G

    1996-07-01

    Overexpression of vsr in Escherichia coli stimulates transition and frameshift mutations. The pattern of mutations suggests that mutagenesis is due to saturation or inactivation of dam-directed mismatch repair.

  10. Shigella strains are not clones of Escherichia coli but sister species in the genus Escherichia.

    PubMed

    Zuo, Guanghong; Xu, Zhao; Hao, Bailin

    2013-02-01

    Shigella species and Escherichia coli are closely related organisms. Early phenotyping experiments and several recent molecular studies put Shigella within the species E. coli. However, the whole-genome-based, alignment-free and parameter-free CVTree approach shows convincingly that four established Shigella species, Shigella boydii, Shigella sonnei, Shigella felxneri and Shigella dysenteriae, are distinct from E. coli strains, and form sister species to E. coli within the genus Escherichia. In view of the overall success and high resolution power of the CVTree approach, this result should be taken seriously. We hope that the present report may promote further in-depth study of the Shigella-E. coli relationship.

  11. Infected hepatic Echinococcus cyst presenting as recurrent Escherichia coli empyema.

    PubMed

    Chang, R; Higgins, M; DiLisio, R; Hawasli, A; Camaro, L G; Khatib, R

    1993-03-01

    An 81-year-old man, previously a shepherd in Italy, presented with recurrent Escherichia coli empyema over an 8-month period. His empyema was caused by an infected, nonviable hepatic Echinococcus cyst that eroded the diaphragm and led to intermittent spillage and pleural seeding. This case demonstrates that when dealing with Escherichia coli empyema, a subdiaphragmatic source ought to be suspected, and among immigrants from areas with prevalent hydatid disease, infected hepatic Echinococcus cyst might rarely be the cause.

  12. ENERGY REQUIREMENT FOR THYMINELESS DEATH IN CELLS OF ESCHERICHIA COLI.

    PubMed

    FREIFELDER, D; MAALOE, O

    1964-10-01

    Freifelder, David (University of California, Berkeley), and Ole Maaløe. Energy requirement for thymineless death in cells of Escherichia coli. J. Bacteriol. 88:987-990. 1964.-Thymineless death in thymine-requiring Escherichia coli is arrested immediately and reversibly by nitrogenation if the bacterial population is growing in a medium containing a carbon source that can only be metabolized aerobically. The mechanism of death, therefore, involves a metabolic process.

  13. ENERGY REQUIREMENT FOR THYMINELESS DEATH IN CELLS OF ESCHERICHIA COLI

    PubMed Central

    Freifelder, David; Maaløe, Ole

    1964-01-01

    Freifelder, David (University of California, Berkeley), and Ole Maaløe. Energy requirement for thymineless death in cells of Escherichia coli. J. Bacteriol. 88:987–990. 1964.—Thymineless death in thymine-requiring Escherichia coli is arrested immediately and reversibly by nitrogenation if the bacterial population is growing in a medium containing a carbon source that can only be metabolized aerobically. The mechanism of death, therefore, involves a metabolic process. PMID:14219063

  14. The Escherichia coli Peripheral Inner Membrane Proteome*

    PubMed Central

    Papanastasiou, Malvina; Orfanoudaki, Georgia; Koukaki, Marina; Kountourakis, Nikos; Sardis, Marios Frantzeskos; Aivaliotis, Michalis; Karamanou, Spyridoula; Economou, Anastassios

    2013-01-01

    Biological membranes are essential for cell viability. Their functional characteristics strongly depend on their protein content, which consists of transmembrane (integral) and peripherally associated membrane proteins. Both integral and peripheral inner membrane proteins mediate a plethora of biological processes. Whereas transmembrane proteins have characteristic hydrophobic stretches and can be predicted using bioinformatics approaches, peripheral inner membrane proteins are hydrophilic, exist in equilibria with soluble pools, and carry no discernible membrane targeting signals. We experimentally determined the cytoplasmic peripheral inner membrane proteome of the model organism Escherichia coli using a multidisciplinary approach. Initially, we extensively re-annotated the theoretical proteome regarding subcellular localization using literature searches, manual curation, and multi-combinatorial bioinformatics searches of the available databases. Next we used sequential biochemical fractionations coupled to direct identification of individual proteins and protein complexes using high resolution mass spectrometry. We determined that the proposed cytoplasmic peripheral inner membrane proteome occupies a previously unsuspected ∼19% of the basic E. coli BL21(DE3) proteome, and the detected peripheral inner membrane proteome occupies ∼25% of the estimated expressed proteome of this cell grown in LB medium to mid-log phase. This value might increase when fleeting interactions, not studied here, are taken into account. Several proteins previously regarded as exclusively cytoplasmic bind membranes avidly. Many of these proteins are organized in functional or/and structural oligomeric complexes that bind to the membrane with multiple interactions. Identified proteins cover the full spectrum of biological activities, and more than half of them are essential. Our data suggest that the cytoplasmic proteome displays remarkably dynamic and extensive communication with

  15. Microdiesel: Escherichia coli engineered for fuel production.

    PubMed

    Kalscheuer, Rainer; Stölting, Torsten; Steinbüchel, Alexander

    2006-09-01

    Biodiesel is an alternative energy source and a substitute for petroleum-based diesel fuel. It is produced from renewable biomass by transesterification of triacylglycerols from plant oils, yielding monoalkyl esters of long-chain fatty acids with short-chain alcohols such as fatty acid methyl esters and fatty acid ethyl esters (FAEEs). Despite numerous environmental benefits, a broader use of biodiesel is hampered by the extensive acreage required for sufficient production of oilseed crops. Therefore, processes are urgently needed to enable biodiesel production from more readily available bulk plant materials like sugars or cellulose. Toward this goal, the authors established biosynthesis of biodiesel-adequate FAEEs, referred to as Microdiesel, in metabolically engineered Escherichia coli. This was achieved by heterologous expression in E. coli of the Zymomonas mobilis pyruvate decarboxylase and alcohol dehydrogenase and the unspecific acyltransferase from Acinetobacter baylyi strain ADP1. By this approach, ethanol formation was combined with subsequent esterification of the ethanol with the acyl moieties of coenzyme A thioesters of fatty acids if the cells were cultivated under aerobic conditions in the presence of glucose and oleic acid. Ethyl oleate was the major constituent of these FAEEs, with minor amounts of ethyl palmitate and ethyl palmitoleate. FAEE concentrations of 1.28 g l(-1) and a FAEE content of the cells of 26 % of the cellular dry mass were achieved by fed-batch fermentation using renewable carbon sources. This novel approach might pave the way for industrial production of biodiesel equivalents from renewable resources by employing engineered micro-organisms, enabling a broader use of biodiesel-like fuels in the future.

  16. Annual Surveillance Summary: Escherichia coli (E. coli) Infections in the Military Health System (MHS), 2016

    DTIC Science & Technology

    2017-06-30

    women.5 Screening practices may also contribute to higher rates of E. coli infections among females of reproductive age, as the Infectious Disease...Annual Surveillance Summary: Escherichia coli (E. coli) Infections in the Military Health System (MHS...and prevalence among all beneficiaries seeking care within the Military Health System (MHS). This report describes demographics, clinical

  17. 21 CFR 866.3255 - Escherichia coli serological reagents.

    Code of Federal Regulations, 2011 CFR

    2011-04-01

    ... 21 Food and Drugs 8 2011-04-01 2011-04-01 false Escherichia coli serological reagents. 866.3255 Section 866.3255 Food and Drugs FOOD AND DRUG ADMINISTRATION, DEPARTMENT OF HEALTH AND HUMAN SERVICES (CONTINUED) MEDICAL DEVICES IMMUNOLOGY AND MICROBIOLOGY DEVICES Serological Reagents § 866.3255 Escherichia...

  18. 21 CFR 866.3255 - Escherichia coli serological reagents.

    Code of Federal Regulations, 2010 CFR

    2010-04-01

    ... 21 Food and Drugs 8 2010-04-01 2010-04-01 false Escherichia coli serological reagents. 866.3255 Section 866.3255 Food and Drugs FOOD AND DRUG ADMINISTRATION, DEPARTMENT OF HEALTH AND HUMAN SERVICES (CONTINUED) MEDICAL DEVICES IMMUNOLOGY AND MICROBIOLOGY DEVICES Serological Reagents § 866.3255 Escherichia...

  19. Biocontrol of Escherichia coli O157

    PubMed Central

    Boyacioglu, Olcay; Sharma, Manan; Sulakvelidze, Alexander; Goktepe, Ipek

    2013-01-01

    The effect of a bacteriophage cocktail (EcoShield™) that is specific against Escherichia coli O157:H7 was evaluated against a nalidixic acid-resistant enterohemorrhagic E. coli O157:H7 RM4407 (EHEC) strain on leafy greens stored under either (1) ambient air or (2) modified atmosphere (MA; 5% O2/35% CO2/60% N2). Pieces (~2 × 2 cm2) of leafy greens (lettuce and spinach) inoculated with 4.5 log CFU/cm2 EHEC were sprayed with EcoShield™ (6.5 log PFU/cm2). Samples were stored at 4 or 10°C for up to 15 d. On spinach, the level of EHEC declined by 2.38 and 2.49 log CFU/cm2 at 4 and 10°C, respectively, 30 min after phage application (p ≤ 0.05). EcoShield™ was also effective in reducing EHEC on the surface of green leaf lettuce stored at 4°C by 2.49 and 3.28 log units in 30 min and 2 h, respectively (p ≤ 0.05). At 4°C under atmospheric air, the phage cocktail significantly (p ≤ 0.05) lowered the EHEC counts in one day by 1.19, 3.21 and 3.25 log CFU/cm2 on spinach, green leaf and romaine lettuce, respectively compared with control (no bacteriophage) treatments. When stored under MA at 4°C, phages reduced (p ≤ 0.05) EHEC populations by 2.18, 3.50 and 3.13 log CFU/cm2, on spinach, green leaf and romaine lettuce. At 10°C, EHEC reductions under atmospheric air storage were 1.99, 3.90 and 3.99 log CFU/cm2 (p ≤ 0.05), while population reductions under MA were 3.08, 3.89 and 4.34 logs on spinach, green leaf and romaine lettuce, respectively, compared with controls (p ≤ 0.05). The results of this study showed that bacteriophages were effective in reducing the levels of E. coli O157:H7 on fresh leafy produce, and that the reduction was further improved when produce was stored under the MA conditions. PMID:23819107

  20. Oligosaccharide Binding in Escherichia coli Glycogen Synthase

    SciTech Connect

    Sheng, Fang; Yep, Alejandra; Feng, Lei; Preiss, Jack; Geiger, James H.

    2010-11-17

    Glycogen/starch synthase elongates glucan chains and is the key enzyme in the synthesis of glycogen in bacteria and starch in plants. Cocrystallization of Escherichia coli wild-type glycogen synthase (GS) with substrate ADPGlc and the glucan acceptor mimic HEPPSO produced a closed form of GS and suggests that domain-domain closure accompanies glycogen synthesis. Cocrystallization of the inactive GS mutant E377A with substrate ADPGlc and oligosaccharide results in the first oligosaccharide-bound glycogen synthase structure. Four bound oligosaccharides are observed, one in the interdomain cleft (G6a) and three on the N-terminal domain surface (G6b, G6c, and G6d). Extending from the center of the enzyme to the interdomain cleft opening, G6a mostly interacts with the highly conserved N-terminal domain residues lining the cleft of GS. The surface-bound oligosaccharides G6c and G6d have less interaction with enzyme and exhibit a more curled, helixlike structural arrangement. The observation that oligosaccharides bind only to the N-terminal domain of GS suggests that glycogen in vivo probably binds to only one side of the enzyme to ensure unencumbered interdomain movement, which is required for efficient, continuous glucan-chain synthesis.

  1. Endonuclease IV (nfo) mutant of Escherichia coli.

    PubMed Central

    Cunningham, R P; Saporito, S M; Spitzer, S G; Weiss, B

    1986-01-01

    A cloned gene, designated nfo, caused overproduction of an EDTA-resistant endonuclease specific for apurinic-apyrimidinic sites in DNA. The sedimentation coefficient of the enzyme was similar to that of endonuclease IV. An insertion mutation was constructed in vitro and transferred from a plasmid to the Escherichia coli chromosome. nfo mutants had an increased sensitivity to the alkylating agents methyl methanesulfonate and mitomycin C and to the oxidants tert-butyl hydroperoxide and bleomycin. The nfo mutation enhanced the killing of xth (exonuclease III) mutants by methyl methanesulfonate, H2O2, tert-butyl hydroperoxide, and gamma rays, and it enhanced their mutability by methyl methanesulfonate. It also increased the temperature sensitivity of an xth dut (dUTPase) mutant that is defective in the repair of uracil-containing DNA. These results are consistent with earlier findings that endonuclease IV and exonuclease III both cleave DNA 5' to an apurinic-apyrimidinic site and that exonuclease III is more active. However, nfo mutants were more sensitive to tert-butyl hydroperoxide and to bleomycin than were xth mutants, suggesting that endonuclease IV might recognize some lesions that exonuclease III does not. The mutants displayed no marked increase in sensitivity to 254-nm UV radiation, and the addition of an nth (endonuclease III) mutation to nfo or nfo xth mutants did not significantly increase their sensitivity to any of the agents tested. Images PMID:2430946

  2. Completion of DNA replication in Escherichia coli.

    PubMed

    Wendel, Brian M; Courcelle, Charmain T; Courcelle, Justin

    2014-11-18

    The mechanism by which cells recognize and complete replicated regions at their precise doubling point must be remarkably efficient, occurring thousands of times per cell division along the chromosomes of humans. However, this process remains poorly understood. Here we show that, in Escherichia coli, the completion of replication involves an enzymatic system that effectively counts pairs and limits cellular replication to its doubling point by allowing converging replication forks to transiently continue through the doubling point before the excess, over-replicated regions are incised, resected, and joined. Completion requires RecBCD and involves several proteins associated with repairing double-strand breaks including, ExoI, SbcDC, and RecG. However, unlike double-strand break repair, completion occurs independently of homologous recombination and RecA. In some bacterial viruses, the completion mechanism is specifically targeted for inactivation to allow over-replication to occur during lytic replication. The results suggest that a primary cause of genomic instabilities in many double-strand-break-repair mutants arises from an impaired ability to complete replication, independent from DNA damage.

  3. 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

  4. The eclipse period of Escherichia coli

    PubMed Central

    von Freiesleben, Ulrik; Krekling, Martin A.; Hansen, Flemming G.; Løbner-Olesen, Anders

    2000-01-01

    The minimal time between successive initiations on the same origin (the eclipse) in Escherichia coli was determined to be ∼25–30 min. An inverse relationship was found between the length of the eclipse and the amount of Dam methyltransferase in the cell, indicating that the eclipse corresponds to the period of origin hemimethylation. The SeqA protein was absolutely required for the eclipse, and DnaA titration studies suggested that the SeqA protein prevented the binding of multiple DnaA molecules on oriC (initial complex formation). No correlation between the amount of SeqA and eclipse length was revealed, but increased SeqA levels affected chromosome partitioning and/or cell division. This was corroborated further by an aberrant nucleoid distribution in SeqA-deficient cells. We suggest that the SeqA protein’s role in maintaining the eclipse is tied to a function in chromosome organization. PMID:11080169

  5. Biochemistry of homologous recombination in Escherichia coli.

    PubMed Central

    Kowalczykowski, S C; Dixon, D A; Eggleston, A K; Lauder, S D; Rehrauer, W M

    1994-01-01

    Homologous recombination is a fundamental biological process. Biochemical understanding of this process is most advanced for Escherichia coli. At least 25 gene products are involved in promoting genetic exchange. At present, this includes the RecA, RecBCD (exonuclease V), RecE (exonuclease VIII), RecF, RecG, RecJ, RecN, RecOR, RecQ, RecT, RuvAB, RuvC, SbcCD, and SSB proteins, as well as DNA polymerase I, DNA gyrase, DNA topoisomerase I, DNA ligase, and DNA helicases. The activities displayed by these enzymes include homologous DNA pairing and strand exchange, helicase, branch migration, Holliday junction binding and cleavage, nuclease, ATPase, topoisomerase, DNA binding, ATP binding, polymerase, and ligase, and, collectively, they define biochemical events that are essential for efficient recombination. In addition to these needed proteins, a cis-acting recombination hot spot known as Chi (chi: 5'-GCTGGTGG-3') plays a crucial regulatory function. The biochemical steps that comprise homologous recombination can be formally divided into four parts: (i) processing of DNA molecules into suitable recombination substrates, (ii) homologous pairing of the DNA partners and the exchange of DNA strands, (iii) extension of the nascent DNA heteroduplex; and (iv) resolution of the resulting crossover structure. This review focuses on the biochemical mechanisms underlying these steps, with particular emphases on the activities of the proteins involved and on the integration of these activities into likely biochemical pathways for recombination. Images PMID:7968921

  6. Regulation of alcohol fermentation by Escherichia coli

    SciTech Connect

    Clark, D.P.

    1986-03-01

    The purpose of this project is to elucidate the way in which the fermentative synthesis of ethanol is regulated in the facultative anaerobe Escherichia coli. Focus is on the two final steps in alcohol synthesis, which are catalyzed by alcohol dehydrogenase and acetaldehyde CoA dehydrogenase. We have isolated a series of mutations affecting the expression of these enzymes. Some of these mutations are in the structural genes for these enzymes; others affect the regulation of the adh operon. We have recently cloned the genes coding for these enzymes and are now studying the effect of multiple copies of the adh gene on fermentative growth and its regulation. A recently invented technique, proton suicide has allowed the selection of a variety of novel mutants affecting fermentation which are presently being characterized. We have isolated a comprehensive collection of operon fusions in which the lacZ structural gene is fused to promoters that are inactive aerobically but active anaerobically. Although these genes (like adh) are only expressed under anaerobic conditions, the level of induction varies from two-fold to nearly 100-fold. The nitrogen source, medium pH, nature of the buffer, presence of alternative electron acceptors (e.g., nitrate), and other factors exert a great effect on the expression of many of these genes. In the near future we will investigate control mechanisms common to the adh operon and other anaerobically regulated genes.

  7. Regulation of Glutamine Transport in Escherichia coli.

    PubMed Central

    Willis, R C; Iwata, K K; Furlong, C E

    1975-01-01

    The formation of the high-affinity (Km equal to 0.2 muM) L-glutamine transport system of Escherichia coli strain 7 (Lin) appears to be subject to the same major control as the glutamine synthetase (EC 6.3.1.2) of this gram-negative organism. Culture of cells under nitrogen-limited conditions provides maximum derepression of both the glutamine synthetase and the glutamine transport system. Nutritional conditions providing a rich supply of ammonium salts or available sources of nitrogen, i.e., conditions which repress the formation of glutamine synthetase, provide three- and 20-fold repression, respectively, of the glutamine transport system. Culture of cells with glutamine supplements of 2 mM does not increase the repression of high-affinity glutamine transport system beyond the level observed in the absence of glutamine. A second kinetically distinct low-affinity component of glutamine. A second kinetically distinct low-affinity component of glutamine uptake is observed in cells cultured with a glutamine-depleted nutrient broth. This second component is associated with the appearance of glutaminase A (EC 3.5.1.2) and asparaginase I (EC 3.5.1.1), a periplasmic enzyme. Parallel changes were observed in the levels of the high-affinity glutamine transport system and the glutamine synthetase when cells were cultured with the carbon sources: glucose, glycerol, or succinate. PMID:238938

  8. ESCHERICHIA COLI Gene Induction by Alkylation Treatment

    PubMed Central

    Volkert, Michael R.; Nguyen, Dinh C.; Beard, K. Christopher

    1986-01-01

    Searches for alkylation-inducible (aid) genes of Escherichia coli have been conducted by screening random fusions of the Mu-dl(ApR lac) phage for fusions showing increased β-galactosidase activity after treatment with methylating agents, but not after treatments with UV-irradiation. In this report we describe gene fusions that are specifically induced by alkylation treatments. Nine new mutants are described, and their properties are compared with the five mutants described previously. The total of 14 fusion mutants map at five distinct genetic loci. They can be further subdivided on the basis of their induction by methyl methanesulfonate (MMS) and N-methyl-N'-nitro-N-nitrosoguanidine (MNNG). alkA, aidB and aidD are induced by both agents and appear to be regulated by ada. Neither aidC nor aidI is regulated by ada. Moreover, since aidC is induced only by MNNG and aidI is induced only by MMS, these two genes are likely to be individually regulated. Thus, there appear to be at least three different regulatory mechanisms controlling aid genes. PMID:3080354

  9. Escherichia coli gene induction by alkylation treatment.

    PubMed

    Volkert, M R; Nguyen, D C; Beard, K C

    1986-01-01

    Searches for alkylation-inducible (aid) genes of Escherichia coli have been conducted by screening random fusions of the Mu-dl(ApR lac) phage for fusions showing increased beta-galactosidase activity after treatment with methylating agents, but not after treatments with UV-irradiation. In this report we describe gene fusions that are specifically induced by alkylation treatments. Nine new mutants are described, and their properties are compared with the five mutants described previously. The total of 14 fusion mutants map at five distinct genetic loci. They can be further subdivided on the basis of their induction by methyl methanesulfonate (MMS) and N-methyl-N' -nitro-N-nitrosoguanidine (MNNG). alkA, aidB and aidD are induced by both agents and appear to be regulated by ada. Neither aidC nor aidI is regulated by ada. Moreover, since aidC is induced only by MNNG and aidI is induced only by MMS, these two genes are likely to be individually regulated. Thus, there appear to be at least three different regulatory mechanisms controlling aid genes.

  10. Antimicrobial-resistant Invasive Escherichia coli, Spain

    PubMed Central

    Oteo, Jesús; Lázaro, Edurne; de Abajo, Francisco J.; Baquero, Fernando; Campos, José

    2005-01-01

    To address the public health problem of antimicrobial resistance, the European Union founded the European Antimicrobial Resistance Surveillance System. A network of 32 Spanish hospitals, serving ≈9.6 million persons, submitted antimicrobial-susceptibility data on 7,098 invasive Escherichia coli species (2001–2003). Resistance to ampicillin, cotrimoxazole, ciprofloxacin, gentamicin, and tobramycin was found at rates of 59.9%, 32.6%, 19.3%, 6.8%, and 5.3%, respectively. Resistance to multiple drugs increased from 13.8% in 2001 to 20.6% in 2003 (p <0.0001). Antimicrobial consumption data were obtained from the Spanish National Health System. In spite of decreased cephalosporin and β-lactam use, overall extended-spectrum β-lactamase production increased from 1.6% (2001) to 4.1% (2003) (p <0.0001), mainly due to the rising prevalence of cefotaximases. Resistance to ciprofloxacin significantly increased, mostly in community-onset infections, which coincided with a rise in community quinolone use. Cotrimoxazole resistance remained stable at ≈30%, even though its use was dramatically reduced. PMID:15829192

  11. Ribonuclease Sensitivity of Escherichia coli Ribosomes

    PubMed Central

    Santer, Melvin; Smith, Josephine R.

    1966-01-01

    Santer, Melvin (Haverford College, Haverford, Pa.), and Josephine R. Smith. Ribonuclease sensitivity of Escherichia coli ribosomes. J. Bacteriol. 92:1099–1110. 1966.—The ribonucleic acid (RNA) contained in 70S ribosomes and in 50S and 30S subunits was hydrolyzed by pancreatic ribonuclease. A 7% amount of the RNA was removed from the 70S particle; at 10−4m magnesium concentration, a maximum of 24 and 30% of the RNA in the 50S and the 30S fractions, respectively, was removed by ribonuclease. At the two lower magnesium ion concentrations, 50S ribosomes did not lose any protein, whereas 30S ribosomes lost protein as a result of ribonuclease treatment. A number of proteins were removed from the 30S particles by ribonuclease, and these proteins were antigenically related to proteins present in 50S ribosomes. The differential effect of ribonuclease on 50S and 30S ribosomes suggested that they have structural dissimilarities. Images PMID:5332866

  12. Genotoxicity of Graphene in Escherichia coli

    NASA Astrophysics Data System (ADS)

    Sharma, Ananya

    Rapid advances in nanotechnology necessitate assessment of the safety of nanomaterials in the resulting products and applications. One key nanomaterial attracting much interest in many areas of science and technology is graphene. Graphene is a one atom thick carbon allotrope arranged in a two-dimensional honeycomb lattice. In addition to being extremely thin, graphene has several extraordinary physical properties such as its exceptional mechanical strength, thermal stability, and high electrical conductivity. Graphene itself is relatively chemically inert and therefore pristine graphene must undergo a process called functionalization, which is combination of chemical and physical treatments that change the properties of graphene, to make it chemically active. Functionalization of graphene is of crucial importance as the end application of graphene depends on proper functionalization. In the field of medicine, graphene is currently a nanomaterial of high interest for building biosensors, DNA transistors, and probes for cancer detection. Despite the promising applications of graphene in several areas of biomedicine, there have been only few studies in recent years that focus on evaluating cytotoxicity of graphene on cells, and almost no studies that investigate how graphene exposure affects cellular genetic material. Therefore, in this study we used a novel approach to evaluate the genotoxicity, i.e., the effects of graphene on DNA, using Escherichia coli as a prokaryotic model organism.

  13. The DNA exonucleases of Escherichia coli

    PubMed Central

    Lovett, Susan T.

    2014-01-01

    DNA exonucleases, enzymes that hydrolyze phosphodiester bonds in DNA from a free end, play important cellular roles in DNA repair, genetic recombination and mutation avoidance in all organisms. This article reviews the structure, biochemistry and biological functions of the 17 exonucleases currently identified in the bacterium Escherichia coli. These include the exonucleases associated with DNA polymerases I (polA), II (polB) and III (dnaQ/mutD), Exonucleases I (xonA/sbcB), III (xthA), IV, VII (xseAB), IX (xni/xgdG) and X (exoX), the RecBCD, RecJ, and RecE exonucleases, SbcCD endo/exonuclease, the DNA exonuclease activities of RNase T (rnt) and Endonuclease IV (nfo) and TatD. These enzymes are diverse in terms of substrate specificity and biochemical properties and have specialized biological roles. Most of these enzymes fall into structural families with characteristic sequence motifs, and members of many of these families can be found in all domains of life. PMID:26442508

  14. Stringency of substrate specificity of Escherichia coli malate dehydrogenase.

    SciTech Connect

    Boernke, W. E.; Millard, C. S.; Stevens, P. W.; Kakar, S. N.; Stevens, F. J.; Donnelly, M. I.; Nebraska Wesleyan Univ.

    1995-09-10

    Malate dehydrogenase and lactate dehydrogenase are members of the structurally and functionally homologous family of 2-ketoacid dehydrogenases. Both enzymes display high specificity for their respective keto substrates, oxaloacetate and pyruvate. Closer analysis of their specificity, however, reveals that the specificity of malate dehydrogenase is much stricter and less malleable than that of lactate dehydrogenase. Site-specific mutagenesis of the two enzymes in an attempt to reverse their specificity has met with contrary results. Conversion of a specific active-site glutamine to arginine in lactate dehydrogenase from Bacillus stearothermophilus generated an enzyme that displayed activity toward oxaloacetate equal to that of the native enzyme toward pyruvate (H. M. Wilks et al. (1988) Science 242, 1541-1544). We have constructed a series of mutants in the mobile, active site loop of the Escherichia coli malate dehydrogenase that incorporate the complementary change, conversion of arginine 81 to glutamine, to evaluate the role of charge distribution and conformational flexibility within this loop in defining the substrate specificity of these enzymes. Mutants incorporating the change R81Q all had reversed specificity, displaying much higher activity toward pyruvate than to the natural substrate, oxaloacetate. In contrast to the mutated lactate dehydrogenase, these reversed-specificity mutants were much less active than the native enzyme. Secondary mutations within the loop of the E. coli enzyme (A80N, A80P, A80P/M85E/D86T) had either no or only moderately beneficial effects on the activity of the mutant enzyme toward pyruvate. The mutation A80P, which can be expected to reduce the overall flexibility of the loop, modestly improved activity toward pyruvate. The possible physiological relevance of the stringent specificity of malate dehydrogenase was investigated. In normal strains of E. coli, fermentative metabolism was not affected by expression of the mutant

  15. Enterotoxigenic Escherichia coli Multilocus Sequence Types in Guatemala and Mexico

    PubMed Central

    Klena, John; Rodas, Claudia; Bourgeois, August Louis; Torres, Olga; Svennerholm, Ann-Mari; Sjöling, Åsa

    2010-01-01

    The genetic backgrounds of 24 enterotoxigenic Escherichia coli (ETEC) strains from Mexico and Guatemala expressing heat-stable toxin (ST) and coli surface antigen 6 (CS6) were analyzed. US travelers to these countries and resident children in Guatemala were infected by ETEC strains of sequence type 398, expressing STp and carrying genetically identical CS6 sequences. PMID:20031063

  16. Characterization of enterohemorrhagic Escherichia coli on veal hides and carcasses

    USDA-ARS?s Scientific Manuscript database

    Enterohemorrhagic E. coli (EHEC) are Shiga toxin–producing Escherichia coli (STEC) associated with the most severe forms of foodborne illnesses. The United States Department of Agriculture (USDA) Food Safety Inspection Service (FSIS) has identified a higher percentage of non-O157 EHEC compared to E....

  17. Escherichia coli Pathotypes Occupy Distinct Niches in the Mouse Intestine

    PubMed Central

    Meador, Jessica P.; Caldwell, Matthew E.; Cohen, Paul S.

    2014-01-01

    Since the first step of the infection process is colonization of the host, it is important to understand how Escherichia coli pathogens successfully colonize the intestine. We previously showed that enterohemorrhagic O157:H7 strain E. coli EDL933 colonizes a niche in the streptomycin-treated mouse intestine that is distinct from that of human commensal strains, which explains how E. coli EDL933 overcomes colonization resistance imparted by some, but not all, commensal E. coli strains. Here we sought to determine if other E. coli pathogens use a similar strategy. We found that uropathogenic E. coli CFT073 and enteropathogenic E. coli E2348/69 occupy intestinal niches that are distinct from that of E. coli EDL933. In contrast, two enterohemorrhagic strains, E. coli EDL933 and E. coli Sakai, occupy the same niche, suggesting that strategies to prevent colonization by a given pathotype should be effective against other strains of the same pathotype. However, we found that a combination of commensal E. coli strains that can prevent colonization by E. coli EDL933 did not prevent colonization by E. coli CFT073 or E. coli E2348/69. Our results indicate that development of probiotics to target multiple E. coli pathotypes will be problematic, as the factors that govern niche occupation and hence stable colonization vary significantly among strains. PMID:24566621

  18. Draft Genome Sequence of Uropathogenic Escherichia coli Strain NB8

    PubMed Central

    Mi, Zu-huang; Wang, Chun-xin; Zhu, Jian-ming

    2016-01-01

    Escherichia coli NB8 is a clinical pyelonephritis isolate. Here, we report the draft genome sequence of uropathogenic E. coli NB8, which contains drug resistance genes encoding resistance to beta-lactams, aminoglycosides, quinolones, macrolides, colistin, sulfonamide-trimethoprim, and tetracycline. NB8 infects the kidney and bladder, making it an important tool for studying E. coli pathogenesis. PMID:27609920

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

    PubMed

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

    2017-09-01

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

  20. Soil solarization reduces Escherichia coli O157:H7 and total Escherichia coli on cattle feedlot pen surfaces

    USDA-ARS?s Scientific Manuscript database

    Feedlot pen soils are a source for transmission of Escherichia coli O157:H7, and therefore a target for preharvest strategies to reduce this pathogen in cattle. The objective of this study was to determine the ability of soil solarization to reduce E. coli O157:H7 in feedlot surface material (FSM)....

  1. Free RNA polymerase in Escherichia coli.

    PubMed

    Patrick, Michael; Dennis, Patrick P; Ehrenberg, Mans; Bremer, Hans

    2015-12-01

    The frequencies of transcription initiation of regulated and constitutive genes depend on the concentration of free RNA polymerase holoenzyme [Rf] near their promoters. Although RNA polymerase is largely confined to the nucleoid, it is difficult to determine absolute concentrations of [Rf] at particular locations within the nucleoid structure. However, relative concentrations of free RNA polymerase at different growth rates, [Rf]rel, can be estimated from the activities of constitutive promoters. Previous studies indicated that the rrnB P2 promoter is constitutive and that [Rf]rel in the vicinity of rrnB P2 increases with increasing growth rate. Recently it has become possible to directly visualize Rf in growing Escherichia coli cells. Here we examine some of the important issues relating to gene expression based on these new observations. We conclude that: (i) At a growth rate of 2 doublings/h, there are about 1000 free and 2350 non-specifically DNA-bound RNA polymerase molecules per average cell (12 and 28%, respectively, of 8400 total) which are in rapid equilibrium. (ii) The reversibility of the non-specific binding generates more than 1000 free RNA polymerase molecules every second in the immediate vicinity of the DNA. Of these, most rebind non-specifically to the DNA within a few ms; the frequency of non-specific binding is at least two orders of magnitude greater than specific binding and transcript initiation. (iii) At a given amount of RNA polymerase per cell, [Rf] and the density of non-specifically DNA-bound RNA polymerase molecules along the DNA both vary reciprocally with the amount of DNA in the cell. (iv) At 2 doublings/h an E. coli cell contains, on the average, about 1 non-specifically bound RNA polymerase per 9 kbp of DNA and 1 free RNA polymerase per 20 kbp of DNA. However some DNA regions (i.e. near active rRNA operons) may have significantly higher than average [Rf].

  2. The Melibiose Transporter of Escherichia coli

    PubMed Central

    Fuerst, Oliver; Lin, Yibin; Granell, Meritxell; Leblanc, Gérard; Padrós, Esteve; Lórenz-Fonfría, Víctor A.; Cladera, Josep

    2015-01-01

    We examine the role of Lys-377, the only charged residue in helix XI, on the functional mechanism of the Na+-sugar melibiose symporter from Escherichia coli. Intrinsic fluorescence, FRET, and Fourier transform infrared difference spectroscopy reveal that replacement of Lys-377 with either Cys, Val, Arg, or Asp disables both Na+ and melibiose binding. On the other hand, molecular dynamics simulations extending up to 200–330 ns reveal that Lys-377 (helix XI) interacts with the anionic side chains of two of the three putative ligands for cation binding (Asp-55 and Asp-59 in helix II). When Asp-59 is protonated during the simulations, Lys-377 preferentially interacts with Asp-55. Interestingly, when a Na+ ion is positioned in the Asp-55-Asp-59 environment, Asp-124 in helix IV (a residue essential for melibiose binding) reorients and approximates the Asp-55-Asp-59 pair, and all three acidic side chains act as Na+ ligands. Under these conditions, the side chain of Lys-377 interacts with the carboxylic moiety of these three Asp residues. These data highlight the crucial role of the Lys-377 residue in the spatial organization of the Na+ binding site. Finally, the analysis of the second-site revertants of K377C reveals that mutation of Ile-22 (in helix I) preserves Na+ binding, whereas that of melibiose is largely abolished according to spectroscopic measurements. This amino acid is located in the border of the sugar-binding site and might participate in sugar binding through apolar interactions. PMID:25971963

  3. Polyamine transport inEscherichia coli.

    PubMed

    Igarashi, K; Kashiwagi, K

    1996-03-01

    The polyamine content in cells is regulated by both polyamine biosynthesis and its transport. We recently obtained and characterized three clones of polyamine transport genes (pPT104, pPT79 and pPT71) inEscherichia coli. The system encoded by pPT104 was the spermidine-preferential uptake system and that encoded by pPT79 the putrescine-specific uptake system. Furthermore, these two systems were periplasmic transport systems consisting of four kinds of proteins: pPT104 clone encoded potA, -B,-C, and -D proteins and pPT79 clone encoded potF, -G, -H, and -I proteins, judging from the deduced amino acid sequences of the nucleotide sequences of these clones. PotD and -F proteins were periplasmic substrate binding proteins and potA and -G proteins membrane associated proteins having the nucleotide binding site. PotB and -C proteins, and potH and -I proteins were transmembrane proteins probably forming channels for spermidine and putrescine, respectively. Their amino acid sequences in the corresponding proteins were similar to each other. The functions of potA and -D proteins in the spermidine-preferential uptake system encoded by pPT104 clone were studied in detail through a combined biochemical and genetic approach. In contrast, the putrescine transport system encoded by pPT71 consisted of one membrane protein (potE protein) haveing twelve transmembrane segments, and was active in both the uptake and excretion of putrescine. The uptake was dependent on membrane potential, and the excretion was due to the exchange reaction between putrescine and ornithine.

  4. Novel Mechanism of Escherichia coli Porin Regulation

    PubMed Central

    Castillo-Keller, Maria; Vuong, Phu; Misra, Rajeev

    2006-01-01

    A novel mechanism of Escherichia coli porin regulation was discovered from multicopy suppressors that permitted growth of cells expressing a mutant OmpC protein in the absence of DegP. Analyses of two suppressors showed that both substantially lowered OmpC expression. Suppression activities were confined to a short DNA sequence, which we designated ipeX for inhibition of porin expression, and to DNA containing a 3′-truncated ompR gene. The major effect of ipeX on ompC expression was exerted posttranscriptionally, whereas the truncated OmpR protein reduced ompC transcription. ipeX was localized within an untranslated region of 247 base pairs between the stop codon of nmpC—a remnant porin gene from the cryptic phage qsr′ (DLP12) genome—and its predicted Rho-independent transcriptional terminator. Interestingly, another prophage, PA-2, which encodes a porin similar to NmpC, known as Lc, has sequences downstream from lc identical to that of ipeX. PA-2 lysogenization leads to Lc expression and OmpC inhibition. Our data show that the synthesis of the lc transcript, whose 3′ end contains the corresponding ipeX sequence, inhibits OmpC expression. Overexpression of ipeX RNA inhibited both OmpC and OmpF expression but not that of OmpA. ompC-phoA chimeric gene constructs revealed a 248-bp untranslated region of ompC required for ipeX-mediated inhibition. However, no sequence complementarity was found between ipeX and this region of ompC, indicating that inhibition may not involve simple base pairing between the two RNA molecules. The effect of ipeX on ompC, but not on ompF, was independent of the RNA chaperone Hfq. PMID:16385048

  5. Mutational Consequences of Ciprofloxacin in Escherichia coli.

    PubMed

    Song, Lisa Yun; Goff, Marisa; Davidian, Christina; Mao, Zhiyuan; London, Marisa; Lam, Karen; Yung, Madeline; Miller, Jeffrey H

    2016-10-01

    We examined the mutagenic specificity of the widely used antibiotic ciprofloxacin (CPR), which displays weak to moderate mutagenic activity in several bacteria and generates short in-frame deletions in rpoB in Staphylococcus aureus To determine the spectrum of mutations in a system where any gene knockout would result in a recovered mutant, including frameshifts and both short and long deletions, we examined CPR-induced mutations in the thymidylate synthase-encoding thyA gene. Here, any mutation resulting in loss of thymidylate synthase activity generates trimethoprim (Trm) resistance. We found that deletions and insertions in all three reading frames predominated in the spectrum. They tend to be short deletions and cluster in two regions, one being a GC-rich region with potential extensive secondary structures. We also exploited the well-characterized rpoB-Rif(r) system in Escherichia coli to determine that cells grown in the presence of sublethal doses of CPR not only induced short in-frame deletions in rpoB, but also generated base substitution mutations resulting from induction of the SOS system. Some of the specific point mutations prominent in the spectrum of a strain that overproduces the dinB-encoded Pol IV were also present after growth in CPR. However, these mutations disappeared in CPR-treated dinB mutants, whereas the deletions remained. Moreover, CPR-induced deletions also occurred in a strain lacking all three SOS-induced polymerases. We discuss the implications of these findings for the consequences of overuse of CPR and other antibiotics. Copyright © 2016, American Society for Microbiology. All Rights Reserved.

  6. Serogroups of Escherichia coli from drinking water.

    PubMed

    Ramteke, P W; Tewari, Suman

    2007-07-01

    Fifty seven isolates of thermotolerant E. coli were recovered from 188 drinking water sources, 45 (78.9%) were typable of which 15 (26.3%) were pathogenic serotypes. Pathogenic serogroup obtained were 04 (Uropathogenic E. coli, UPEC), 025 (Enterotoxigenic E. coli, ETEC), 086 (Enteropathogenic E. coli, EPEC), 0103 (Shiga-toxin producing E. coli, STEC), 0157 (Shiga-toxin producing E. coli, STEC), 08 (Enterotoxigenic E. coli, ETEC) and 0113 (Shiga-toxin producing E. coli, STEC). All the pathogenic serotypes showed resistance to bacitracin and multiple heavy metal ions. Resistance to streptomycin and cotrimazole was detected in two strains whereas resistance to cephaloridine, polymixin-B and ampicillin was detected in one strain each. Transfer of resistances to drugs and metallic ions was observed in 9 out of 12 strains studied. Resistances to bacitracin were transferred in all nine strains. Among heavy metals resistance to As(3+) followed by Cr(6+) were transferred more frequently.

  7. Control of Acid Resistance in Escherichia coli

    PubMed Central

    Castanie-Cornet, Marie-Pierre; Penfound, Thomas A.; Smith, Dean; Elliott, John F.; Foster, John W.

    1999-01-01

    Acid resistance (AR) in Escherichia coli is defined as the ability to withstand an acid challenge of pH 2.5 or less and is a trait generally restricted to stationary-phase cells. Earlier reports described three AR systems in E. coli. In the present study, the genetics and control of these three systems have been more clearly defined. Expression of the first AR system (designated the oxidative or glucose-repressed AR system) was previously shown to require the alternative sigma factor RpoS. Consistent with glucose repression, this system also proved to be dependent in many situations on the cyclic AMP receptor protein. The second AR system required the addition of arginine during pH 2.5 acid challenge, the structural gene for arginine decarboxylase (adiA), and the regulator cysB, confirming earlier reports. The third AR system required glutamate for protection at pH 2.5, one of two genes encoding glutamate decarboxylase (gadA or gadB), and the gene encoding the putative glutamate:γ-aminobutyric acid antiporter (gadC). Only one of the two glutamate decarboxylases was needed for protection at pH 2.5. However, survival at pH 2 required both glutamate decarboxylase isozymes. Stationary phase and acid pH regulation of the gad genes proved separable. Stationary-phase induction of gadA and gadB required the alternative sigma factor ςS encoded by rpoS. However, acid induction of these enzymes, which was demonstrated to occur in exponential- and stationary-phase cells, proved to be ςS independent. Neither gad gene required the presence of volatile fatty acids for induction. The data also indicate that AR via the amino acid decarboxylase systems requires more than an inducible decarboxylase and antiporter. Another surprising finding was that the ςS-dependent oxidative system, originally thought to be acid induced, actually proved to be induced following entry into stationary phase regardless of the pH. However, an inhibitor produced at pH 8 somehow interferes with the

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

    USGS Publications Warehouse

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

    2017-01-01

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

  9. Current pathogenic Escherichia coli foodborne outbreak cases and therapy development.

    PubMed

    Yang, Shih-Chun; Lin, Chih-Hung; Aljuffali, Ibrahim A; Fang, Jia-You

    2017-08-01

    Food contamination by pathogenic microorganisms has been a serious public health problem and a cause of huge economic losses worldwide. Foodborne pathogenic Escherichia coli (E. coli) contamination, such as that with E. coli O157 and O104, is very common, even in developed countries. Bacterial contamination may occur during any of the steps in the farm-to-table continuum from environmental, animal, or human sources and cause foodborne illness. To understand the causes of the foodborne outbreaks by E. coli and food-contamination prevention measures, we collected and investigated the past 10 years' worldwide reports of foodborne E. coli contamination cases. In the first half of this review article, we introduce the infection and symptoms of five major foodborne diarrheagenic E. coli pathotypes: enteropathogenic E. coli (EPEC), Shiga toxin-producing E. coli/enterohemorrhagic E. coli (STEC/EHEC), Shigella/enteroinvasive E. coli (EIEC), enteroaggregative E. coli (EAEC), and enterotoxigenic E. coli (ETEC). In the second half of this review article, we introduce the foodborne outbreak cases caused by E. coli in natural foods and food products. Finally, we discuss current developments that can be applied to control and prevent bacterial food contamination.

  10. Investigation of ’Escherichia coli’ Enterotoxins

    DTIC Science & Technology

    1978-05-01

    E . coli diarrheal disease in man and domestic animals. Fundamentally, the design of the vaccine is based on the well- documented ability of cholera antitoxin to neutralize both cholera and heat- labile E . coli enterotoxins and on the ability of certain E . coli antigens to enhance the immune response to cholera toxoid and possibly whole-cell Cholera Vaccine, as

  11. Rapid Sterilization of Escherichia coli by Solution Plasma Process

    NASA Astrophysics Data System (ADS)

    Andreeva, Nina; Ishizaki, Takahiro; Baroch, Pavel; Saito, Nagahiro

    2012-12-01

    Solution plasma (SP), which is a discharge in the liquid phase, has the potential for rapid sterilization of water without chemical agents. The discharge showed a strong sterilization performance against Escherichia coli bacteria. The decimal value (D value) of the reduction time for E. coli by this system with an electrode distance of 1.0 mm was estimated to be approximately 1.0 min. Our discharge system in the liquid phase caused no physical damage to the E. coli and only a small increase in the temperature of the aqueous solution. The UV light generated by the discharge was an important factor in the sterilization of E. coli.

  12. Human Meningitis-Associated Escherichia coli

    PubMed Central

    KIM, KWANG SIK

    2016-01-01

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

  13. [Frequency, risk factors and vaginal colonization due to Escherichia coli].

    PubMed

    González Pedraza Avilés, Alberto; Sánchez Hernández, Gabriela; Ponce Rosas, Raúl Efrén

    2004-02-01

    Recent studies associate Escherichia coli with symptomatic infections at vaginal level, mainly associated to changes in the normal flora taken place by a series of factors characteristic of the host. To recognize their colonization frequency and these factors, it becomes important due to their association with perinatal complications, besides considering this colonization like the critical step preceding urinary tract infection. To determine the frequency of colonization of Escherichia coli in 519 female patients, the role of the bacterium in the vaginal ecology likes probable cause of clinical manifestations and to recognize the associate's factors of risk with its vaginal colonization. 519 women were studied: 350 symptomatic and 169 asymptomatic. Vaginal swab specimens were inoculated onto the routine mediums. Associations of Escherichia coli with various risk factors were examined by using odds ratios (ORs) and 95% confidence intervals, and statistical significance was assessed by the Chi statistic or Fischer's exact test. Overall Escherichia coli was isolated from 95 (18.3%) of the women. Factors that were significantly associated with vaginal carriage of E. coli were the age extreme groups, the climacteric, and the bad genital habits. The highest frequency of vaginal colonization for Escherichia coli was presented in the population groups where there is hormonal deficiency, mainly of estrogens of the type estradiol. The vaginal colonization for E. coli doesn't associate to sexual behavior. Although E. coli doesn't produce defined symptoms at vaginal level, the relatively low carriage rate indicates that this organism should not be considered as part of the normal indigenous vaginal flora and that it should take into account due to the perinatal complication it is associated.

  14. The Biology of the Escherichia coli Extracellular Matrix

    PubMed Central

    Hufnagel, David A.; DePas, William H.; Chapman, Matthew R.

    2015-01-01

    Chapter Summary Escherichia coli (E. coli) is one of the world’s best-characterized organisms, as it has been extensively studied for over a century. However, most of this work has focused on E. coli grown under laboratory conditions that do not faithfully simulate its natural environments. Therefore, the historical perspectives on E. coli physiology and life cycle are somewhat skewed toward experimental systems that feature E. coli growing logarithmically in a test tube. Typically a commensal bacterium, E. coli resides in the lower intestines of a slew of animals. Outside of the lower intestine, E. coli can adapt and survive in a very different set of environmental conditions. Biofilm formation allows E. coli to survive, and even thrive, in environments that do not support the growth of planktonic populations. E. coli can form biofilms virtually everywhere; in the bladder during a urinary tract infection, on in-dwelling medical devices, and outside of the host on plants and in the soil. The E. coli extracellular matrix, primarily composed of the protein polymer named curli and the polysaccharide cellulose, promotes adherence to organic and inorganic surfaces, and resistance to desiccation, the host immune system and other antimicrobials. The pathways that govern E. coli biofilm formation, cellulose production, and curli biogenesis will be discussed in this book chapter, which concludes with insights into the future of E. coli biofilm research and potential therapies. PMID:26185090

  15. 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

  16. Short-Chain Chromate Ion Transporter Proteins from Bacillus subtilis Confer Chromate Resistance in Escherichia coli▿

    PubMed Central

    Díaz-Magaña, Amada; Aguilar-Barajas, Esther; Moreno-Sánchez, Rafael; Ramírez-Díaz, Martha I.; Riveros-Rosas, Héctor; Vargas, Eréndira; Cervantes, Carlos

    2009-01-01

    Tandem paired genes encoding putative short-chain monodomain protein members of the chromate ion transporter (CHR) superfamily (ywrB and ywrA) were cloned from genomic DNA of Bacillus subtilis strain 168. The transcription of the paired genes, renamed chr3N and chr3C, respectively, was shown to occur via a bicistronic mRNA generated from a promoter upstream of the chr3N gene. The chr3N and chr3C genes conferred chromate resistance when expressed in Escherichia coli strain W3110. The cloned chr3N gene alone did not confer chromate resistance on E. coli, suggesting that both chr3N and chr3C genes are required for function. E. coli cells expressing paired chr3N and chr3C genes demonstrated diminished uptake of chromate compared to that by a vector-only control strain. These results suggest that short-chain CHR proteins form heterodimer transporters which efflux chromate ions from the cytoplasm. PMID:19581367

  17. Intestinal Colonization by Enterotoxigenic Escherichia coli.

    DTIC Science & Technology

    1980-09-01

    E . coli is mediated by specific types of pili. These pili are antigenic and can be used in diagnosing enterotoxigenic E . coli infections. They are also good protective antigens. When pregnant dams are vaccinated parenterally or orally with pili on live piliated bacteria, they secrete antibodies against the pili in their milk. Neonates suckling dams so vaccinated are passively protected against fatal challenge by enterotoxigenic E . coli . Pili are also good candidate protective antigens for the development of vaccines to protect by

  18. Transcription of foreign DNA in Escherichia coli.

    PubMed

    Warren, René L; Freeman, John D; Levesque, Roger C; Smailus, Duane E; Flibotte, Stephane; Holt, Robert A

    2008-11-01

    Propagation of heterologous DNA in E. coli host cells is central to molecular biology. DNA constructs are often engineered for expression of recombinant protein in E. coli, but the extent of incidental transcription arising from natural regulatory sequences in cloned DNA remains underexplored. Here, we have used programmable microarrays and RT-PCR to measure, comprehensively, the transcription of H. influenzae, P. aeruginosa, and human DNA propagating in E. coli as bacterial artificial chromosomes. We find evidence that at least half of all H. influenzae genes are transcribed in E. coli. Highly transcribed genes are principally involved in energy metabolism, and their proximal promoter regions are significantly enriched with E. coli sigma(70) (also known as RpoD) binding sites. H. influenzae genes acquired from an ancient bacteriophage Mu insertion are also highly transcribed. Compared with H. influenzae, a smaller proportion of P. aeruginosa genes are transcribed in E. coli, and in E. coli there is punctuated transcription of human DNA. The presence of foreign DNA in E. coli disturbs the host transcriptional profile, with expression of the E. coli phage shock protein operon and the flagellar gene cluster being particularly strongly up-regulated. While cross-species transcriptional activation is expected to be enabling for horizontal gene transfer in bacteria, incidental expression of toxic genes can be problematic for DNA cloning. Ongoing characterization of cross-expression will help inform the design of biosynthetic gene clusters and synthetic microbial genomes.

  19. [Expression of Photobacterium leiognathi bioluminescence system genes in Escherichia coli].

    PubMed

    Ptitsyn, L R; Fatova, M A; Stepanov, A I

    1990-02-01

    Expression of Photobacterium leiognathi bioluminescence genes under the control of lac, tac, tet promoters in Escherichia coli cells has been studied. The position of the genes for aliphatic aldehyde biosynthesis and for the synthesis of luciferase subunits was identified. The plasmid pBRPL1 has been constructed containing the system of bioluminescence genes devoid of promoter following the polylinker DNA fragment. The plasmid can be used for selection of promoter containing DNA sequences as well as for studying the promoters regulation in process of Escherichia coli cells growth.

  20. Recurrent Hemolytic and Uremic Syndrome Induced by Escherichia Coli

    PubMed Central

    Commereuc, Morgane; Weill, Francois-Xavier; Loukiadis, Estelle; Gouali, Malika; Gleizal, Audrey; Kormann, Raphaël; Ridel, Christophe; Frémeaux-Bacchi, Véronique; Rondeau, Eric; Hertig, Alexandre

    2016-01-01

    Abstract A widespread belief is that typical hemolytic and uremic syndrome (HUS) does not recur. We report the case of a patient infected twice with raw milk taken from his own cow and containing a Shiga toxin–producing Escherichia coli O174:H21 that induced recurrent HUS causing severe renal and cerebral disorders. A genomic comparison of the human and bovine Shiga toxin–producing Escherichia coli O174:H21 isolates revealed that they were identical. Typical HUS may recur. Since milk from this animal was occasionally distributed locally, thereby posing a serious threat for the whole village, this particular cow was destroyed. PMID:26735524

  1. Diarrheagenic Escherichia coli in Children from Costa Rica

    PubMed Central

    Pérez, Cristian; Gómez-Duarte, Oscar G.; Arias, María L.

    2010-01-01

    More than 5,000 diarrheal cases per year receive medical care at the National Children's Hospital of Costa Rica, and nearly 5% of them require hospitalization. A total of 173 Escherichia coli strains isolated from children with diarrhea were characterized at the molecular, serologic, and phenotypic level. Multiplex and duplex polymerase chain reactions were used to detect the six categories of diarrheagenic E. coli. Thirty percent (n = 52) of the strains were positive, indicating a high prevalence among the pediatric population. Enteropathogenic E. coli and enteroinvasive E. coli pathotypes were the most prevalent (21% and 19%, respectively). Pathogenic strains were distributed among the four E. coli phylogenetic groups A, B1, B2, and D, with groups A and B1 the most commonly found. This study used molecular typing to evaluate the prevalence of diarrheagenic E. coli reported in Costa Rica and demonstrated the importance of these pathotypes in the pediatric population. PMID:20682870

  2. Selection of antibiotics in detection procedure of Escherichia coli O157:H7 in vegetables

    NASA Astrophysics Data System (ADS)

    Hoang, Hoang A.; Nhung, Nguyen T. T.

    2017-09-01

    Detection of Escherichia coli O157:H7 in ready-to-eat fresh vegetables is important since this bacteria is considered as one of the most important pathogens in relation to public health. However, it could be a big challenge for detection of initial low concentrations of E. coli O157:H7 in the samples. In this study, selection of antibiotics that suppress growth of background bacteria to enable detection of E. coli O157:H7 in ready-to-eat fresh vegetables was investigated. Firstly, different combinations of two antibiotics, i.e. novobiocin (N) and vancomycin (V), in BHI broth were conducted. The three antibiotic combinations were preliminary examined their effect on the growth of E. coli O157:H7 and Bacillus spp. in broth based on OD600nm measurement. The combination of both the antibiotics was selected to examine their possibility to support detection of E. coli O157:H7 in vegetables. It was successful when two antibiotics showed their support in detection of E. coli O157:H7 at very low concentration of 2 CFU per one gram of lettuce. Usage of these antibiotics is simple and cheap in the detection procedure and could be applied to other types of ready-to-eat fresh vegetables popular in Vietnam.

  3. Measuring the Viscosity of the Escherichia coli Plasma Membrane Using Molecular Rotors.

    PubMed

    Mika, Jacek T; Thompson, Alexander J; Dent, Michael R; Brooks, Nicholas J; Michiels, Jan; Hofkens, Johan; Kuimova, Marina K

    2016-10-04

    The viscosity is a highly important parameter within the cell membrane, affecting the diffusion of small molecules and, hence, controlling the rates of intracellular reactions. There is significant interest in the direct, quantitative assessment of membrane viscosity. Here we report the use of fluorescence lifetime imaging microscopy of the molecular rotor BODIPY C10 in the membranes of live Escherichia coli bacteria to permit direct quantification of the viscosity. Using this approach, we investigated the viscosity in live E. coli cells, spheroplasts, and liposomes made from E. coli membrane extracts. For live cells and spheroplasts, the viscosity was measured at both room temperature (23°C) and the E. coli growth temperature (37°C), while the membrane extract liposomes were studied over a range of measurement temperatures (5-40°C). At 37°C, we recorded a membrane viscosity in live E. coli cells of 950 cP, which is considerably higher than that previously observed in other live cell membranes (e.g., eukaryotic cells, membranes of Bacillus vegetative cells). Interestingly, this indicates that E. coli cells exhibit a high degree of lipid ordering within their liquid-phase plasma membranes. Copyright © 2016 Biophysical Society. Published by Elsevier Inc. All rights reserved.

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

    PubMed

    Riley, M

    1998-01-01

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

  5. Large Surface Blebs on Escherichia coli Heated to Inactivating Temperatures

    PubMed Central

    Scheie, Paul; Ehrenspeck, Susan

    1973-01-01

    Large surface blebs were observed with phase-contrast optics on Escherichia coli B/r and Bs-1 heated to temperatures at which colony-forming ability was lost. Characterization of such blebs was consistent with the view that they were formed by a physical process and were bounded by the outer membrane of the cell. A hypothesis for thermal inactivation of E. coli is presented that places membrane damage near the primary lethal event. Images PMID:4196258

  6. Expression of staphylococcal enterotoxin C1 in Escherichia coli.

    PubMed Central

    Bohach, G A; Schlievert, P M

    1987-01-01

    The structural gene encoding staphylococcal enterotoxin C1 was cloned into Escherichia coli and localized on a 1.5-kilobase HindIII-ClaI DNA fragment by subcloning. The toxin was partially purified from E. coli clones and shown to be immunologically identical to enterotoxin C1 from Staphylococcus aureus. The cloned toxin also had the same molecular weight (26,000) and charge heterogeneity as staphylococcus-derived enterotoxin. Toxins from both sources were equally biologically active. Images PMID:3542834

  7. Acetolactate Synthase from Bacillus subtilis Serves as a 2-Ketoisovalerate Decarboxylase for Isobutanol Biosynthesis in Escherichia coli▿

    PubMed Central

    Atsumi, Shota; Li, Zhen; Liao, James C.

    2009-01-01

    A pathway toward isobutanol production previously constructed in Escherichia coli involves 2-ketoacid decarboxylase (Kdc) from Lactococcus lactis that decarboxylates 2-ketoisovalerate (KIV) to isobutyraldehyde. Here, we showed that a strain lacking Kdc is still capable of producing isobutanol. We found that acetolactate synthase from Bacillus subtilis (AlsS), which originally catalyzes the condensation of two molecules of pyruvate to form 2-acetolactate, is able to catalyze the decarboxylation of KIV like Kdc both in vivo and in vitro. Mutational studies revealed that the replacement of Q487 with amino acids with small side chains (Ala, Ser, and Gly) diminished only the decarboxylase activity but maintained the synthase activity. PMID:19684168

  8. Lysis of Escherichia coli by Ethylenediaminetetraacetate and Phospholipases as Measured by β-Galactosidase Activity

    PubMed Central

    Slein, Milton W.; Logan, Gerald F.

    1967-01-01

    A permeaseless mutant of Escherichia coli, which produces β-galactosidase constitutively, was treated briefly with ethylenediaminetetraacetate and then with the phospholipases of Bacillus cereus. Cell lysis occurred, as indicated by an increase in β-galactosidase activity and a decrease in absorbancy of the cell suspension. The susceptibility of the cells to attack by ethylenediaminetetraacetate and the phospholipases was markedly affected by the age of the cells when harvested. The results suggest that permeability changes may be associated with the activity of a phospholipase that specifically degrades phosphatidyl ethanolamine. A sonic-treatment method for determining the total β-galactosidase content of E. coli cells, which is independent of their age when harvested, is described. PMID:4168313

  9. Multidrug-resistant Escherichia coli in Asia: epidemiology and management.

    PubMed

    Sidjabat, Hanna E; Paterson, David L

    2015-05-01

    Escherichia coli has become multiresistant by way of production of a variety of β-lactamases. The prevalence of CTX-M-producing E. coli has reached 60-79% in certain parts of Asia. The acquisition of CTX-M plasmids by E. coli sequence type 131, a successful clone of E. coli, has caused further dissemination of CTX-M-producing E. coli. The prevalence of carbapenemase-producing E. coli, especially Klebsiella pneumoniae carbapenemase, and New Delhi metallo-β-lactamase (NDM)-producing E. coli has been increasing in Asia. K. pneumoniae carbapenemase and NDM have now been found in E. coli sequence type 131. The occurrence of NDM-producing E. coli is a major concern particularly in the Indian subcontinent, but now elsewhere in Asia as well. There are multiple reasons why antibiotic resistance in E. coli in Asia has reached such extreme levels. Approaches beyond antibiotic therapy, such as prevention of antibiotic resistance by antibiotic stewardship and protecting natural microbiome, are strategies to avoid further spread of antibiotic resistance.

  10. Heat-stable Escherichia coli enterotoxin production in vivo.

    PubMed Central

    Whipp, S C; Moon, H W; Lyon, N C

    1975-01-01

    Hysterectomy-derived, colostrum-deprived piglets were infected with enterotoxigenic Escherichia coli on day 4 of life. Samples of feces and intestinal contents were collected and tested in infant mice for enterotoxic activity. Positive enterotoxic responses were observed in mice given filtrates of feces and intestinal contents from piglets infected withe enterotoxigenic E. coli known to produce heat-stable enterotoxin but not heat-liabile enterotoxin in vitro. It is concluded that heat-stable enterotoxigenic E. coli induce diarrhea by production of heat-stable enterotoxin in vivo. PMID:1097335

  11. An integrated database to support research on Escherichia coli

    SciTech Connect

    Baehr, A.; Dunham, G.; Matsuda, Hideo; Michaels, G.; Taylor, R.; Overbeek, R.; Rudd, K.E.; Ginsburg, A.; Joerg, D.; Kazic, T.; Hagstrom, R.; Zawada, D.; Smith, C.; Yoshida, Kaoru

    1992-01-01

    We have used logic programming to design and implement a prototype database of genomic information for the model bacterial organism Escherichia coli. This report presents the fundamental database primitives that can be used to access and manipulate data relating to the E. coli genome. The present system, combined with a tutorial manual, provides immediate access to the integrated knowledge base for E. coli chromosome data. It also serves as the foundation for development of more user-friendly interfaces that have the same retrieval power and high-level tools to analyze complex chromosome organization.

  12. The quantitative and condition-dependent Escherichia coli proteome

    PubMed Central

    Schmidt, Alexander; Kochanowski, Karl; Vedelaar, Silke; Ahrné, Erik; Volkmer, Benjamin; Callipo, Luciano; Knoops, Kèvin; Bauer, Manuel; Aebersold, Ruedi; Heinemann, Matthias

    2016-01-01

    Measuring precise concentrations of proteins can provide insights into biological processes. Here, we use efficient protein extraction and sample fractionation and state-of-the-art quantitative mass spectrometry techniques to generate a comprehensive, condition-dependent protein abundance map of Escherichia coli. We measure cellular protein concentrations for 55% of predicted E. coli genes (>2300 proteins) under 22 different experimental conditions and identify methylation and N-terminal protein acetylations previously not known to be prevalent in bacteria. We uncover system-wide proteome allocation, expression regulation, and post-translational adaptations. These data provide a valuable resource for the systems biology and broader E. coli research communities. PMID:26641532

  13. YeeO from Escherichia coli exports flavins.

    PubMed

    McAnulty, Michael J; Wood, Thomas K

    2014-01-01

    Multidrug and toxic compound extrusion (MATE) proteins help maintain cellular homeostasis by secreting metabolic wastes. Flavins may occur as cellular waste products, with their production and secretion providing potential benefit for industrial applications related to biofuel cells. Here we find that MATE protein YeeO from Escherichia coli exports both flavin mononucleotide (FMN) and flavin adenine dinucleotide (FAD). Significant amounts of flavins were trapped intracellularly when YeeO was produced indicating transport limits secretion of flavins. Wild-type E. coli secreted 3 flavins (riboflavin, FMN, and FAD), so E. coli likely produces additional flavin transporters.

  14. YeeO from Escherichia coli exports flavins

    PubMed Central

    McAnulty, Michael J; Wood, Thomas K

    2014-01-01

    Multidrug and toxic compound extrusion (MATE) proteins help maintain cellular homeostasis by secreting metabolic wastes. Flavins may occur as cellular waste products, with their production and secretion providing potential benefit for industrial applications related to biofuel cells. Here we find that MATE protein YeeO from Escherichia coli exports both flavin mononucleotide (FMN) and flavin adenine dinucleotide (FAD). Significant amounts of flavins were trapped intracellularly when YeeO was produced indicating transport limits secretion of flavins. Wild-type E. coli secreted 3 flavins (riboflavin, FMN, and FAD), so E. coli likely produces additional flavin transporters. PMID:25482085

  15. Reassessing Escherichia coli as a cell factory for biofuel production.

    PubMed

    Wang, Chonglong; Pfleger, Brian F; Kim, Seon-Won

    2017-03-11

    Via metabolic engineering, industrial microorganisms have the potential to convert renewable substrates into a wide range of biofuels that can address energy security and environmental challenges associated with current fossil fuels. The user-friendly bacterium, Escherichia coli, remains one of the most frequently used hosts for demonstrating production of biofuel candidates including alcohol-, fatty acid- and terpenoid-based biofuels. In this review, we summarize the metabolic pathways for synthesis of these biofuels and assess enabling technologies that assist in regulating biofuel synthesis pathways and rapidly assembling novel E. coli strains. These advances maintain E. coli's position as a prominent host for developing cell factories for biofuel production.

  16. Enteropathogenic Escherichia coli Serotypes and Endemic Diarrhea in Infants

    PubMed Central

    Toledo, M. Regina F.; Alvariza, M. do Carmo B.; Murahovschi, Jayme; Ramos, Sonia R. T. S.; Trabulsi, Luiz R.

    1983-01-01

    Enteropathogenic Escherichia coli serotypes were searched for in feces of 550 children with endemic diarrhea and in 129 controls, in São Paulo, in 1978 and 1979; serotypes O111ab:H−, O111ab:H2, and O119:H6 were significantly associated with diarrhea in children 0 to 5 months old and were the most frequent agents of diarrhea in this age group as compared with enterotoxigenic and enteroinvasive E. coli, Salmonella sp., Shigella sp., and Yersinia enterocolitica. It is concluded that various enteropathogenic E. coli serotypes may be agents of endemic infantile diarrhea. PMID:6339384

  17. Sources of Escherichia coli in a Coastal Subtropical Environment

    PubMed Central

    Solo-Gabriele, Helena M.; Wolfert, Melinda A.; Desmarais, Timothy R.; Palmer, Carol J.

    2000-01-01

    Sources of Escherichia coli in a coastal waterway located in Ft. Lauderdale, Fla., were evaluated. The study consisted of an extensive program of field measurements designed to capture spatial and temporal variations in E. coli concentrations as well as experiments conducted under laboratory-controlled conditions. E. coli from environmental samples was enumerated by using a defined substrate technology (Colilert-18). Field sampling tasks included sampling the length of the North Fork to identify the river reach contributing high E. coli levels, autosampler experiments at two locations, and spatially intense sampling efforts at hot spots. Laboratory experiments were designed to simulate tidal conditions within the riverbank soils. The results showed that E. coli entered the river in a large pulse during storm conditions. After the storm, E. coli levels returned to baseline levels and varied in a cyclical pattern which correlated with tidal cycles. The highest concentrations were observed during high tide, whereas the lowest were observed at low tide. This peculiar pattern of E. coli concentrations between storm events was caused by the growth of E. coli within riverbank soils which were subsequently washed in during high tide. Laboratory analysis of soil collected from the riverbanks showed increases of several orders of magnitude in soil E. coli concentrations. The ability of E. coli to multiply in the soil was found to be a function of soil moisture content, presumably due to the ability of E. coli to outcompete predators in relatively dry soil. The importance of soil moisture in regulating the multiplication of E. coli was found to be critical in tidally influenced areas due to periodic wetting and drying of soils in contact with water bodies. Given the potential for growth in such systems, E. coli concentrations can be artificially elevated above that expected from fecal impacts alone. Such results challenge the use of E. coli as a suitable indicator of water

  18. Structure of Water in Escherichia Coli B

    DTIC Science & Technology

    structure broadening of the NMR water spectrum. Using bacteria grown in the special chemically defined medium, we showed that the water in E. coli B was highly ordered and was very different from ’free’ water and from polywater .

  19. Slugs: Potential Novel Vectors of Escherichia coli O157

    PubMed Central

    Sproston, Emma L.; Macrae, M.; Ogden, Iain D.; Wilson, Michael J.; Strachan, Norval J. C.

    2006-01-01

    Field and laboratory studies were performed to determine whether slugs could act as novel vectors for pathogen (e.g., Escherichia coli O157) transfer from animal feces to salad vegetables. Escherichia coli O157 was isolated from 0.21% of field slugs from an Aberdeenshire sheep farm. These isolates carried the verocytotoxin genes (vt1 and vt2) and the attaching and effacing gene (eae), suggesting that they are potentially pathogenic to humans. Strain typing using multilocus variable number tandem repeats analysis showed that slug and sheep isolates were indistinguishable. Laboratory experiments using an E. coli mutant resistant to nalidixic acid showed that the ubiquitous slug species Deroceras reticulatum could carry viable E. coli on its external surface for up to 14 days. Slugs that had been fed E. coli shed viable bacteria in their feces with numbers showing a short but statistically significant linear log decline. Further, it was found that E. coli persisted for up to 3 weeks in excreted slug feces, and hence, we conclude that slugs have the potential to act as novel vectors of E. coli O157. PMID:16391036

  20. 76 FR 58157 - Shiga Toxin-Producing Escherichia coli

    Federal Register 2010, 2011, 2012, 2013, 2014

    2011-09-20

    ... infections.\\1\\ \\1\\ U.S. Centers for Disease Control and Prevention. 2005. Shiga toxin-producing Escherichia coli (STEC). National Notifiable Diseases Surveillance System (NNDSS), 2005 Case Definition. http://www...) 1422-1429. \\6\\ Centers for Disease Control and Prevention. Bacterial Foodborne and Diarrheal Disease...

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

    USDA-ARS?s Scientific Manuscript database

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

  2. Genome Sequence of Enterotoxigenic Escherichia coli Strain FMU073332

    PubMed Central

    Saldaña-Ahuactzi, Zeus; Cruz-Córdova, Ariadnna; Rodea, Gerardo E.; Porta, Helena; Navarro-Ocaña, Armando; Eslava-Campos, Carlos

    2017-01-01

    ABSTRACT   Enterotoxigenic Escherichia coli (ETEC) is an important cause of bacterial diarrheal illness, affecting practically every population worldwide, and was estimated to cause 120,800 deaths in 2010. Here, we report the genome sequence of ETEC strain FMU073332, isolated from a 25-month-old girl from Tlaltizapán, Morelos, México. PMID:28232434

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

    EPA Science Inventory

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

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

    USDA-ARS?s Scientific Manuscript database

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

  5. Stringent control of FLP recombinase in Escherichia coli.

    PubMed

    Bowden, Steven D; Palani, Nagendra P; Libourel, Igor G L

    2017-02-01

    Site specific recombinases are invaluable tools in molecular biology, and are emerging as powerful recorders of cellular events in synthetic biology. We have developed a stringently controlled FLP recombinase system in Escherichia coli using an arabinose inducible promoter combined with a weak ribosome binding site.

  6. Enteroinvasive Escherichia coli severe dysentery complicated by rotavirus gastroenteritis.

    PubMed

    Pacheco-Gil, Leova; Ochoa, Theresa J; Flores-Romo, Leopoldo; DuPont, Herbert L; Estrada-Garcia, Teresa

    2006-11-01

    Enteroinvasive Escherichia coli (EIEC) is an important agent of pediatric diarrhea and dysentery in developing countries. We report a life-threatening severe dysentery case due to EIEC in a malnourished 4-month-old male, native Indian infant co-infected with rotavirus. The severe gastrointestinal bleeding anemia and hypovolemic shock was successfully treated with IV blood transfusions, rehydration and antibiotic therapy.

  7. Division Planes Alternate in Spherical Cells of Escherichia coli

    PubMed Central

    Begg, K. J.; Donachie, W. D.

    1998-01-01

    In the spherical cells of Escherichia coli rodA mutants, division is initiated at a single point, from which a furrow extends progressively around the cell. Using “giant” rodA ftsA cells, we confirmed that each new division furrow is initiated at the midpoint of the previous division plane and runs perpendicular to it. PMID:9573213

  8. More than a locomotive organelle: flagella in Escherichia coli.

    PubMed

    Zhou, Mingxu; Yang, Yang; Chen, Panlin; Hu, Huijie; Hardwidge, Philip R; Zhu, Guoqiang

    2015-11-01

    The flagellum is a locomotive organelle that allows bacteria to respond to chemical gradients. This review summarizes the current knowledge regarding Escherichia coli flagellin variants and the role of flagella in bacterial functions other than motility, including the relationship between flagella and bacterial virulence.

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

    EPA Science Inventory

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

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

    EPA Science Inventory

    The increase in resistance rates to trimehtoprim-sulfamethoxazole (TMP/SMX) in isolates of Escherichia coli has become a matter of increasing concern. This has been particularly true in reference to community acquired urinary tract infections (UTI). This study utilized sewage i...

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

    USDA-ARS?s Scientific Manuscript database

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

  12. New types of Escherichia coli recombination-deficient mutants.

    PubMed

    Freifelder, D

    1976-11-01

    A set of Escherichia coli mutants deficient in intramolecular recombination and different from those previously found is described. All have temperature-sensitive lethal mutations. The mutants have been characterized with respect to the following properties: the Pap phenotype, deoxyribonucleic acid synthesis, sensitivity to ultraviolet light, ability to support the growth of phage lambda, filament formation, and mutation frequency.

  13. New types of Escherichia coli recombination-deficient mutants.

    PubMed Central

    Freifelder, D

    1976-01-01

    A set of Escherichia coli mutants deficient in intramolecular recombination and different from those previously found is described. All have temperature-sensitive lethal mutations. The mutants have been characterized with respect to the following properties: the Pap phenotype, deoxyribonucleic acid synthesis, sensitivity to ultraviolet light, ability to support the growth of phage lambda, filament formation, and mutation frequency. PMID:789362

  14. Escherichia coli growth studied by dual-parameter flow cytophotometry.

    PubMed Central

    Steen, H B; Boye, E

    1981-01-01

    The growth of Escherichia coli cells has been analyzed for the first time by dual-parameter flow cytophotometry, in which the deoxyribonucleic acid and protein contents of single bacteria have been measured simultaneously with an accuracy of a few percent and at a rate of 3,000 cells/s. PMID:7007339

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

    EPA Science Inventory

    The increase in resistance rates to trimehtoprim-sulfamethoxazole (TMP/SMX) in isolates of Escherichia coli has become a matter of increasing concern. This has been particularly true in reference to community acquired urinary tract infections (UTI). This study utilized sewage i...

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

    USDA-ARS?s Scientific Manuscript database

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

  17. Isolation of an Lc-specific Escherichia coli bacteriophage.

    PubMed Central

    Fralick, J A; Diedrich, D L; Casey-Wood, S

    1990-01-01

    We isolated an OmpF-specific bacteriophage whose host range mutant, SQ108h2, requires the presence of the Lc porin for its attachment and which can be used to screen or select for Lc-defective mutants among Escherichia coli K-12 strains lysogenic for the PA-2 converting phage. Images FIG. 1 PMID:1689719

  18. Plasmolysis of Escherichia coli B-r with sucrose.

    PubMed

    Scheie, P O

    1969-05-01

    Escherichia coli B/r cells were plasmolyzed in sucrose solutions and observed under phase contrast. The prevalence of plasmolysis under various conditions was noted, and the degree of plasmolysis was categorized as slight, extensive, or severe. The presence of ions reduced the prevalence of plasmolysis. Survival curves showed that extensive plasmolysis was not lethal to colony-forming ability.

  19. Naturally Occurring Extended-Spectrum Cephalosporinases in Escherichia coli

    PubMed Central

    Mammeri, Hedi; Poirel, Laurent; Fortineau, Nicolas; Nordmann, Patrice

    2006-01-01

    Genetic and functional characterization of the cephalosporinases produced by 65 clonally unrelated clinical Escherichia coli isolates revealed genetic diversity of the ampC genes and showed that Gln287, Cys287, Pro296, Leu298, and Phe350 substitutions were involved in extension of the hydrolysis spectrum to include ceftazidime and cefepime. PMID:16801449

  20. armA and aminoglycoside resistance in Escherichia coli.

    PubMed

    González-Zorn, Bruno; Teshager, Tirushet; Casas, María; Porrero, María C; Moreno, Miguel A; Courvalin, Patrice; Domínguez, Lucas

    2005-06-01

    We report armA in an Escherichia coli pig isolate from Spain. The resistance gene was borne by self-transferable IncN plasmid pMUR050. Molecular analysis of the plasmid and of the armA locus confirmed the spread of this resistance determinant.

  1. armA and Aminoglycoside Resistance in Escherichia coli

    PubMed Central

    González-Zorn, Bruno; Teshager, Tirushet; Casas, María; Porrero, María C.; Courvalin, Patrice; Domínguez, Lucas

    2005-01-01

    We report armA in an Escherichia coli pig isolate from Spain. The resistance gene was borne by self-transferable IncN plasmid pMUR050. Molecular analysis of the plasmid and of the armA locus confirmed the spread of this resistance determinant. PMID:15963296

  2. Norfloxacin resistance in a clinical isolate of Escherichia coli.

    PubMed Central

    Aoyama, H; Sato, K; Kato, T; Hirai, K; Mitsuhashi, S

    1987-01-01

    Analysis of DNA gyrase supercoiling and of norfloxacin uptake in Escherichia coli GN14176, a moderately norfloxacin-resistant clinical isolate, indicated that resistance was associated with both an altered drug target and a reduction in drug uptake. Images PMID:2829712

  3. Enterotoxigenic Escherichia coli and Vibrio cholerae diarrhea, Bangladesh, 2004.

    PubMed

    Qadri, Firdausi; Khan, Ashraful I; Faruque, Abu Syed G; Begum, Yasmin Ara; Chowdhury, Fahima; Nair, Gopinath B; Salam, Mohammed A; Sack, David A; Svennerholm, Ann-Mari

    2005-07-01

    Flooding in Dhaka in July 2004 caused epidemics of diarrhea. Enterotoxigenic Escherichia coli (ETEC) was almost as prevalent as Vibrio cholerae O1 in diarrheal stools. ETEC that produced heat-stable enterotoxin alone was most prevalent, and 78% of strains had colonization factors. Like V. cholerae O1, ETEC can cause epidemic diarrhea.

  4. Plasmolysis of Escherichia coli B/r with Sucrose

    PubMed Central

    Scheie, Paul O.

    1969-01-01

    Escherichia coli B/r cells were plasmolyzed in sucrose solutions and observed under phase contrast. The prevalence of plasmolysis under various conditions was noted, and the degree of plasmolysis was categorized as slight, extensive, or severe. The presence of ions reduced the prevalence of plasmolysis. Survival curves showed that extensive plasmolysis was not lethal to colony-forming ability. PMID:4891252

  5. Multidrug-Resistant Escherichia coli in Bovine Animals, Europe

    PubMed Central

    Brennan, Evan; Martins, Marta; McCusker, Matthew P.; Wang, Juan; Alves, Bruno Martins; Hurley, Daniel; El Garch, Farid; Woehrlé, Frédérique; Miossec, Christine; McGrath, Leisha; Srikumar, Shabarinath; Wall, Patrick

    2016-01-01

    Of 150 Escherichia coli strains we cultured from specimens taken from cattle in Europe, 3 had elevated MICs against colistin. We assessed all 3 strains for the presence of the plasmid-mediated mcr-1 gene and identified 1 isolate as mcr-1–positive and co-resistant to β-lactam, florfenicol, and fluoroquinolone antimicrobial compounds. PMID:27533105

  6. rRNA transcription rate in Escherichia coli.

    PubMed Central

    Gotta, S L; Miller, O L; French, S L

    1991-01-01

    The rate of in vivo transcription elongation for Escherichia coli rRNA operons was determined by electron microscopy following addition of rifampin to log-phase cultures. Direct observation of RNA polymerase positions along rRNA operons 30, 40, and 70 s after inhibition of transcription initiation yielded a transcription elongation rate of 42 nucleotides per s. Images FIG. 1 PMID:1717439

  7. Immunologic Control of Diarrheal Disease Due to Enterotoxigenic Escherichia coli

    DTIC Science & Technology

    1984-01-01

    Classical Enteropathogenic (Serotyped) Escherichia coli Strains of Proven Pathogenicity. Infect. Immun. 38:798-801, 1982. 8. Levine, M.M. Vacunas Contra...Microbiol., 18:808-815, 1983. 8 15. Levine, M.M., Lanata, C. Progresos en Vacunas Contra Diarrea Bacteriana. Adelantos Microbiol. Enferm. Inf., 2:67-117

  8. EcoCyc: Encyclopedia of Escherichia coli genes and metabolism.

    PubMed

    Karp, P D; Riley, M; Paley, S M; Pellegrini-Toole, A; Krummenacker, M

    1998-01-01

    The encyclopedia of Escherichia coli genes and metabolism (EcoCyc) is a database that combines information about the genome and the intermediary metabolism of E.coli. The database describes 3030 genes of E.coli , 695 enzymes encoded by a subset of these genes, 595 metabolic reactions that occur in E.coli, and the organization of these reactions into 123 metabolic pathways. The EcoCyc graphical user interface allows scientists to query and explore the EcoCyc database using visualization tools such as genomic-map browsers and automatic layouts of metabolic pathways. EcoCyc can be thought of as an electronic review article because of its copious references to the primary literature, and as a (qualitative) computational model of E.coli metabolism. EcoCyc is available at URL http://ecocyc.PangeaSystems.com/ecocyc/

  9. Phylogenetic Group Determination of Escherichia coli Isolated from Animals Samples

    PubMed Central

    Morcatti Coura, Fernanda; Diniz, Soraia de Araújo; Silva, Marcos Xavier; Mussi, Jamili Maria Suhet; Barbosa, Silvia Minharro; Lage, Andrey Pereira; Heinemann, Marcos Bryan

    2015-01-01

    This study analyzes the occurrence and distribution of phylogenetic groups of 391 strains of Escherichia coli isolated from poultry, cattle, and water buffalo. The frequency of the phylogroups was A = 19%, B1 = 57%, B2 = 2.3%, C = 4.6%, D = 2.8%, E = 11%, and F = 3.3%. Phylogroups A (P < 0.001) and F (P = 0.018) were associated with E. coli strains isolated from poultry, phylogroups B1 (P < 0.001) and E (P = 0.002) were associated with E. coli isolated from cattle, and phylogroups B2 (P = 0.003) and D (P = 0.017) were associated with E. coli isolated from water buffalo. This report demonstrated that some phylogroups are associated with the host analyzed and the results provide knowledge of the phylogenetic composition of E. coli from domestic animals. PMID:26421310

  10. Glycerol elicits energy taxis of Escherichia coli and Salmonella typhimurium.

    PubMed

    Zhulin, I B; Rowsell, E H; Johnson, M S; Taylor, B L

    1997-05-01

    Escherichia coli and Salmonella typhimurium show positive chemotaxis to glycerol, a chemical previously reported to be a repellent for E. coli. The threshold of the attractant response in both species was 10(-6) M glycerol. Glycerol chemotaxis was energy dependent and coincident with an increase in membrane potential. Metabolism of glycerol was required for chemotaxis, and when lactate was present to maintain energy production in the absence of glycerol, the increases in membrane potential and chemotactic response upon addition of glycerol were abolished. Methylation of a chemotaxis receptor was not required for positive glycerol chemotaxis in E. coli or S. typhimurium but is involved in the negative chemotaxis of E. coli to high concentrations of glycerol. We propose that positive chemotaxis to glycerol in E. coli and S. typhimurium is an example of energy taxis mediated via a signal transduction pathway that responds to changes in the cellular energy level.

  11. Using zebra mussels to monitor Escherichia coli in environmental waters.

    PubMed

    Selegean, J P; Kusserow, R; Patel, R; Heidtke, T M; Ram, J L

    2001-01-01

    Use of the zebra mussel (Dreissena polymorpha) as an indicator of previously elevated bacteria concentrations in a watershed was examined. The ability of the zebra mussel to accumulate and purge Escherichia coli over several days was investigated in both laboratory and field experiments. In laboratory experiments, periodic enumeration of E. coli in mussels that had been exposed to a dilute solution of raw sewage demonstrated that (i) maximum concentrations of E. coli are reached within a few hours of exposure to sewage, (ii) the tissue concentration attained is higher than the concentration in the ambient water, and (iii) the E. coli concentrations take several days to return to preexposure concentrations when mussels are subsequently placed in sterile water. In field experiments conducted in southeast Michigan in the Clinton River watershed, brief increases in E. coli concentrations in the water were accompanied by increases in mussel concentrations of E. coli that lasted 2 or 3 d. The ability of mussels to retain and to concentrate E. coli made it possible to detect E. coli in the environment under conditions that conventional monitoring may often miss. Sampling caged mussels in a river and its tributaries may enable watershed managers to reduce the sampling frequency normally required to identify critical E. coli sources, thereby providing a more cost-effective river monitoring strategy for bacterial contamination.

  12. Cytotoxic Escherichia coli strains encoding colibactin colonize laboratory mice.

    PubMed

    García, Alexis; Mannion, Anthony; Feng, Yan; Madden, Carolyn M; Bakthavatchalu, Vasudevan; Shen, Zeli; Ge, Zhongming; Fox, James G

    2016-12-01

    Escherichia coli strains have not been fully characterized in laboratory mice and are not currently excluded from mouse colonies. Colibactin (Clb), a cytotoxin, has been associated with inflammation and cancer in humans and animals. We performed bacterial cultures utilizing rectal swab, fecal, and extra intestinal samples from clinically unaffected or affected laboratory mice. Fifty-one E. coli were isolated from 45 laboratory mice, identified biochemically, and selected isolates were serotyped. The 16S rRNA gene was amplified and sequenced for specific isolates, PCR used for clbA and clbQ gene amplification, and phylogenetic group identification was performed on all 51 E. coli strains. Clb genes were sequenced and selected E. coli isolates were characterized using a HeLa cell cytotoxicity assay. Forty-five of the 51 E. coli isolates (88%) encoded clbA and clbQ and belonged to phylogenetic group B2. Mouse E. coli serotypes included: O2:H6, O-:H-, OM:H+, and O22:H-. Clb-encoding O2: H6 mouse E. coli isolates were cytotoxic in vitro. A Clb-encoding E. coli was isolated from a clinically affected genetically modified mouse with cystic endometrial hyperplasia. Our findings suggest that Clb-encoding E. coli colonize laboratory mice and may induce clinical and subclinical diseases that may impact experimental mouse models.

  13. Evolution of the iss gene in Escherichia coli.

    PubMed

    Johnson, Timothy J; Wannemuehler, Yvonne M; Nolan, Lisa K

    2008-04-01

    The increased serum survival gene iss has long been recognized for its role in extraintestinal pathogenic Escherichia coli (ExPEC) virulence. iss has been identified as a distinguishing trait of avian ExPEC but not of human ExPEC. This gene has been localized to large virulence plasmids and shares strong similarities with the bor gene from bacteriophage lambda. Here, we demonstrate that three alleles of iss occur among E. coli isolates that appear to have evolved from a common lambda bor precursor. In addition to the occurrence of iss on the ColV/BM virulence plasmids, at least two iss alleles occur within the E. coli chromosome. One of these alleles (designated type 3) was found to occur in the genomes of all currently sequenced ExPEC strains on a similar prophage element that also harbors the Sit iron and manganese transport system. When the prevalence of the three iss types was examined among 487 E. coli isolates, the iss type 3 gene was found to occur at a high frequency among ExPEC isolates, irrespective of the host source. The plasmid-borne iss allele (designated type 1) was highly prevalent among avian pathogenic E. coli and neonatal meningitis-associated E. coli isolates but not among uropathogenic E. coli isolates. This study demonstrates the evolution of iss in E. coli and provides an additional tool for discriminating among E. coli pathotypes through the differentiation of the three iss allele types and bor.

  14. EFFECT OF DIHYDROSTREPTOMYCIN ON TETRAZOLIUM DYE REDUCTION IN ESCHERICHIA COLI

    PubMed Central

    Bragg, P. D.; Polglase, W. J.

    1963-01-01

    Bragg, P. D. (University of British Columbia, Vancouver, British Columbia, Canada) and W. J. Polglase. Effect of dihydrostreptomycin on tetrazolium dye reduction in Escherichia coli. J. Bacteriol. 85:795–800. 1963.—Sonic-disrupted extracts of Escherichia coli, grown without added antibiotic (sensitive and resistant), contained (in supernatant of fraction centrifuged at 100,000 × g) a dihydrostreptomycin-inhibitable, succinate-triphenyltetrazolium chloride (TTC) reductase activity. The succinate-TTC reductase activities of extracts of E. coli grown in the presence of dihydrostreptomycin (resistant and dependent) were relatively low and were not inhibited by the antibiotic. At a moderate magnesium concentration, the degree of inhibition by dihydrostreptomycin of succinate-TTC reductase activity was sufficiently marked to indicate an important site of action of the antibiotic. Magnesium, putrescine, and spermidine antagonized the action of dihydrostreptomycin in the succinate-TTC reductase system. PMID:14044945

  15. EFFECT OF DIHYDROSTREPTOMYCIN ON TETRAZOLIUM DYE REDUCTION IN ESCHERICHIA COLI.

    PubMed

    BRAGG, P D; POLGLASE, W J

    1963-04-01

    Bragg, P. D. (University of British Columbia, Vancouver, British Columbia, Canada) and W. J. Polglase. Effect of dihydrostreptomycin on tetrazolium dye reduction in Escherichia coli. J. Bacteriol. 85:795-800. 1963.-Sonic-disrupted extracts of Escherichia coli, grown without added antibiotic (sensitive and resistant), contained (in supernatant of fraction centrifuged at 100,000 x g) a dihydrostreptomycin-inhibitable, succinate-triphenyltetrazolium chloride (TTC) reductase activity. The succinate-TTC reductase activities of extracts of E. coli grown in the presence of dihydrostreptomycin (resistant and dependent) were relatively low and were not inhibited by the antibiotic. At a moderate magnesium concentration, the degree of inhibition by dihydrostreptomycin of succinate-TTC reductase activity was sufficiently marked to indicate an important site of action of the antibiotic. Magnesium, putrescine, and spermidine antagonized the action of dihydrostreptomycin in the succinate-TTC reductase system.

  16. Polyerositis and Arthritis Due to Escherichia coli in Gnotobiotic Pigs

    PubMed Central

    Waxler, G. L.; Britt, A. L.

    1972-01-01

    Forty gnotobiotic pigs from six litters were exposed orally to Escherichia coli 083:K·:NM at 69 to 148 hours of age, while 17 pigs from the same litters served as unexposed controls. Clinical signs of infection included fever, anorexia, diarrhea, lameness, and reluctance to move. Eighty-four percent of the exposed pigs in four litters died, while only 13% in two litters died. Gross and microscopic lesions included serofibrinous to fibrinopurulent polyserositis in 96% of the exposed pigs in four litters and 33% of the exposed pigs in two litters. A few pigs had gross and/or microscopic lesions of arthritis. Escherichia coli was routinely isolated from the serous and synovial cavities of infected pigs. Anti-hog cholera serum administered orally as a colostrum substitute gave partial protection against E. coli infection. ImagesFig. 1.Fig. 2.Fig. 3.Fig. 4.Fig. 5.Fig. 6.Fig. 7.Fig. 8. PMID:4261837

  17. Colonization and infection with fluoroquinolone-resistant Escherichia coli among cancer patients: clonal analysis.

    PubMed

    Oethinger, M; Jellen-Ritter, A S; Conrad, S; Marre, R; Kern, W V

    1998-01-01

    Escherichia coli with high-level fluoroquinolone resistance were isolated from feces and/or various body sites of 16 cancer patients who were on oral fluoroquinolone prophylaxis. Population analysis of fecal isolates in 11 patients showed that fluoroquinolone-resistant E. coli was the only aerobic gram-negative bacillus present and exhibited a relatively homogenous fluoroquinolone MIC distribution. Molecular typing by pulsed field gel electrophoresis of chromosomal DNA digests or by random amplified polymorphic DNA fingerprinting confirmed the clonal nature of gastrointestinal tract colonization with E. coli. Genotyping of ten colonies picked from the same fecal culture demonstrated identical strains in four of four patients examined. Identical genotypes from the same patient were isolated over prolonged periods of time in 12 of 12 cases examined, with one patient (with the longest follow-up of 14 months) who lost his initial genotype and became persistently colonized with a new genotype. In the 11 patients who developed infection due to fluoroquinolone-resistant E. coli, molecular typing also indicated that fecal colonization was associated with, and presumably preceded infection due to an indistinguishable genotype of fluoroquinolone-resistant E. coli.

  18. Production of itaconic acid in Escherichia coli expressing recombinant α-amylase using starch as substrate.

    PubMed

    Okamoto, Shusuke; Chin, Taejun; Nagata, Keisuke; Takahashi, Tetsuya; Ohara, Hitomi; Aso, Yuji

    2015-05-01

    Several studies on fermentative production of a vinyl monomer itaconic acid from hydrolyzed starch using Aspergillus terreus have been reported. Herein, we report itaconic acid production by Escherichia coli expressing recombinant α-amylase, using soluble starch as its sole carbon source. To express α-amylase in E. coli, we first constructed recombinant plasmids expressing α-amylases by using cell surface display technology derived from two amylolytic bacteria, Bacillus amyloliquefaciens NBRC 15535(T) and Streptococcus bovis NRIC 1535. The recombinant α-amylase from S. bovis (SBA) showed activity at 28°C, which is the optimal temperature for production of itaconic acid, while α-amylase from B. amyloliquefaciens displayed no noticeable activity. E. coli cells expressing SBA produced 0.15 g/L itaconic acid after 69 h cultivation under pH-stat conditions, using 1% starch as the sole carbon source. In fact, E. coli cells expressing SBA had similar growth rates when grown in the presence of 1% glucose or starch, thereby highlighting the expression of an active α-amylase that enabled utilization of starch to produce itaconic acid in E. coli.

  19. Expression of antimicrobial peptide LH multimers in Escherichia coli C43(DE3).

    PubMed

    Tian, Zi-gang; Dong, Tian-tang; Yang, Ya-lin; Teng, Da; Wang, Jian-hua

    2009-05-01

    The tandem repeats of LFB15(W4,10)-HP(4-16) (LH) gene were cloned into vector pET32a(+) for recombinant expression in Escherichia coli. The E. coli C43(DE3) was successfully used as the expression host to avoid the cell death during induction in E. coli BL21(DE3). Fusion LH dimer was expressed as inclusion body at a portion of 35% of total cell protein and could be well purified by Ni(2+)-chelating chromatography. The recombinant LH was released by the cleavage of 50% formic acid, and its yield reached 11.3 mg/l with purity of 95%. The MIC(50) of 3.6 and 1.9 microM of recombinant LH against E. coli CMCC 44102 and Bacillus subtilis ATCC 6633 were determined, respectively. The results demonstrated that expression of tandem LH gene in E. coli C43(DE3) and formic acid cleavage would provide a potent efficient platform for the production of interested peptides.

  20. Study of the effects of high-energy proton beams on escherichia coli

    NASA Astrophysics Data System (ADS)

    Park, Jeong Chan; Jung, Myung-Hwan

    2015-10-01

    Antibiotic-resistant bacterial infection is one of the most serious risks to public health care today. However, discouragingly, the development of new antibiotics has progressed little over the last decade. There is an urgent need for alternative approaches to treat antibiotic-resistant bacteria. Novel methods, which include photothermal therapy based on gold nano-materials and ionizing radiation such as X-rays and gamma rays, have been reported. Studies of the effects of high-energy proton radiation on bacteria have mainly focused on Bacillus species and its spores. The effect of proton beams on Escherichia coli (E. coli) has been limitedly reported. Escherichia coli is an important biological tool to obtain metabolic and genetic information and is a common model microorganism for studying toxicity and antimicrobial activity. In addition, E. coli is a common bacterium in the intestinal tract of mammals. In this research, the morphological and the physiological changes of E. coli after proton irradiation were investigated. Diluted solutions of cells were used for proton beam radiation. LB agar plates were used to count the number of colonies formed. The growth profile of the cells was monitored by using the optical density at 600 nm. The morphology of the irradiated cells was observed with an optical microscope. A microarray analysis was performed to examine the gene expression changes between irradiated samples and control samples without irradiation. E coli cells have observed to be elongated after proton irradiation with doses ranging from 13 to 93 Gy. Twenty-two were up-regulated more than twofold in proton-irradiated samples (93 Gy) compared with unexposed one.

  1. Lytic bacteriophages reduce Escherichia coli O157

    PubMed Central

    Ferguson, Sean; Roberts, Cheryl; Handy, Eric; Sharma, Manan

    2013-01-01

    The role of lytic bacteriophages in preventing cross contamination of produce has not been evaluated. A cocktail of three lytic phages specific for E. coli O157:H7 (EcoShield™) or a control (phosphate buffered saline, PBS) was applied to lettuce by either; (1) immersion of lettuce in 500 ml of EcoShield™ 8.3 log PFU/ml or 9.8 log PFU/ml for up to 2 min before inoculation with E. coli O157:H7; (2) spray-application of EcoShield™ (9.3 log PFU/ml) to lettuce after inoculation with E. coli O157:H7 (4.10 CFU/cm2) following exposure to 50 μg/ml chlorine for 30 sec. After immersion studies, lettuce was spot-inoculated with E. coli O157:H7 (2.38 CFU/cm2). Phage-treated, inoculated lettuce pieces were stored at 4°C for and analyzed for E. coli O157:H7 populations for up to 7 d. Immersion of lettuce in 9.8 log PFU/ml EcoShield™ for 2 min significantly (p < 0.05) reduced E. coli O157:H7 populations after 24 h when stored at 4°C compared with controls. Immersion of lettuce in suspensions containing high concentrations of EcoShield™ (9.8 log PFU/ml) resulted in the deposition of high concentrations (7.8 log log PFU/cm2) of bacteriophages on the surface of fresh cut lettuce, potentially contributing to the efficacy of the lytic phages on lettuce. Spraying phages on to inoculated fresh cut lettuce after being washed in hypochlorite solution was significantly more effective in reducing E. coli O157:H7 populations (2.22 log CFU/cm2) on day 0 compared with control treatments (4.10 log CFU/cm2). Both immersion and spray treatments provided protection from E. coli O157:H7 contamination on lettuce, but spray application of lytic bacteriophages to lettuce was more effective in immediately reducing E. coli O157:H7 populations fresh cut lettuce. PMID:23819106

  2. Adhesive threads of extraintestinal pathogenic Escherichia coli.

    PubMed

    Antão, Esther-Maria; Wieler, Lothar H; Ewers, Christa

    2009-12-10

    The ability to adhere to host surfaces is by far the most vital step in the successful colonization by microbial pathogens. Colonization begins with the attachment of the bacterium to receptors expressed by cells forming the lining of the mucosa. Long hair like extracellular appendages called fimbriae, produced by most Gram-negative pathogens, mediate specific attachment to the epithelial cell surface. Associated with the fimbriae is a protein called an adhesin, which directs high-affinity binding to specific cell surface components. In the last couple of years, an enormous amount of research has been undertaken that deals with understanding how bacterial pathogens adhere to host cells. E. coli in all probability is one of the best studied free-living organisms. A group of E. coli called Extraintestinal pathogenic E. coli (ExPEC) including both human and animal pathogens like Uropathogenic E. coli (UPEC), Newborn meningitic E. coli (NMEC) and Avian pathogenic E. coli (APEC), have been found to harbour many fimbriae including Type 1 fimbriae, P fimbriae, curli fibres, S fimbriae, F1C fimbriae, Dr fimbriae, afimbrial adhesins, temperature-sensitive haemagglutinin and many novel adhesin gene clusters that have not yet been characterized. Each of these adhesins is unique due to the recognition of an adhesin-specific receptor, though as a group these adhesins share common genomic organization. A newly identified putative adhesin temporarily termed ExPEC Adhesin I, encoded by gene yqi, has been recently found to play a significant role in the pathogenesis of APEC infection, thus making it an interesting candidate for future research. The aim of this review is to describe the role of ExPEC adhesins during extraintestinal infections known till date, and to suggest the idea of investigating their potential role in the colonization of the host gut which is said to be a reservoir for ExPEC.

  3. Experimental Escherichia coli O157:H7 carriage in calves.

    PubMed Central

    Brown, C A; Harmon, B G; Zhao, T; Doyle, M P

    1997-01-01

    Nine weaned calves (6 to 8 weeks of age) were given 10(10) CFU of a five-strain mixture of enterohemorrhagic Escherichia coli O157:H7 by oral-gastric intubation. After an initial brief period of pyrexia in three calves and transient mild diarrhea in five calves, calves were clinically normal throughout the 13- to 27-day study. The population of E. coli O157:H7 in the faces decreased dramatically in all calves during the first 2 weeks after inoculation. Thereafter, small populations of E. coli O157:H7 persisted in all calves, where they were detected intermittently in the feces and rumen contents. While withholding food increased fecal shedding of E. coli O157:H7 by 1 to 2 log10/g in three of four calves previously shedding small populations of E. coli O157:H7, the effect of fasting on fecal shedding of E. coli O157:H7 was variable in calves shedding larger populations. At necropsy, E. coli O157:H7 was not isolated from sites outside the alimentary tract. E. coli O157:H7 was isolated from the forestomach or colon of all calves at necropsy. Greater numbers of E. coli O157:H7 were present in the gastrointestinal contents than in the corresponding mucosal sections, and there was no histologic or immunohistochemical evidence of E. coli O157:H7 adhering to the mucosa. In conclusion, under these experimental conditions, E. coli O157:H7 is not pathogenic in weaned calves, and while it does not appear to colonize mucosal surfaces for extended periods, E. coli O157:H7 persists in the contents of the rumen and colon as a source for fecal shedding. PMID:8979335

  4. Travelers' diarrhea and toxigenic Escherichia coli.

    PubMed

    Gorbach, S L; Kean, B H; Evans, D G; Evans, D J; Bessudo, D

    1975-05-01

    In a group of 133 United States students studied for 18 days after arriving in Mexico, diarrhea developed in 38 (29 per cent). Diarrhea rarely began before the fourth day, and the mean onset was 13 days after arrival. Symptoms lasted an average of 3.4 days but persisted in 21 per cent of sick students. Heat-labile enterotoxin-producing Escheria coli was found in the stools of 72 per cent of sick and 15 per cent of healthy students. None had heat-labile Esch. coli when they entered Mexico. The incubation period was short, generally 24 to 48 hours, and the carrier state was five days or less in 82 per cent of students surveyed. Entamoeba histolytica was found in 6 per cent of cases of diarrhea, but not salmonella, shigella or penetrating Esch. coli. These studies suggest that approximately 70 per cent of travelers' diarrhea in Mexico is associated with heat-labile toxigenic strains of Esch. coli.

  5. In vivo formation of glutamyl-tRNA(Gln) in Escherichia coli by heterologous glutamyl-tRNA synthetases.

    PubMed

    Núñez, Harold; Lefimil, Claudia; Min, Bokkee; Söll, Dieter; Orellana, Omar

    2004-01-16

    Two types of glutamyl-tRNA synthetase exist: the discriminating enzyme (D-GluRS) forms only Glu-tRNA(Glu), while the non-discriminating one (ND-GluRS) also synthesizes Glu-tRNA(Gln), a required intermediate in protein synthesis in many organisms (but not in Escherichia coli). Testing the capacity to complement a thermosensitive E. coli gltX mutant and to suppress an E. coli trpA49 missense mutant we examined the properties of heterologous gltX genes. We demonstrate that while Acidithiobacillus ferrooxidans GluRS1 and Bacillus subtilis Q373R GluRS form Glu-tRNA(Glu), A. ferrooxidans and Helicobacter pylori GluRS2 form Glu-tRNA(Gln) in E. coli in vivo.

  6. Genomic Comparative Study of Bovine Mastitis Escherichia coli

    PubMed Central

    Kempf, Florent; Slugocki, Cindy; Blum, Shlomo E.; Leitner, Gabriel; Germon, Pierre

    2016-01-01

    Escherichia coli, one of the main causative agents of bovine mastitis, is responsible for significant losses on dairy farms. In order to better understand the pathogenicity of E. coli mastitis, an accurate characterization of E. coli strains isolated from mastitis cases is required. By using phylogenetic analyses and whole genome comparison of 5 currently available mastitis E. coli genome sequences, we searched for genotypic traits specific for mastitis isolates. Our data confirm that there is a bias in the distribution of mastitis isolates in the different phylogenetic groups of the E. coli species, with the majority of strains belonging to phylogenetic groups A and B1. An interesting feature is that clustering of strains based on their accessory genome is very similar to that obtained using the core genome. This finding illustrates the fact that phenotypic properties of strains from different phylogroups are likely to be different. As a consequence, it is possible that different strategies could be used by mastitis isolates of different phylogroups to trigger mastitis. Our results indicate that mastitis E. coli isolates analyzed in this study carry very few of the virulence genes described in other pathogenic E. coli strains. A more detailed analysis of the presence/absence of genes involved in LPS synthesis, iron acquisition and type 6 secretion systems did not uncover specific properties of mastitis isolates. Altogether, these results indicate that mastitis E. coli isolates are rather characterized by a lack of bona fide currently described virulence genes. PMID:26809117

  7. Abundance of culturable versus viable Escherichia coli in freshwater.

    PubMed

    Servais, Pierre; Prats, Josué; Passerat, Julien; Garcia-Armisen, Tamara

    2009-07-01

    Approved methods traditionally used for Escherichia coli enumeration in waters are culture-based. However, these methods can underestimate the E. coli abundance in aquatic systems because they do not take into account cells that remain viable but have lost the ability to grow in or on culture media. We investigated, in freshwater samples, the abundance of (i) culturable E. coli, enumerated by the most probable number microplate method and (ii) viable E. coli, estimated using a procedure called DVC-FISH, which couples fluorescent in situ hybridization (FISH) and a viability testing technique (direct viable count (DVC)). The ratio of culturable to viable E. coli was close to 1 in highly contaminated waters (samples with a high concentration of culturable E. coli), but decreased drastically for weakly contaminated samples. This indicates a large fraction of viable but nonculturable (VBNC) E. coli in the latter samples. Microcosm experiments showed that some environmental factors, such as nutrient scarcity and solar irradiation, could lead to the presence of a high proportion of VBNC E. coli.

  8. Estimation of Escherichia coli in raw ground beef.

    PubMed Central

    Stiles, M E; Ng, L K

    1980-01-01

    This study was undertaken to establish and evaluate more rapid methods of estimating Escherichia coli in ground beef than the standard most probable number (MPN) technique. Direct inoculation of and modifications to EC medium gave unreliable estimates of the presumptive E. coli count. Solid media incubated at an elevated temperature were compared to the MPN technique. Anderson and Baird-Parker's tryptone bile agar (TBA) method and prepoured plates of Endo, Levine eosin methylene blue (EMB), and violet red bile (VRBA) agars incubated at 44 degree C gave equivalent counts to the standard MPN method. Anderson and Baird-Parker TBA was the most selective solid medium for E. coli estimation, but all selective media incubated at elevated temperature reduced apparent E. coli counts by as much as 50%. Indole-producing and lactose-fermenting Enterobacteriaceae, capable of growth at elevated temperature, were tested for their growth on TBA, EMB, and VRBA at elevated temperature. TBA was selective for E. coli biotype I compared to other Enterobacteriaceae that predominate in meats. VRBA and EMB incubated at elevated temperature were not as selective as TBA, but differences in colonies could be observed between typical E. coli colonies and other Enterobacteriaceae on these media. Therefore, VRBA incubated at elevated temperature is proposed as a quality assurance screening test for presumptive E. coli in ground meat. Resuscitation techniques and prepoured plates with VRBA increased recovery levels of presumptive E. coli, but, under the conditions of this study, not to levels that represented a significant practical difference. PMID:7008695

  9. Genomic Comparative Study of Bovine Mastitis Escherichia coli.

    PubMed

    Kempf, Florent; Slugocki, Cindy; Blum, Shlomo E; Leitner, Gabriel; Germon, Pierre

    2016-01-01

    Escherichia coli, one of the main causative agents of bovine mastitis, is responsible for significant losses on dairy farms. In order to better understand the pathogenicity of E. coli mastitis, an accurate characterization of E. coli strains isolated from mastitis cases is required. By using phylogenetic analyses and whole genome comparison of 5 currently available mastitis E. coli genome sequences, we searched for genotypic traits specific for mastitis isolates. Our data confirm that there is a bias in the distribution of mastitis isolates in the different phylogenetic groups of the E. coli species, with the majority of strains belonging to phylogenetic groups A and B1. An interesting feature is that clustering of strains based on their accessory genome is very similar to that obtained using the core genome. This finding illustrates the fact that phenotypic properties of strains from different phylogroups are likely to be different. As a consequence, it is possible that different strategies could be used by mastitis isolates of different phylogroups to trigger mastitis. Our results indicate that mastitis E. coli isolates analyzed in this study carry very few of the virulence genes described in other pathogenic E. coli strains. A more detailed analysis of the presence/absence of genes involved in LPS synthesis, iron acquisition and type 6 secretion systems did not uncover specific properties of mastitis isolates. Altogether, these results indicate that mastitis E. coli isolates are rather characterized by a lack of bona fide currently described virulence genes.

  10. [Acute diarrheal disease caused by enteropathogenic Escherichia coli in Colombia].

    PubMed

    Gómez-Duarte, Oscar G

    2014-10-01

    Intestinal Escherichia coli pathogens are leading causes of acute diarrheal disease in children less than 5 years in Latin America, Africa and Asia and a leading cause of death in children living in poorest communities in Africa and South East Asia. Studies on the role of E. coli pathogens in childhood diarrhea in Colombia and other countries in Latin America are limited due to the lack of detection assays in clinical laboratories at the main urban medical centers. Recent studies report that enterotoxigenic E. coli is the most common E. coli pathogens associated with diarrhea in children less than 5 years of age. Other E. coli pathotypes have been detected in children with diarrhea including enteropathogenic, enteroaggregative, shiga-toxin producing and diffusely adherent E. coli. It was also found that meat and vegetables at retail stores are contaminated with Shiga-toxin producing E. coli and enteroaggregative E. coli, suggesting that food products are involved in transmission and infection of the susceptible host. More studies are necessary to evaluate the mechanisms of transmission, the impact on the epidemiology of diarrheal disease, and management strategies and prevention of these pathogens affecting the pediatric population in Colombia.

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

    PubMed

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

    2017-01-01

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

  12. Yeast DNA sequences initiating gene expression in Escherichia coli.

    PubMed

    Lewin, Astrid; Tran, Thi Tuyen; Jacob, Daniela; Mayer, Martin; Freytag, Barbara; Appel, Bernd

    2004-01-01

    DNA transfer between pro- and eukaryotes occurs either during natural horizontal gene transfer or as a result of the employment of gene technology. We analysed the capacity of DNA sequences from a eukaryotic donor organism (Saccharomyces cerevisiae) to serve as promoter region in a prokaryotic recipient (Escherichia coli) by creating fusions between promoterless luxAB genes from Vibrio harveyi and random DNA sequences from S. cerevisiae and measuring the luminescence of transformed E. coli. Fifty-four out of 100 randomly analysed S. cerevisiae DNA sequences caused considerable gene expression in E. coli. Determination of transcription start sites within six selected yeast sequences in E. coli confirmed the existence of bacterial -10 and -35 consensus sequences at appropriate distances upstream from transcription initiation sites. Our results demonstrate that the probability of transcription of transferred eukaryotic DNA in bacteria is extremely high and does not require the insertion of the transferred DNA behind a promoter of the recipient genome.

  13. Proton-linked D-xylose transport in Escherichia coli.

    PubMed Central

    Lam, V M; Daruwalla, K R; Henderson, P J; Jones-Mortimer, M C

    1980-01-01

    The addition of xylose to energy-depleted cells of Escherichia coli elicited an alkaline pH change which failed to appear in the presence of uncoupling agents. Accumulation of [14C]xylose by energy-replete cells was also inhibited by uncoupling agents, but not by fluoride or arsenate. Subcellular vesicles of E. coli accumulated [14C]xylose provided that ascorbate plus phenazine methosulfate were present for respiration, and this accumulation was inhibited by uncoupling agents or valinomycin. Therefore, the transport of xylose into E. coli appears to be energized by a proton-motive force, rather than by a phosphotransferase or directly energized mechanism. Its specificity for xylose as inducer and substrate and the genetic location of a xylose-H+ transport-negative mutation near mtl showed that the xylose-H+ system is distinct from other proton-linked sugar transport systems of E. coli. PMID:6995439

  14. Escherichia coli control in a surface flow treatment wetland.

    PubMed

    MacIntyre, M E; Warner, B G; Slawson, R M

    2006-06-01

    A field experiment showed that numbers of Escherichia coli declined significantly when floating Lemna spp. plants were removed to create open water areas in a typical newly constructed surface flow treatment wetland in southern Ontario. It is suggested that E. coli declined immediately after Lemna removal because the Lemna was shading the water column from penetration by natural UV radiation, it was providing favourable attachment sites for the E. coli, and it was not allowing effective free exchange of oxygen from surface winds to the water column to maintain high enough dissolved oxygen supplies for predator zooplankton populations. Operators of wetland systems must have the specialized skills required to recognize the cause and the appropriate maintenance requirements to maintain efficient operation of such unconventional systems should E. coli numbers increase during the course of operation.

  15. Biosynthesis of Two Flavones, Apigenin and Genkwanin, in Escherichia coli.

    PubMed

    Lee, Hyejin; Kim, Bong Gyu; Kim, Mihyang; Ahn, Joong-Hoon

    2015-09-01

    The flavonoid apigenin and its O-methyl derivative, genkwanin, have various biological activities and can be sourced from some vegetables and fruits. Microorganisms are an alternative for the synthesis of flavonoids. Here, to synthesize genkwanin from tyrosine, we first synthesized apigenin from p-coumaric acid using four genes (4CL, CHS, CHI, and FNS) in Escherichia coli. After optimization of different combinations of constructs, the yield of apigenin was increased from 13 mg/l to 30 mg/l. By introducing two additional genes (TAL and POMT7) into an apigenin-producing E. coli strain, we were able to synthesize 7-O-methyl apigenin (genkwanin) from tyrosine. In addition, the tyrosine content in E. coli was modulated by overexpressing aroG and tyrA. The engineered E. coli strain synthesized approximately 41 mg/l genkwanin.

  16. Escherichia coli early-onset sepsis: trends over two decades.

    PubMed

    Mendoza-Palomar, Natalia; Balasch-Carulla, Milena; González-Di Lauro, Sabina; Céspedes, Maria Concepció; Andreu, Antònia; Frick, Marie Antoinette; Linde, Maria Ángeles; Soler-Palacin, Pere

    2017-08-02

    Escherichia coli early-onset sepsis (EOS) is an important cause of mortality and morbidity in neonates, especially in preterm and very low birth weight (VLBW) newborns. The aim of our study was to evaluate potential changes in the clinical and microbiological characteristics of E. coli EOS in our setting. Epidemiological, clinical, and microbiological data from all neonates with proven E. coli EOS from January 1994 to December 2014 were retrospectively collected in a single tertiary care hospital in Barcelona (Spain). Seventy-eight E. coli EOS cases were analyzed. A slight increase in the incidence of E. coli EOS was observed during the study period. VLBW newborns remained the group with higher incidence (10.4 cases per 1000 live births) and mortality (35.3%). Systematic use of PCR increased E. coli EOS diagnosis, mainly in the term newborn group. There was an increase in resistant E. coli strains causing EOS, with especially high resistance to ampicillin and gentamicin (92.8 and 28.6%, respectively). Nonetheless, resistant strains were not associated with poorer clinical outcomes. There is an urgent need to reconsider the empirical therapy used in neonatal EOS, particularly in VLBW newborns. What is Known: • E. coli early-onset sepsis (EOS) and E. coli resistant strains have been described as overall stable but increasing in VLBW neonates (< 1.500 g) in previous studies. What is New: • Our study shows an increasing incidence of E. coli EOS in all age groups, overruling group B Streptoccocus for the last 10 years. E. coli resistant strains also increased equally in all age groups, with high resistance rates to our first line antibiotics (ampicillin and gentamicin). • Empiric antibiotic therapy of EOS, mainly in VLBW newborns, should be adapted to this new scenario.

  17. Compilation of DNA sequences of Escherichia coli

    PubMed Central

    Kröger, Manfred

    1989-01-01

    We have compiled the DNA sequence data for E.coli K12 available from the GENBANK and EMBO databases and over a period of several years independently from the literature. We have introduced all available genetic map data and have arranged the sequences accordingly. As far as possible the overlaps are deleted and a total of 940,449 individual bp is found to be determined till the beginning of 1989. This corresponds to a total of 19.92% of the entire E.coli chromosome consisting of about 4,720 kbp. This number may actually be higher by some extra 2% derived from the sequence of lysogenic bacteriophage lambda and the various insertion sequences. This compilation may be available in machine readable form from one of the international databanks in some future. PMID:2654890

  18. Resistance and virulence factors of Escherichia coli isolated from chicken.

    PubMed

    Pavlickova, Silvie; Dolezalova, Magda; Holko, Ivan

    2015-01-01

    Chicken meat has become an important part of the human diet and besides contamination by pathogenic Escherichia coli there is a risk of antibiotic resistance spreading via the food chain. The purpose of this study was to examine the prevalence of resistance against eight antibiotics and the presence of 14 virulence factors among 75 Escherichia coli strains isolated from chicken meat in the Czech Republic after classification into phylogenetic groups by the multiplex PCR method. More than half of strains belonged to A phylogroup, next frequently represented was B1 phylogroup, which suggests the commensal strains. The other strains were classified into phylogroups B2 and D, which had more virulence factors. Almost half of all E. coli strains were resistant to at least one of eight-tested antibiotics. A multidrug resistance was observed in 13% of strains. The most prevalent virulence genes were iucD, iss and tsh. None of genes encoding toxins was detected. Most of E. coli strains isolated from chicken meat can be considered as nonpathogenic on the basis of analysis of virulence factors, antibiotic resistance and phylogroups assignment. It can provide a useful tool for prediction of a potential risk from food contaminated by E. coli.

  19. Escherichia coli as a model active colloid: A practical introduction.

    PubMed

    Schwarz-Linek, Jana; Arlt, Jochen; Jepson, Alys; Dawson, Angela; Vissers, Teun; Miroli, Dario; Pilizota, Teuta; Martinez, Vincent A; Poon, Wilson C K

    2016-01-01

    The flagellated bacterium Escherichia coli is increasingly used experimentally as a self-propelled swimmer. To obtain meaningful, quantitative results that are comparable between different laboratories, reproducible protocols are needed to control, 'tune' and monitor the swimming behaviour of these motile cells. We critically review the knowledge needed to do so, explain methods for characterising the colloidal and motile properties of E. coli cells, and propose a protocol for keeping them swimming at constant speed at finite bulk concentrations. In the process of establishing this protocol, we use motility as a high-throughput probe of aspects of cellular physiology via the coupling between swimming speed and the proton motive force.

  20. PROPERTIES OF A BACTERIOPHAGE DERIVED FROM ESCHERICHIA COLI K235

    PubMed Central

    Jesaitis, Margeris A.; Hutton, John J.

    1963-01-01

    A temperate bacteriophage was isolated from the colicinogenic strain of Escherichia coli K235 and characterized. This phage, termed PK, is related to P2 virus morphologically, serologically, and, possibly, genetically and it bears no relationship to the T-even phages. It was also demonstrated that PK virus and colicine K differ both in their host range and in their immunological specificity, and that PK prophage does not induce the colicinogenesis in its host bacterium. It was concluded that the formation of colicine K. and PK phage in E. coli K235 are controlled by different genetic determinants. PMID:14029160

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

    PubMed

    Riley, M; Space, D B

    1996-01-01

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

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

    PubMed Central

    Riley, M; Space, D B

    1996-01-01

    GenProtEc is a database of Escherichia coli genes and their gene products, classified by type of function and physiological role and with citations to the literature for each. Also present are data on sequence similarities among E.coli proteins with PAM values, percent identity of amino acids, length of alignment and percent aligned. The database is available as a PKZip file by ftp from mbl.edu/pub/ecoli.exe. The program runs under MS-DOS on IMB-compatible machines. GenProtEc can also be accessed through the World Wide Web at URL http://mbl.edu/html/ecoli.html. PMID:8594596

  3. Functional role of bdm during flagella biogenesis in Escherichia coli.

    PubMed

    Kim, Ji-Sun; Kim, Yu Jin; Seo, Sojin; Seong, Maeng-Je; Lee, Kangseok

    2015-03-01

    The biofilm-dependent modulation gene (bdm) has recently been shown to play a role in osmotic-induced formation of biofilm in Escherichia coli. In this study, we demonstrated that deletion of bdm results in down-regulation of flagella biosynthesis genes and, consequently, a defect in E. coli motility. In addition, we employed atomic force microscopy to confirm the absence of flagella-like structures on the surface of bdm-null cells. These findings indicate that bdm plays a key role in regulatory pathway for the formation of flagella.

  4. Nitric oxide donor-mediated killing of bioluminescent Escherichia coli.

    PubMed Central

    Virta, M; Karp, M; Vuorinen, P

    1994-01-01

    The antimicrobial activities of two nitric oxide-releasing compounds against Escherichia coli were investigated by using recombinant E. coli cloned with a luciferase gene from Pyrophorus plagiophthalamus. Since luciferase uses intracellular ATP to generate visible light which can be measured from living cells in real time, we wanted to compare the extent to which cell viability parallels light emission. Results from luminescence measurements and CFU counts were in good agreement, and the decrease in light emission was shown to provide a rapid and more sensitive indication of cytotoxicity. PMID:7695261

  5. Bacterial self-defence: how Escherichia coli evades serum killing.

    PubMed

    Miajlovic, Helen; Smith, Stephen G

    2014-05-01

    The ability to survive the bactericidal action of serum is advantageous to extraintestinal pathogenic Escherichia coli that gain access to the bloodstream. Evasion of the innate defences present in serum, including complement and antimicrobial peptides, involves multiple factors. Serum resistance mechanisms utilized by E. coli include the production of protective extracellular polysaccharide capsules and expression of factors that inhibit or interfere with the complement cascade. Recent studies have also highlighted the importance of structural integrity of the cell envelope in serum survival. These survival strategies are outlined in this review with particular attention to novel findings and recent insights into well-established resistance mechanisms.

  6. Accelerated glycerol fermentation in Escherichia coli using methanogenic formate consumption.

    PubMed

    Richter, Katrin; Gescher, Johannes

    2014-06-01

    Escherichia coli can ferment glycerol anaerobically only under very defined restrictive conditions. Hence, it was the aim of this study to overcome this limitation via a co-cultivation approach. Anaerobic glycerol fermentation by a pure E. coli culture was compared to a co-culture that also contained the formate-oxidizing methanogen Methanobacterium formicicum. Co-cultivation of the two strains led to a more than 11-fold increased glycerol consumption. Furthermore, it supported a constantly neutral pH and a shift from ethanol to succinate production. Moreover, M. formicicum was analyzed for its ability to grow on different standard media and a surprising versatility could be demonstrated.

  7. Recombinant protein expression in Escherichia coli: advances and challenges

    PubMed Central

    Rosano, Germán L.; Ceccarelli, Eduardo A.

    2014-01-01

    Escherichia coli is one of the organisms of choice for the production of recombinant proteins. Its use as a cell factory is well-established and it has become the most popular expression platform. For this reason, there are many molecular tools and protocols at hand for the high-level production of heterologous proteins, such as a vast catalog of expression plasmids, a great number of engineered strains and many cultivation strategies. We review the different approaches for the synthesis of recombinant proteins in E. coli and discuss recent progress in this ever-growing field. PMID:24860555

  8. Ribitol and D-arabitol catabolism in Escherichia coli.

    PubMed Central

    Scangos, G A; Reiner, A M

    1978-01-01

    In Escherichia coli C, the catabolism of the pentitols ribitol and D-arabitol proceeds through separate, inducible operons, each consisting of a dehydrogenase and a kinase. The ribitol operon is induced in response to ribulose, and the D-arabitol operon is induced in response to D-arabitol. Each operon is under negative control. The genes of the ribitol and D-arabitol operons are very closely linked and lie in a mirror image arrangement, rtlB-rtlA-rtlC-atlC-atlA-atlB, between metG and his on the E. coli chromosome. PMID:350825

  9. Further studies of Escherichia coli in babies after normal delivery

    PubMed Central

    Bettelheim, K. A.; Teoh-Chan, Ching Haan; Chandler, Mary E.; O'Farrell, Sheila M.; Rahamin, Layla; Shaw, Elizabeth J.; Shooter, R. A.

    1974-01-01

    Previous work showed that on the basis of O serotyping alone of Escherichia coli, the majority of babies acquired the same O serotype as was found in the stools of their respective mothers. Further characterization of the E. coli by H serotyping, determination of their antibiotic resistance and ability to ferment six carbohydrates showed that in the majority of cases the previous results were confirmed. In a minority of cases this further testing showed that the strains were not identical. In some instances a number of strains isolated from the same pair showed different combinations of the markers used. PMID:4608224

  10. Metabolic engineering of Escherichia coli for 1-butanol production.

    PubMed

    Atsumi, Shota; Cann, Anthony F; Connor, Michael R; Shen, Claire R; Smith, Kevin M; Brynildsen, Mark P; Chou, Katherine J Y; Hanai, Taizo; Liao, James C

    2008-11-01

    Compared to ethanol, butanol offers many advantages as a substitute for gasoline because of higher energy content and higher hydrophobicity. Typically, 1-butanol is produced by Clostridium in a mixed-product fermentation. To facilitate strain improvement for specificity and productivity, we engineered a synthetic pathway in Escherichia coli and demonstrated the production of 1-butanol from this non-native user-friendly host. Alternative genes and competing pathway deletions were evaluated for 1-butanol production. Results show promise for using E. coli for 1-butanol production.

  11. Inducible repair of oxidative DNA damage in Escherichia coli.

    PubMed

    Demple, B; Halbrook, J

    Hydrogen peroxide is lethal to many cell types, including the bacterium Escherichia coli. Peroxides yield transient radical species that can damage DNA and cause mutations. Such partially reduced oxygen species are occasionally released during cellular respiration and are generated by lethal and mutagenic ionizing radiation. Because cells live in an environment where the threat of oxidative DNA damage is continual, cellular mechanisms may have evolved to avoid and repair this damage. Enzymes are known which evidently perform these functions. We report here that resistance to hydrogen peroxide toxicity can be induced in E. coli, that this novel induction is specific and occurs, in part, at the level of DNA repair.

  12. Sedimentation and gravitational instability of Escherichia coli Suspension

    NASA Astrophysics Data System (ADS)

    Douarche, Carine; Salin, Dominique; Collaboration between Laboratory FAST; LPS Collaboration

    2016-11-01

    The successive run and tumble of Escherichia coli bacteria provides an active matter suspension of rod-like particles with a large swimming diffusion. As opposed to inactive elongated particles, this diffusion prevents clustering and instability in the gravity field. We measure the time dependent E . coli concentration profile during their sedimentation. After some hours, due to the dioxygen consumption, a motile / non-motile front forms leading to a Rayleigh-Taylor type gravitational instability. Analyzing both sedimentation and instability in the framework of active particle suspensions, we can measure the relevant bacteria hydrodynamic characteristics such as its single particle sedimentation velocity and its hindrance volume.

  13. [Drug resistance of Escherichia coli strains isolated from poultry].

    PubMed

    Giurov, B; Korudzhiĭski, N; Bineva, I

    1981-01-01

    Studied was the sensitivity of a total of 143 strains of Escherichia coli, isolated from young birds and broilers died from coli septicaemia, to antibiotics and chemotherapeutics. The following descending order was established: gentamycin, carbenicillin, ampicillin, furazolidon, borgal, kanamycin, strep tomycin, chloramphenicol, neomycin sulphathiazole, and tetracycline. Markers of resistance were established with all strains with regard to the therapeutic agents in current and prospective use in industrial poultry farming. It is stated that a preliminary antibiogram is indispensable in order to obtain dependable results in the treatment of animals affected with colibacteriosis. An alternative is to apply directly those drugs to which the strains have shown highest sensitivity.

  14. Advances in molecular serotyping and subtyping of Escherichia coli

    DOE PAGES

    Fratamico, Pina M.; DebRoy, Chitrita; Liu, Yanhong; ...

    2016-05-03

    Escherichia coli plays an important role as a member of the gut microbiota; however, pathogenic strains also exist, including various diarrheagenic E. coli pathotypes and extraintestinal pathogenic E. coli that cause illness outside of the GI-tract. E. coli have traditionally been serotyped using antisera against the ca. 186 O-antigens and 53 H-flagellar antigens. Phenotypic methods, including bacteriophage typing and O- and H- serotyping for differentiating and characterizing E. coli have been used for many years; however, these methods are generally time consuming and not always accurate. Advances in next generation sequencing technologies have made it possible to develop genetic-based subtypingmore » and molecular serotyping methods for E. coli, which are more discriminatory compared to phenotypic typing methods. Furthermore, whole genome sequencing (WGS) of E. coli is replacing established subtyping methods such as pulsedfield gel electrophoresis, providing a major advancement in the ability to investigate food-borne disease outbreaks and for trace-back to sources. Furthermore, a variety of sequence analysis tools and bioinformatic pipelines are being developed to analyze the vast amount of data generated by WGS and to obtain specific information such as O- and H-group determination and the presence of virulence genes and other genetic markers.« less

  15. Inactivation of Escherichia coli using atmospheric-pressure plasma jet

    NASA Astrophysics Data System (ADS)

    Kuwahata, Hiroshi; Yamaguchi, Takeshi; Ohyama, Ryu-ichiro; Ito, Atsushi

    2015-01-01

    An atmospheric-pressure argon (Ar) plasma jet was applied to the inactivation of Escherichia coli. The Ar plasma jet was generated at a frequency of 10 kHz, an applied voltage of 10 kV, and an Ar gas flow rate of 10 L/min at atmospheric pressure. E. coli cells seeded on an agar medium in a Petri dish were inactivated by Ar plasma jet irradiation for 1 s. Scanning electron microscopy (SEM) revealed that E. coli cells were killed because their cell wall and membrane were disrupted. To determine the causes of the disruption of the cell wall and membrane of E. coli, we performed the following experiments: the measurement of the surface temperature of an agar medium using a thermograph, the analysis of an emission spectrum of a plasma jet obtained using a multichannel spectrometer, and the determination of the distribution of the concentration of hydrogen peroxide (H2O2) generated on an agar medium by plasma jet irradiation using semiquantitative test strips. Moreover, H2O2 solutions of different concentrations were dropped onto an agar medium seeded with E. coli cells to examine the contribution of H2O2 to the death of E. coli. The results of these experiments showed that the cell wall and membrane of E. coli were disrupted by electrons in the plasma jet, as well as by electroneutral excited nitrogen molecules (N2) and hydroxyl (OH) radicals in the periphery of the plasma jet.

  16. Gentamicin resistance among Escherichia coli strains isolated in neonatal sepsis.

    PubMed

    Hasvold, J; Bradford, L; Nelson, C; Harrison, C; Attar, M; Stillwell, T

    2013-01-01

    Neonatal sepsis is a significant cause of morbidity and mortality among term and preterm infants. Ampicillin and gentamicin are standard empiric therapy for early onset sepsis. Four cases of neonatal sepsis secondary to Escherichia coli (E. coli) found to be gentamicin resistant occurred within a five week period in one neonatal intensive care unit (NICU). To determine whether these cases could be tied to a single vector of transmission, and to more broadly evaluate the incidence of gentamicin resistant strains of E. coli in the neonatal population at our institution compared to other centers, we reviewed the charts of the four neonates (Infants A through D) and their mothers. The E. coli isolates were sent for Pulse Field Gel Electrophoresis (PFGE) to evaluate for genetic similarity between strains. We also reviewed all positive E. coli cultures from one NICU over a two year period. Infants A and B had genetically indistinguishable strains which matched that of urine and placental cultures of Infant B's mother. Infant C had a genetically distinct organism. Infant D, the identical twin of Infant C, did not have typing performed. Review of all cultures positive for E. coli at our institution showed a 12.9 percent incidence of gentamicin-resistance. A review of other studies showed that rates of resistance vary considerably by institution. We conclude that gentamicin-resistant E. coli is a relatively uncommon cause of neonatal sepsis, but should remain a consideration in patients who deteriorate despite initiation of empiric antibiotics.

  17. Phosphorylation of streptozotocin during uptake via the phosphoenolpyruvate: sugar phosphotransferase system in Escherichia coli.

    PubMed Central

    Ammer, J; Brennenstuhl, M; Schindler, P; Höltje, J V; Zähner, H

    1979-01-01

    Mutants of Escherichia coli K-12, Staphylococcus aureus, and Bacillus subtilis defective in the general components (enzyme I, or HPr, or both) of the phosphoenolpyruvate:sugar phosphotransferase system are shown to be resistant to the antibiotic streptozotocin. It is shown here, employing 32P-labeled phosphoenolpyruvate, that wild-type cells of E. coli phosphorylate streptozotocin, whereas with a phosphotransferase system-defective mutant of E. coli the drug is recovered in an unaltered, free form. The internal accumulation of streptozotocin at the steady-state level was about 70 times that of the concentration in the external medium. The antibacterial action of streptozotocin, as well as the uptake of the drug, was inhibited by N-acetyl-D-glucosamine. The uptake of the antibiotic was extremely sensitive to p-chloromercuribenzoate. It is concluded that streptozotocin is taken up by E. coli via the phosphoenolpyruvate:sugar phosphotransferase system and consequently accumulates in the cell at first as streptozotocin-phosphate. PMID:161156

  18. Penem derivatives: beta-lactamase stability and affinity for penicillin-binding proteins in Escherichia coli.

    PubMed

    Ohya, S; Utsui, Y; Sugawara, S; Yamazaki, M

    1982-03-01

    Penem derivatives, a new group of beta-lactam antibiotics with potent activities against a wide range of bacteria, including Pseudomonas aeruginosa, were tested for their stability against hydrolysis by beta-lactamases purified from clinical isolates of Morganella morganii. Proteus vulgaris, and Escherichia coli and by a penicillinase from Bacillus cereus. Penems having 6 alpha substituents, such as hydroxyethyl, hydroxymethyl, and ethyl groups, were very stable against hydrolysis by each of the enzymes. Penems having no 6 alpha substituents were easily hydrolyzed by P. vulgaris and E. coli enzymes, whereas they were rather stable against hydrolysis by M. morganii and B. cereus enzymes, a typical cephalosporinase and penicillinase, respectively. Affinity of the penems for E. coli penicillin-binding proteins (PBPs) was also tested. beta-Lactamase-stable penems having a 6 alpha-hydroxyethyl group showed high affinity for PBP-4, -5, and -6 as well as for PBP-1A, -1Bs, and -2. However, the penems having no 6 alpha substituents showed a far lower affinity for PBP-4, -5, and -6 than that shown by the corresponding 6 alpha-hydroxyethyl penems. Among the penems tested, affinity for PBP-4, -5, and -6 was closely related to their beta-lactamase stability, as was the case among cephamycins and cephalosporins. Effects of the penems on the morphology of a strain of E. coli are also described.

  19. Engineering of Escherichia coli to facilitate efficient utilization of isomaltose and panose in industrial glucose feedstock.

    PubMed

    Abe, Kenji; Kuroda, Akio; Takeshita, Ryo

    2017-03-01

    Industrial glucose feedstock prepared by enzymatic digestion of starch typically contains significant amounts of disaccharides such as maltose and isomaltose and trisaccharides such as maltotriose and panose. Maltose and maltosaccharides can be utilized in Escherichia coli fermentation using industrial glucose feedstock because there is an intrinsic assimilation pathway for these sugars. However, saccharides that contain α-1,6 bonds, such as isomaltose and panose, are still present after fermentation because there is no metabolic pathway for these sugars. To facilitate more efficient utilization of glucose feedstock, we introduced glvA, which encodes phospho-α-glucosidase, and glvC, which encodes a subunit of the phosphoenolpyruvate-dependent maltose phosphotransferase system (PTS) of Bacillus subtilis, into E. coli. The heterologous expression of glvA and glvC conferred upon the recombinant the ability to assimilate isomaltose and panose. The recombinant E. coli assimilated not only other disaccharides but also trisaccharides, including alcohol forms of these saccharides, such as isomaltitol. To the best of our knowledge, this is the first report to show the involvement of the microbial PTS in the assimilation of trisaccharides. Furthermore, we demonstrated that an L-lysine-producing E. coli harboring glvA and glvC converted isomaltose and panose to L-lysine efficiently. These findings are expected to be beneficial for industrial fermentation.

  20. No evidence for a bovine mastitis Escherichia coli pathotype.

    PubMed

    Leimbach, Andreas; Poehlein, Anja; Vollmers, John; Görlich, Dennis; Daniel, Rolf; Dobrindt, Ulrich

    2017-05-08

    Escherichia coli bovine mastitis is a disease of significant economic importance in the dairy industry. Molecular characterization of mastitis-associated E. coli (MAEC) did not result in the identification of common traits. Nevertheless, a mammary pathogenic E. coli (MPEC) pathotype has been proposed suggesting virulence traits that differentiate MAEC from commensal E. coli. The present study was designed to investigate the MPEC pathotype hypothesis by comparing the genomes of MAEC and commensal bovine E. coli. We sequenced the genomes of eight E. coli isolated from bovine mastitis cases and six fecal commensal isolates from udder-healthy cows. We analyzed the phylogenetic history of bovine E. coli genomes by supplementing this strain panel with eleven bovine-associated E. coli from public databases. The majority of the isolates originate from phylogroups A and B1, but neither MAEC nor commensal strains could be unambiguously distinguished by phylogenetic lineage. The gene content of both MAEC and commensal strains is highly diverse and dominated by their phylogenetic background. Although individual strains carry some typical E. coli virulence-associated genes, no traits important for pathogenicity could be specifically attributed to MAEC. Instead, both commensal strains and MAEC have very few gene families enriched in either pathotype. Only the aerobactin siderophore gene cluster was enriched in commensal E. coli within our strain panel. This is the first characterization of a phylogenetically diverse strain panel including several MAEC and commensal isolates. With our comparative genomics approach we could not confirm previous studies that argue for a positive selection of specific traits enabling MAEC to elicit bovine mastitis. Instead, MAEC are facultative and opportunistic pathogens recruited from the highly diverse bovine gastrointestinal microbiota. Virulence-associated genes implicated in mastitis are a by-product of commensalism with the primary function

  1. EFFECT OF VISIBLE RANGE ELECTROMAGNETIC RADIATIONS ON ESCHERICHIA COLI.

    PubMed

    Azeemi, Samina T Yousuf; Shaukat, Saleem Farooq; Azeemi, Khawaja Shamsuddin; Khan, Idrees; Mahmood, Khalid; Naz, Farah

    2017-01-01

    Escherichia coli is the agent responsible for a range of clinical diseases. With emerging antimicrobial resistance, other treatment options including solar/photo-therapy are becoming increasingly common. Visible Range Radiation Therapy/Colour Therapy is an emerging technique in the field of energy/vibrational medicine that uses visible spectrum of Electromagnetic Radiations to cure different diseases. In this study, our goal was to understand the effect of Visible Range Electromagnetic Radiations on E. coli (in vitro) and therefore find out the most appropriate visible range radiation for the treatment of diseases caused by E. coli. A total of 6 non-repetitive E. coli isolates were obtained from urine samples obtained from hospitalized patients with UTI. Single colony of E. coli was inoculated in 3 ml of Lysogeny Broth (LB) and 40 μl of this E. coli suspension was poured into each of the plastic tubes which were then irradiated with six different wavelengths in the visible region (Table. 1) after 18 hours with one acting as a control. The Optical Densities of these irradiated samples were then measured. Furthermore, scanning electron microscopy (TEFCAN ZEGA3) was carried out. The analysis of the microscopic and SEM images of irradiated E. coli samples with six different visible range radiations is representative of The fact that E. coli responded differently to every applied radiation in the visible region and the most profound inhibitory effects were that of 538nm Visible Range Radiation (Green) which proved to be bactericidal and 590nm Visible Range Radiation (yellow) which was bacteriostatic. The enhanced growth of E. coli with varying degrees was clearly observed in 610nm (orange), 644nm (red), 464nm (Purple) and 453nm (blue). It can be concluded that 538nm (Green) and 590nm (Yellow) can effectively be used for treating E. coli borne diseases.

  2. Formation of nonculturable Escherichia coli in drinking water.

    PubMed

    Bjergbaek, L A; Roslev, P

    2005-01-01

    To examine whether incubation of Escherichia coli in nondisinfected drinking water result in development of cells that are not detectable using standard procedures but maintain a potential for metabolic activity and cell division. Survival and detectability of four different E. coli strains were studied using drinking water microcosms and samples from contaminated drinking water wells. Recovery of E. coli was compared using different cultivation-dependent methods, fluorescence in situ hybridization (FISH) using specific oligonucleotide probes, direct viable counts (DVC), and by enumeration of gfp-tagged E. coli (green fluorescent protein, GFP). Two levels of stress responses were observed after incubation of E. coli in nondisinfected drinking water: (i) the presence of cells that were not detected using standard cultivation methods but could be cultivated after gentle resuscitation on nonselective nutrient-rich media, and (ii) the presence of cells that responded to nutrient addition but could only be detected by cultivation-independent methods (DVC, FISH and GFP). Collectively, the experiments demonstrated that incubation for 20-60 days in nondisinfected drinking water resulted in detection of only 0.7-5% of the initial E. coli population using standard cultivation methods, whereas 1-20% could be resuscitated to a culturable state, and 17-49% could be clearly detected using cultivation-independent methods. Resuscitation of stressed E. coli on nonselective nutrient-rich media increased cell counts in drinking water using both traditional (CFU), and cultivation-independent methods (DVC, FISH and GFP). The cultivation-independent methods resulted in detection of 10-20 times more E. coli than the traditional methods. The results indicate that a subpopulation of substrate-responsive but apparent nonculturable E. coli may develop in drinking water during long-term starvation survival. The existence of substrate-responsive but nonculturable cells should be considered

  3. Biosynthesis of phosphatidyl glycerophosphate in Escherichia coli.

    PubMed

    Chang, Y Y; Kennedy, E P

    1967-09-01

    An enzyme (L-glycerol 3-phosphate: CMP phosphatidyltransferase) catalyzing the synthesis of phosphatidyl glycerophosphate from CDP-diglyceride and L-glycerol 3-phosphate has been rendered soluble by treatment of the particulate, membrane-containing fraction of E. coli with Triton X-100 and has been partially purified. The enzyme, devoid of phosphatidyl glycerophosphatase activity, is specific for L-glycerol 3-phosphate and is completely dependent upon added Mg(++) or Mn(++) for activity. It has high affinity for CDP-diglyceride and can be used for the assay of this nucleotide. Other properties of the enzyme are also described.

  4. Specific mistranslation in hisT mutants of Escherichia coli.

    PubMed

    Parker, J

    1982-01-01

    Certain strains of Escherichia coli mistranslate at very high frequencies when starved for asparagine or histidine. This mistranslation is the result of misreading events on the ribosome. The introduction of a hisT mutation into such a strain decreases the frequency of mistranslation during histidine starvation but not during asparagine starvation. The most likely explanation is that the replacement of the pseudouridine residue in the anticodon loop of glutamine specific transfer ribonucleic acid by uridine in hisT mutants leads to an increase in fidelity of transfer ribonucleic acid function. The hisT gene in Escherichia coli has also been more accurately mapped, giving the gene order purF-hisT-aroC-fadL-dsdA.

  5. Mechanobiology of Antimicrobial Resistant Escherichia coli and Listeria innocua.

    PubMed

    Tajkarimi, Mehrdad; Harrison, Scott H; Hung, Albert M; Graves, Joseph L

    2016-01-01

    A majority of antibiotic-resistant bacterial infections in the United States are associated with biofilms. Nanoscale biophysical measures are increasingly revealing that adhesive and viscoelastic properties of bacteria play essential roles across multiple stages of biofilm development. Atomic Force Microscopy (AFM) applied to strains with variation in antimicrobial resistance enables new opportunities for investigating the function of adhesive forces (stickiness) in biofilm formation. AFM force spectroscopy analysis of a field strain of Listeria innocua and the strain Escherichia coli K-12 MG1655 revealed differing adhesive forces between antimicrobial resistant and nonresistant strains. Significant increases in stickiness were found at the nanonewton level for strains of Listeria innocua and Escherichia coli in association with benzalkonium chloride and silver nanoparticle resistance respectively. This advancement in the usage of AFM provides for a fast and reliable avenue for analyzing antimicrobial resistant cells and the molecular dynamics of biofilm formation as a protective mechanism.

  6. LYSIS OF ESCHERICHIA COLI BY SULFHYDRYL-BINDING REAGENTS

    PubMed Central

    Schaechter, M.; Santomassino, Katherine A.

    1962-01-01

    Schaechter, M. (College of Medicine, University of Florida, Gainesville) and K. Santomassino. Lysis of Escherichia coli by sulfhydryl-binding reagents. J. Bacteriol. 84:318–325. 1962—Washed suspensions of gram-negative rods were lysed by low concentrations of some sulfhydryl-binding and oxidizing reagents, but not by reducing agents. Some kinetic aspects of this phenomenon were studied with p-chloromercuribenzoate and Escherichia coli B/r. Structures resulting from the action of this reagent consisted of impure cell walls. These could be purified by treatment with trypsin. Cell walls prepared mechanically and cell membranes obtained by lysing protoplasts were not overtly affected by this chemical. Images PMID:14497913

  7. Enterotoxigenic Escherichia coli infection in captive black-footed ferrets.

    PubMed

    Bradley, G A; Orr, K; Reggiardo, C; Glock, R D

    2001-07-01

    Enterotoxigenic Escherichia coli with genes for heat stabile toxins Sta and STb was isolated from the gastrointestinal tract and multiple visceral organs of three adult and three juvenile black-footed ferrets (Mustela nigripes) that died in a captive breeding colony between 24 May 1998 and 2 July 1998. Similar isolates were obtained from rectal swabs of one adult and one juvenile that were clinically ill. All were fed a diet composed of mink chow, raw rabbit meat, beef liver powder, blood meal and lard. Escherichia coli of the same toxin genotype was isolated from the mixed ration. Clinical signs included sudden death, dehydration, anorexia and diarrhea. Necropsy lesions included acute enteritis with large numbers of rod shaped bacteria microscopically visible on intestinal villi.

  8. Growth and Division of Filamentous Forms of Escherichia coli.

    PubMed

    Adler, H I; Hardigree, A A

    1965-07-01

    Adler, Howard I. (Oak Ridge National Laboratory, Oak Ridge, Tenn.), and Alice A. Hardigree. Growth and division of filamentous forms of Escherichia coli. J. Bacteriol. 90:223-226. 1965.-Cells of certain mutant strains of Escherichia coli grow into long multinucleate filaments after exposure to radiation. Deoxyribonucleic acid, ribonucleic acid, and protein synthesis proceed, but cytokinesis does not occur. Cytokinesis (cross-septation) can be initiated by exposure of the filaments to pantoyl lactone or a temperature of 42 C. If growing filaments are treated with mitomycin C, nuclear division does not occur, and nuclear material is confined to the central region of the filament. Cytokinesis cannot be induced in mitomycin C-treated filaments by pantoyl lactone or treatment at 42 C.

  9. TRYPTOPHANASE-TRYPTOPHAN SYNTHETASE SYSTEMS IN ESCHERICHIA COLI III.

    PubMed Central

    Freundlich, Martin; Lichstein, Herman C.

    1962-01-01

    Freundlich, Martin (University of Minnesota, Minneapolis) and Herman C. Lichstein. Tryptophanase-tryptophan synthetase systems in Escherichia coli. III. Requirements for enzyme synthesis. J. Bacteriol. 84:996–1006. 1962.—The requirements for the formation of tryptophanase and tryptophan synthetase in Escherichia coli during repression release were studied. The kinetics of the formation of tryptophan synthetase differed in the two strains examined; this was attributed to differences in the endogenous level of tryptophan in the bacterial cells. The formation of both enzymes was inhibited by chloramphenicol, and by the absence of arginine in an arginine-requiring mutant. These results are indicative of a requirement for protein synthesis for enzyme formation. Requirements for nucleic acid synthesis were examined by use of a uracil- and thymine-requiring mutant, and with purine and pyrimidine analogues. The results obtained suggest that some type of ribonucleic acid synthesis was necessary for the formation of tryptophanase and tryptophan synthetase. PMID:13959620

  10. Mechanobiology of Antimicrobial Resistant Escherichia coli and Listeria innocua

    PubMed Central

    Tajkarimi, Mehrdad; Harrison, Scott H.; Hung, Albert M.; Graves, Joseph L.

    2016-01-01

    A majority of antibiotic-resistant bacterial infections in the United States are associated with biofilms. Nanoscale biophysical measures are increasingly revealing that adhesive and viscoelastic properties of bacteria play essential roles across multiple stages of biofilm development. Atomic Force Microscopy (AFM) applied to strains with variation in antimicrobial resistance enables new opportunities for investigating the function of adhesive forces (stickiness) in biofilm formation. AFM force spectroscopy analysis of a field strain of Listeria innocua and the strain Escherichia coli K-12 MG1655 revealed differing adhesive forces between antimicrobial resistant and nonresistant strains. Significant increases in stickiness were found at the nanonewton level for strains of Listeria innocua and Escherichia coli in association with benzalkonium chloride and silver nanoparticle resistance respectively. This advancement in the usage of AFM provides for a fast and reliable avenue for analyzing antimicrobial resistant cells and the molecular dynamics of biofilm formation as a protective mechanism. PMID:26914334

  11. Thiolases of Escherichia coli: purification and chain length specificities.

    PubMed Central

    Feigenbaum, J; Schulz, H

    1975-01-01

    The presence of only one thiolase (EC 2.3.1.9) in wild-type Escherichia coli induced for enzymes of beta oxidation was demonstrated. A different thiolase was shown to be present in a mutant constitutive for the enzymes of butyrate degradation. The two thiolases were purified to near homogeneity by a simple two-step procedure and were found to be associated with different proteins as shown by gel electrophoresis. The thiolase isolated from induced wild-type Escherichia coli cell was active on beta-ketoacyl-coenzyme A derivatives containing 4 to 16 carbons, but exhibited optimal activity with medium-chain substrates. In contrast, the thiolase isolated from the constitutive mutant was shown to be specific for acetoacetyl-coenzyme A. PMID:236278

  12. Shear alters motility of Escherichia coli

    NASA Astrophysics Data System (ADS)

    Molaei, Mehdi; Jalali, Maryam; Sheng, Jian

    2013-11-01

    Understanding of locomotion of microorganisms in shear flows drew a wide range of interests in microbial related topics such as biological process including pathogenic infection and biophysical interactions like biofilm formation on engineering surfaces. We employed microfluidics and digital holography microscopy to study motility of E. coli in shear flows. We controlled the shear flow in three different shear rates: 0.28 s-1, 2.8 s-1, and 28 s-1 in a straight channel with the depth of 200 μm. Magnified holograms, recorded at 15 fps with a CCD camera over more than 20 minutes, are analyzed to obtain 3D swimming trajectories and subsequently used to extract shear responses of E.coli. Thousands of 3-D bacterial trajectories are tracked. The change of bacteria swimming characteristics including swimming velocity, reorientation, and dispersion coefficient are computed directly for individual trajectory and ensemble averaged over thousands of realizations. The results show that shear suppresses the bacterial dispersions in bulk but promote dispersions near the surface contrary to those in quiescent flow condition. Ongoing analyses are focusing to quantify effect of shear rates on tumbling frequency and reorientation of cell body, and its implication in locating the hydrodynamic mechanisms for shear enhanced angular scattering. NIH, NSF, GoMRI.

  13. Positive regulation of the Escherichia coli glycine cleavage enzyme system.

    PubMed Central

    Wilson, R L; Steiert, P S; Stauffer, G V

    1993-01-01

    A new mutation in Escherichia coli, designated gcvA1, that results in noninducible expression of both gcv and a gcvT-lacZ gene fusion was isolated. A plasmid carrying the wild-type gcvA gene complemented the mutation and restored glycine-inducible gcv and gcvT-lacZ gene expression. These results suggest that gcvA encodes a positive-acting regulatory protein that acts in trans to increase expression of gcv. PMID:8423160

  14. Lipophilic chelator inhibition of electron transport in Escherichia coli.

    PubMed Central

    Crane, R T; Sun, I L; Crane, F L

    1975-01-01

    The lipophilic chelator bathophenanthroline inhibits electron transport in membranes from Escherichia coli. The less lipophilic 1,10-phenanthroline, bathophenanthroline sulfonate, and alpha,alpha-dipyridyl have little effect. Reduced nicotinamide adenine dinucleotide oxidase is more sensitive to bathophenanthroline inhibition than lactate oxidase activity. Evidence for two sites of inhibition comes from the fact that both reduced nicotinamide adenine dinucleotide menadione reductase and duroquinol oxidase activities are inhibited. Addition of uncouplers of phosphorylation before bathophenanthroline protects against inhibition. PMID:1092663

  15. Genome Sequence of Enterotoxigenic Escherichia coli Strain FMU073332.

    PubMed

    Saldaña-Ahuactzi, Zeus; Cruz-Córdova, Ariadnna; Rodea, Gerardo E; Porta, Helena; Navarro-Ocaña, Armando; Eslava-Campos, Carlos; Cevallos, Miguel A; Xicohtencatl-Cortes, Juan

    2017-02-23

    Enterotoxigenic Escherichia coli (ETEC) is an important cause of bacterial diarrheal illness, affecting practically every population worldwide, and was estimated to cause 120,800 deaths in 2010. Here, we report the genome sequence of ETEC strain FMU073332, isolated from a 25-month-old girl from Tlaltizapán, Morelos, México. Copyright © 2017 Saldaña-Ahuactzi et al.

  16. Spurious hydrogen sulfide production by Providencia and Escherichia coli species.

    PubMed Central

    Treleaven, B E; Diallo, A A; Renshaw, E C

    1980-01-01

    Hydrogen sulfide production was noted in two Escherichia coli strands and one Provaidenica alcalifaciens (Proteus inconstans A) strain isolated from clinical stool specimens durin the summer of 1979. An investigation into this phenomenon revealed the predence of Eubacterium lentum, an anaerobe, growing in synergism with the Enterobacteriaceae and producing H2s. The implications of this association are discssed with reference to clinical microbiology laboratory practice. PMID:7000823

  17. Current perspectivesin pathogenesis and antimicrobial resistance of enteroaggregative Escherichia coli.

    PubMed

    Kong, Haishen; Hong, Xiaoping; Li, Xuefen

    2015-08-01

    Enteroaggregative Escherichia coli (EAEC) is an emerging pathogen that causes acute and persistent diarrhea in children and adults. While the pathogenic mechanisms of EAEC intestinal colonization have been uncovered (including bacterial adhesion, enterotoxin and cytotoxin secretion, and stimulation of mucosal inflammation), those of severe extraintestinal infections remain largely unknown. The recent emergence of multidrug resistant EAEC represents an alarming public health threat and clinical challenge, and research on the molecular mechanisms of resistance is urgently needed.

  18. Automated recombinant protein expression screening in Escherichia coli.

    PubMed

    Busso, Didier; Stierlé, Matthieu; Thierry, Jean-Claude; Moras, Dino

    2008-01-01

    To fit the requirements of structural genomics programs, new as well as classical methods have been adapted to automation. This chapter describes the automated procedure developed within the Structural Biology and Genomics Platform, Strasbourg for performing recombinant protein expression screening in Escherichia coli. The procedure consists of parallel competent cells transformation, cell plating, and liquid culture inoculation, implemented for up to 96 samples at a time.

  19. Antibacterial efficacy of silver nanoparticles against Escherichia coli

    NASA Astrophysics Data System (ADS)

    Pattabi, Rani M.; Thilipan, G. Arun Kumar; Bhat, Vinayachandra; Sridhar, K. R.; Pattabi, Manjunatha

    2013-02-01

    Silver nanoparticles (AgNPs) synthesized by subjecting an aqueous solution of AgNO3 and polyvinyl alcohol to irradiation from an UV lamp has been studied for its antibacterial potential against Gram-negative bacteria (Escherichia coli). The diameter of the zone of inhibition is found to depend on both the irradiation time and the nanoparticle concentration. As the synthesis method adopted uses no toxic reagents, these particles may serve as promising candidates in the search for better antibacterial agents.

  20. Polymorphous crystallization and diffraction of threonine deaminase from Escherichia coli.

    PubMed

    Gallagher, D T; Eisenstein, E; Fisher, K E; Zondlo, J; Chinchilla, D; Yu, H D; Dill, J; Winborne, E; Ducote, K; Xiao, G; Gilliland, G L

    1998-05-01

    The biosynthetic threonine deaminase from Escherichia coli, an allosteric tetramer with key regulatory functions, has been crystallized in several crystal forms. Two distinct forms, both belonging to either space group P3121 or P3221, with different sized asymmetric units that both contain a tetramer, grow under identical conditions. Diffraction data sets to 2.8 A resolution (native) and 2. 9 A resolution (isomorphous uranyl derivative) have been collected from a third crystal form in space group I222.

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

    PubMed Central

    Ireland, J C; Klostermann, P; Rice, E W; Clark, R M

    1993-01-01

    Titanium dioxide in the anatase crystalline form was used as a photocatalyst to generate hydroxyl radicals in a flowthrough water reactor. Experiments were performed on pure cultures of Escherichia coli in dechlorinated tap water and a surface water sample to evaluate the disinfection capabilities of the reactor. In water devoid of significant amounts of inorganic-radical scavengers, rapid cell death was observed with both pure cultures and members of the indigenous flora in a natural water sample. PMID:8390819

  2. Effects of Acridine Orange on the Growth of Escherichia coli

    PubMed Central

    Southwick, Frederick S.; Carr, Howard S.; Carden, George A.; D'Alisa, Rose M.; Rosenkranz, Herbert S.

    1972-01-01

    Exposure of Escherichia coli to critical acridine orange (AO) concentrations did not result in loss of viability. However, the deoxyribonucleic acid (DNA) of cells exposed to such agents was rapidly degraded and repolymerized. On the other hand, a bacterium deficient in DNA repair (pol A1−, lacking DNA polymerase) was sensitive to the action of AO. The DNA of such cells was also degraded but it was not repaired. PMID:4553001

  3. Escherichia coli adherence to HEp-2 cells with prefixed cells.

    PubMed Central

    Zepeda-Lopez, H M; Gonzalez-Lugo, G M

    1995-01-01

    We describe a new method which uses cold absolute methanol-prefixed cells for adherence of enteropathogenic Escherichia coli to HEp-2 cells. We found that a method using bacteria grown in Penassay broth to 10(6) to 10(7) CFU/ml and incubated with prefixed cells for 3 h at 37 degrees C, showed 100% sensitivity and specificity against a method using live cells. PMID:7615770

  4. Electric field induced bacterial flocculation of Enteroaggregative Escherichia coli 042

    SciTech Connect

    Kumar, Aloke; Mortensen, Ninell P; Mukherjee, Partha P; Retterer, Scott T; Doktycz, Mitchel John

    2011-01-01

    A response of the aggregation dynamics of enteroaggregative Escherichia coli under low magnitude steady and oscillating electric fields is presented. The presence of uniform electric fields hampered microbial adhesion and biofilm formation on a transverse glass surface, but instead promoted the formation of flocs. Extremely heterogeneous distribution of live and dead cells was observed among the flocs. Moreover, floc formation was largely observed to be independent of the frequency of alternating electric fields.

  5. Role for the female in bacterial conjugation in Escherichia coli.

    PubMed

    Freifelder, D

    1967-08-01

    Hfr and F' Lac male strains of Escherichia coli were mated with purine-requiring females which had been starved for purine. These females formed mating pairs with the males. However, a mating in the absence of purine markedly reduced the yield of recombinants. Transfer of F' Lac or of lambda prophage also occurred infrequently. It was concluded that deoxyribonucleic acid transfer from male to female requires some, as yet unknown, function of the female.

  6. Role for the Female in Bacterial Conjugation in Escherichia coli

    PubMed Central

    Freifelder, David

    1967-01-01

    Hfr and F′ Lac male strains of Escherichia coli were mated with purine-requiring females which had been starved for purine. These females formed mating pairs with the males. However, a mating in the absence of purine markedly reduced the yield of recombinants. Transfer of F′ Lac or of λ prophage also occurred infrequently. It was concluded that deoxyribonucleic acid transfer from male to female requires some, as yet unknown, function of the female. PMID:5341864

  7. Division pattern of a round mutant of Escherichia coli.

    PubMed Central

    Cooper, S

    1997-01-01

    A round mutant of Escherichia coli, when grown in Methocel medium, forms chains of cells and does not form tetrads. This implies that successive division planes of the round mutant are parallel rather than perpendicular. These results differ from a previous proposal that division planes in this round mutant are perpendicular to the prior division plane (W. D. Donachie, S. Addinall, and K. Begg, Bioessays 17:569-576, 1995). PMID:9287016

  8. Some factors affecting cyclopropane acid formation in Escherichia coli

    PubMed Central

    Knivett, V. A.; Cullen, Julia

    1965-01-01

    1. The fatty acid composition of the extractable lipids of Escherichia coli varied with growth conditions. 2. The principal fatty acids were palmitic acid, hexadecenoic acid, octadecenoic acid and the cyclopropane acids, methylenehexadecanoic acid and methyleneoctadecanoic acid. 3. Cyclopropane acid formation from monoenoic acids was increased by acid media, poor oxygen supply, or high growth temperature. 4. Cyclopropane acid formation was decreased by alkaline media, well oxygenated conditions, the presence of citrate, or lack of Mg2+. PMID:5324304

  9. Evidence of Pathogenic Subgroups among Atypical Enteropathogenic Escherichia coli Strains▿

    PubMed Central

    Scaletsky, Isabel C. A.; Aranda, Katia R. S.; Souza, Tamara B.; Silva, Neusa P.; Morais, Mauro B.

    2009-01-01

    We describe the characterization of 126 atypical enteropathogenic Escherichia coli (aEPEC) isolates from 1,749 Brazilian children. Classic aEPEC strains were more frequently found in children with diarrhea than in controls (P < 0.001), showing their importance as acute diarrhea agents in our country. Only aEPEC strains carrying either the ehxA or paa gene were significantly associated with diarrhea. PMID:19759223

  10. Characterization of Aspergillus oryzae aspartyl aminopeptidase expressed in Escherichia coli.

    PubMed

    Watanabe, Jun; Tanaka, Hisaki; Akagawa, Takumi; Mogi, Yoshinobu; Yamazaki, Tatsuo

    2007-10-01

    To characterize aspartyl aminopeptidase from Aspergillus oryzae, the recombinant enzyme was expressed in Escherichia coli. The enzyme cleaves N-terminal acidic amino acids. About 30% activity was retained in 20% NaCl. Digestion of defatted soybean by the enzyme resulted in an increase in the glutamic acid content, suggesting that the enzyme is potentially responsible for the release of glutamic acid in soy sauce mash.

  11. Two Forms of d-Glycerate Kinase in Escherichia coli

    PubMed Central

    Ornston, M. K.; Ornston, L. N.

    1969-01-01

    Escherichia coli K-12 synthesizes two chromatographically distinct forms of glycerate kinase which differ both in their thermolability and in the dependence of their activity upon pH. One enzymatic form, GK I, is found in cells grown with glycerate, glucarate, or glycolate. Of these compounds, glycolate is the only carbon source that elicits the synthesis of the second enzymatic form, GK II. PMID:4887503

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

    PubMed

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

    2014-07-18

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

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

    PubMed Central

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

    2014-01-01

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

  14. Cajanol inhibits the growth of Escherichia coli and Staphylococcus aureus by acting on membrane and DNA damage.

    PubMed

    Liu, Xiao-lei; Zhang, Xin-jian; Fu, Yu-jie; Zu, Yuan-gang; Wu, Nan; Liang, Lu; Efferth, Thomas

    2011-01-01

    In the present study, the mechanism of antibacterial activity of cajanol extracted from the roots of Cajanus cajan (L.) Millsp. towards Escherichia coli (E. coli) and Staphylococcus aureus (S. aureus) was investigated. The antibacterial activity of cajanol was evaluated towards six bacterial strains (Staphylococcus epidermidis, Staphylococcus aureus, Bacillus subtilis, Escherichia coli, Proteus vulgaris, and Pseudomonas aeruginosa) by the broth microdilution method. It showed strong antibacterial activity towards all bacteria tested with minimal inhibition concentration (MIC) values ranging from 98.90 µM to 197.8 µM. Cajanol-induced death rates in the most sensitive strains ( E.COLI, 96.55 % and S. AUREUS, 97.25 %) were analyzed by flow cytometry. Furthermore, the activity of cajanol on the membranes of E. coli and S. aureus was investigated by using lecithin, phosphate groups, and fluorescence microscopy. Cajanol-induced DNA damage was observed by agarose gel electrophoresis. In summary, cajanol inhibited E. coli only by DNA damage, whereas S. aureus was inhibited by affecting both, the lecithin and phosphate groups on the cellular membrane and DNA. The present study shows that cajanol possesses antibacterial activity in vitro towards both gram-negative and gram-positive bacteria and therefore may be a promising candidate as an antibacterial agent for the therapy of microbial infections.

  15. Thymineless Death in Escherichia coli: Inactivation and Recovery

    PubMed Central

    Cummings, Donald J.; Kusy, Alvin R.

    1969-01-01

    The effects of chloramphenicol (CAP) on the progress of thymineless death (TLD), nalidixic acid (NA) inactivation, ultraviolet (UV) irradiation, and mitomycin C (MC) inactivation were studied in Escherichia coli B, Bs−1, Bs−3, Bs−12, and B/r. This was done before, during, and after inactivation. During the progress of inactivation, it was found that at 10 to 20 μg of CAP per ml, up to 50% of the UV-sensitive bacteria survived TLD and about 10% survived NA. In E. coli B/r, at these concentrations of CAP, about 10 to 15% of the cells survived TLD and about 20 to 25% survived NA. Concentrations of CAP greater than 25 μg/ml actually increased the sensitivity of E. coli B, Bs−1, Bs−3, and Bs−12 to inactivation by either TLD or NA; at 150 μg of CAP per ml, the sensitivity of E. coli B/r to inactivation also increased. When E. coli B cells were incubated in CAP prior to inactivation, the longer the preincubation the longer onset of TLD was delayed; NA inactivation was also affected in that the rate of inactivation after CAP incubation was greatly decreased. Preincubation of E. coli B/r with CAP had much less effect on the progress of inactivation. After thymineless death, incubation in CAP plus thymine led to a rapid and almost complete recovery of E. coli B and Bs−12. Lesser recoveries were observed after inactivation due to UV, NA, or MC inactivation. E. coli Bs−1 and B/r did not recover viability after any mode of inactivation, and E. coli Bs−3 and Bs−12 recovered from UV to about 20% of the initial titer. It was suggested that protein synthesis, in particular proteins involved in deoxyribonucleic synthesis, was a determining factor in these inactivating and recovery events. PMID:4897115

  16. Antibiofilm effect of trans-cinnamaldehyde on uropathogenic Escherichia coli.

    PubMed

    Amalaradjou, Mary Anne Roshni; Narayanan, Amoolya; Baskaran, Sangeetha Ananda; Venkitanarayanan, Kumar

    2010-07-01

    Urinary tract infections are the most common hospital acquired infections in humans, caused primarily by uropathogenic Escherichia coli. Indwelling urinary catheters for bladder drainage in humans become encrusted with uropathogenic E. coli biofilms that are resistant to common antibiotics, resulting in chronic infections. We studied the efficacy of the cinnamon ingredient trans-cinnamaldehyde (Sigma) for preventing uropathogenic E. coli biofilm. We also determined the efficacy of trans-cinnamaldehyde as an ingredient in catheter lock solution to inactivate preformed uropathogenic E. coli biofilm. Polystyrene plates and urinary catheters inoculated with uropathogenic E. coli (5 to 6.0 log cfu) were treated with trans-cinnamaldehyde (0%, 0.1%, 0.25% or 0.5%) at 37C. Catheters with uropathogenic E. coli biofilm were also treated with lock solution containing trans-cinnamaldehyde (0%, 1%, 1.25% or 1.5%). Uropathogenic E. coli biofilm on control and trans-cinnamaldehyde treated plates and catheters was determined on incubation days 0, 1, 3 and 5. Trans-cinnamaldehyde potential cytotoxity, if any, was determined in HTB-4 bladder epithelial cells (ATCC). At all concentrations trans-cinnamaldehyde effectively prevented uropathogenic E. coli biofilm on plates and catheters. As a constituent in catheter lock solution, it inactivated uropathogenic E. coli biofilm on catheters. Trans-cinnamaldehyde produced no cytotoxic effects on human bladder epithelial cells at the tested concentrations. Results suggest that trans-cinnamaldehyde may be applied as a catheter surface coating or as an ingredient in catheter lock solution to prevent urinary tract infection in humans. Copyright (c) 2010 American Urological Association Education and Research, Inc. Published by Elsevier Inc. All rights reserved.

  17. Prevalence of Escherichia coli in apple cider manufactured in Connecticut.

    PubMed

    Dingman, D W

    1999-06-01

    Cider samples obtained from 11 cider mills operating in Connecticut during the 1997 to 1998 production season were tested for the presence of Escherichia coli. Cider production began in mid August and continued through March, with peak production in September and October. Of 314 cider samples tested, 11 (4%) were found to contain E. coli. Of the 11 mills, 6 (55%) tested positive for E. coli in the cider at least once during the production year. E. coli was first observed in cider samples produced in mid to late October and was not detected in samples made after January. A trend was observed for cider to decrease in acidity and increase in Brix (soluble sugars) throughout the production season. No correlation between pH and soluble sugars of cider and the presence of E. coli was detected. Eight mills used both dropped apples and tree-picked apples, whereas three mills used tree-picked apples only. The use of dropped apples in cider production began 5 weeks before the first detection of E. coli in cider. E. coli was isolated from cider samples produced using dropped apples and from samples produced using only tree-picked apples. No direct correlation between the use of dropped apples or tree-picked apples and the presence of E. coli in the cider was observed. An association between the time of apple harvest and the appearance of E. coli in cider was noted. For mills providing adequate records, all contaminated cider was produced from apples harvested between mid October and mid November.

  18. Survival of Escherichia coli on strawberries grown under greenhouse conditions.

    PubMed

    Shaw, Angela Laury; Svoboda, Amanda; Jie, Beatrice; Nonnecke, Gail; Mendonca, Aubrey

    2015-04-01

    Strawberries are soft fruit that are not recommended to have a post-harvest wash due to quality concerns. Escherichia coli O157:H7 has been linked to outbreaks with strawberries but little is known about the survival of E. coli during the growth cycle of strawberries. The survival of E. coli on strawberry plants during growing under greenhouses conditions was evaluated. Soil, leaves, and strawberries (if present) were artificially contaminated with an E. coli surrogate either at the time of planting, first runner removal (4 wk), second runner removal (8 wk), or one week prior to harvest. At harvest E. coli was recovered from the leaves, soil, and strawberries regardless of the contamination time. Time of contamination influenced (P < 0.05) numbers of viable E. coli on the plant. The highest survival of E. coli (P < 0.0001) was detected in soil that was contaminated at planting (4.27 log10 CFU g soil(-1)), whereas, the survival of E. coli was maximal at later contamination times (8 wk and 1 wk prior to harvest) for the leaves (4.40 and 4.68 log10 CFU g leaves(-1)) and strawberries (3.37 and 3.53 log10 CFU strawberry(-1)). Cross contamination from leaves to fruit was observed during this study, with the presence of E. coli on strawberries which had not been present at the time of contamination. These results indicate that good agricultural best practices to avoid contamination are necessary to minimize the risk of contamination of these popular fruit with enteric pathogens. Practices should include soil testing prior to harvest and avoiding contamination of the leaves. Copyright © 2014 Elsevier Ltd. All rights reserved.

  19. Fluorogenic assays for immediate confirmation of Escherichia coli.

    PubMed Central

    Feng, P C; Hartman, P A

    1982-01-01

    Rapid assays for Escherichia coli were developed by using the compound 4-methylumbelliferone glucuronide (MUG), which is hydrolyzed by glucuronidase to yield a fluorogenic product. The production of glucuronidase was limited to strains of E. coli and some Salmonella and Shigella strains in the family Enterobacteriaceae. For immediate confirmation of the presence of E. coli in most-probable-number tubes, MUG was incorporated into lauryl tryptose broth at a final concentration of 100 micrograms/ml. Results of both the presumptive test (gas production) and the confirmed test (fluorescence) for E. coli were obtained from a variety of food, water, and milk samples after incubation for only 24 h at 35 degrees C. Approximately 90% of the tubes showing both gas production and fluorescence contained fecal coliforms (they were positive in EC broth incubated at 45 degrees C). Few false-positive reactions were observed. The lauryl tryptose broth-MUG-most-probable-number assay was superior to violet red bile agar for the detection of heat- and chlorine-injured E. coli cells. Anaerogenic strains produced positive reactions, and small numbers of E. coli could be detected in the presence of large numbers of competing bacteria. The fluorogenic assay was sensitive and rapid; the presence of one viable cell was detected within 20 h. E. coli colonies could be distinguished from other coliforms on membrane filters and plates of violet red bile agar if MUG was incorporated into the culture media. A rapid confirmatory test for E. coli that is amenable to automation was developed by using microtitration plates filled with a nonselective medium containing MUG. Pure or mixed cultures containing E. coli produced fluorescence within 4 h (most strains) to 24 h (a few weakly positive strains). Images PMID:7049088

  20. Steady-State Chemotaxis in Escherichia coli

    NASA Astrophysics Data System (ADS)

    Kafri, Yariv; da Silveira, Rava Azeredo

    2008-06-01

    The bacterium E. coli maneuvers itself to regions with high chemoattractant concentrations by performing two stereotypical moves: “runs,” in which it moves in near-straight lines, and “tumbles,” in which it does not advance but changes direction randomly. The duration of each move is stochastic and depends upon the chemoattractant concentration experienced in the recent past. We relate this stochastic behavior to the steady-state density of a bacterium population, and we derive the latter as a function of chemoattractant concentration. In contrast to earlier treatments, here we account for the effects of temporal correlations and variable tumbling durations. A range of behaviors is obtained that depends subtly upon several aspects of the system—memory, correlation, and tumbling stochasticity, in particular.

  1. Preparation of Soluble Proteins from Escherichia coli

    PubMed Central

    Wingfield, Paul T.

    2014-01-01

    Purification of human IL-1β is used in this unit as an example of the preparation of soluble proteins from E. coli. Bacteria containing IL-1β are lysed, and IL-1 β in the resulting supernatant is purified by anion-exchange chromatography, salt precipitation and cation-exchange chromatography, and then concentrated. Finally, the IL-1 β protein is applied to a gel-filtration column to separate it from remaining higher- and lower-molecular-weight contaminants, the purified protein is stored frozen or is lyophilized. The purification protocol described is typical for a protein that is expressed in fairly high abundance (i.e., >5% total protein) and accumulates in a soluble state. Also, the purification procedure serves as an example of how use classical protein purifications methods which may also be used in conjunction with the affinity-based methods now more commonly used. PMID:25367009

  2. Genetic transformation in Escherichia coli K12.

    PubMed

    Cosloy, S D; Oishi, M

    1973-01-01

    An auxotrophic strain of E. coli K12 treated with CaCl(2) was transformed for several markers at a frequency of up to 10(-6) per recipient cell by a DNA preparation isolated from a prototrophic strain. The transforming activity of the DNA preparation was eliminated by treatment with DNase, heat, or sonication, whereas RNase or Pronase treatment had little effect. Two closely linked genetic markers (leu and ara) showed a high degree of cotransformation linkage when high molecular weight DNA was used, but the linkage was almost completely eliminated when sheared, smaller molecular weight DNA was used. There is genetic evidence that the transformation is a result of the replacement of the preexisting genetic marker on the chromosome by that of the donor DNA.

  3. Natural inactivation of Escherichia coli in anaerobic and reduced groundwater.

    PubMed

    Lisle, J T

    2016-06-01

    Inactivation rates of Escherichia coli in groundwater have most often been determined in aerobic and oxidized systems. This study examined E. coli inactivation rates in anaerobic and extremely reduced groundwater systems that have been identified as recharge zones. Groundwater from six artesian wells was diverted to above-ground, flow-through mesocosms that contained laboratory grown E. coli in diffusion chambers. All groundwater was anaerobic and extremely reduced (ORP < -300 mV). Cells were plated onto mTEC agar during 21-day incubation periods. All data fit a bi-phasic inactivation model, with >95% of the E. coli population being inactivated <11·0 h (mean k = 0·488 ±0·188 h(-1) ). The groundwater geochemical conditions enhanced the inactivation of E. coli to rates approx. 21-fold greater than previously published inactivation rate in groundwater (mean k = 0·023 ± 0·030 h(-1) ). Also, mTEC agar inhibits E. coli growth following exposure to anaerobic and reduced groundwater. Aquifer recharge zones with geochemical characteristics observed in this study complement above-ground engineered processes (e.g. filtration, disinfection), while increasing the overall indicator micro-organism log-reduction rate of a facility. Published 2016. This article is a U.S. Government work and is in the public domain in the USA.

  4. Escherichia coli sequence type 131: epidemiology and challenges in treatment.

    PubMed

    Qureshi, Zubair A; Doi, Yohei

    2014-05-01

    Escherichia coli ST131 has emerged as a global epidemic, multidrug-resistant clone of E. coli causing extra-intestinal infections. It is now highly prevalent among fluoroquinolone-resistant and CTX-M ESBL-producing E. coli isolates worldwide. Humans are likely the primary reservoir of ST131. Factors associated with its acquisition include residence in long-term care facilities and recent receipt of antimicrobial agents. E. coli ST131 causes a wide array of infections ranging from cystitis to life-threatening sepsis. Fluoroquinolones and trimethoprim-sulfamethoxazole are no longer adequate options for empiric therapy when E. coli ST131 is suspected from risk factors and local epidemiology. Expanded-spectrum cephalosporins, piperacillin-tazobactam and carbapenems are options to treat serious non-ESBL-producing E. coli ST131 infections, while carbapenems are indicated for ESBL-producing infections. There is a growing interest in reevaluating oral agents including fosfomycin and pivmecillinam for less serious infections such as uncomplicated cystitis.

  5. Ecological and genetic determinants of plasmid distribution in Escherichia coli.

    PubMed

    Medaney, Frances; Ellis, Richard J; Raymond, Ben

    2016-11-01

    Bacterial plasmids are important carriers of virulence and antibiotic resistance genes. Nevertheless, little is known of the determinants of plasmid distribution in bacterial populations. Here the factors affecting the diversity and distribution of the large plasmids of Escherichia coli were explored in cattle grazing on semi-natural grassland, a set of populations with low frequencies of antibiotic resistance genes. Critically, the population genetic structure of bacterial hosts was chararacterized. This revealed structured E. coli populations with high diversity between sites and individuals but low diversity within cattle hosts. Plasmid profiles, however, varied considerably within the same E. coli genotype. Both ecological and genetic factors affected plasmid distribution: plasmid profiles were affected by site, E. coli diversity, E. coli genotype and the presence of other large plasmids. Notably 3/26 E. coli serotypes accounted for half the observed plasmid-free isolates indicating that within species variation can substantially affect carriage of the major conjugative plasmids. The observed population structure suggest that most of the opportunities for within species plasmid transfer occur between different individuals of the same genotype and support recent experimental work indicating that plasmid-host coevolution, and epistatic interactions on fitness costs are likely to be important in determining occupancy. © 2016 The Authors. Environmental Microbiology published by Society for Applied Microbiology and John Wiley & Sons Ltd.

  6. Measuring Escherichia coli Gene Expression during Human Urinary Tract Infections

    PubMed Central

    Mobley, Harry L. T.

    2016-01-01

    Extraintestinal Escherichia coli (E. coli) evolved by acquisition of pathogenicity islands, phage, plasmids, and DNA segments by horizontal gene transfer. Strains are heterogeneous but virulent uropathogenic isolates more often have specific fimbriae, toxins, and iron receptors than commensal strains. One may ask whether it is the virulence factors alone that are required to establish infection. While these virulence factors clearly contribute strongly to pathogenesis, bacteria must survive by metabolizing nutrients available to them. By constructing mutants in all major metabolic pathways and co-challenging mice transurethrally with each mutant and the wild type strain, we identified which major metabolic pathways are required to infect the urinary tract. We must also ask what else is E. coli doing in vivo? To answer this question, we examined the transcriptome of E. coli CFT073 in the murine model of urinary tract infection (UTI) as well as for E. coli strains collected and analyzed directly from the urine of patients attending either a urology clinic or a university health clinic for symptoms of UTI. Using microarrays and RNA-seq, we measured in vivo gene expression for these uropathogenic E. coli strains, identifying genes upregulated during murine and human UTI. Our findings allow us to propose a new definition of bacterial virulence. PMID:26784237

  7. Interaction of Escherichia coli and Soil Particles in Runoff

    PubMed Central

    Muirhead, Richard William; Collins, Robert Peter; Bremer, Philip James

    2006-01-01

    A laboratory-scale model system was developed to investigate the transport mechanisms involved in the horizontal movement of bacteria in overland flow across saturated soils. A suspension of Escherichia coli and bromide tracer was added to the model system, and the bromide concentration and number of attached and unattached E. coli cells in the overland flow were measured over time. Analysis of the breakthrough curves indicated that the E. coli and bromide were transported together, presumably by the same mechanism. This implied that the E. coli was transported by advection with the flowing water. Overland-flow transport of E. coli could be significantly reduced if the cells were preattached to large soil particles (>45 μm). However, when unattached cells were inoculated into the system, the E. coli appeared to attach predominantly to small particles (<2 μm) and hence remained unattenuated during transport. These results imply that in runoff generated by saturation-excess conditions, bacteria are rapidly transported across the surface and have little opportunity to interact with the soil matrix. PMID:16672484

  8. Biochemical characteristic of biofilm of uropathogenic Escherichia coli Dr(+) strains.

    PubMed

    Zalewska-Piątek, Beata; Wilkanowicz, Sabina; Bruździak, Piotr; Piątek, Rafał; Kur, Józef

    2013-07-19

    Urinary tract infections caused by Escherichia coli are very common health problem in the developed countries. The virulence of the uropathogenic E. coli Dr(+) IH11128 is determined by Dr fimbriae, which are homopolymeric structures composed of DraE subunits with the DraD protein capping the fiber. In this study, we have analyzed the structural and biochemical properties of biofilms developed by E. coli strains expressing Dr fimbriae with or without the DraD tip subunit and the surface-exposed DraD protein. We have also demonstrated that these E. coli strains form biofilms on an abiotic surface in a nutrient-dependent fashion. We present evidence that Dr fimbriae are necessary during the first stage of bacterial interaction with the abiotic surface. In addition, we reveal that the DraD alone is also sufficient for the initial surface attachment at an even higher level than Dr fimbriae and that chloramphenicol is able to reduce the normal attachment of the analyzed E. coli. The action of chloramphenicol also shows that protein synthesis is required for the early events of biofilm formation. Additionally, we have identified reduced exopolysaccharide coverage in E. coli that express only Dr fimbrial polyadhesins at the cell surface with or without the DraD capping subunit. Copyright © 2013 Elsevier GmbH. All rights reserved.

  9. Characterization of Shiga toxigenic Escherichia coli isolated from foods.

    PubMed

    Martínez, Aida Juliana; Bossio, Carolina Paba; Durango, Adriana Coral; Vanegas, Maria Consuelo

    2007-12-01

    The aim of this study was to characterize Shiga toxigenic Escherichia coli (STEC) by PCR using strains isolated from ham, beef, and cattle in Colombia. A total of 189 E. coli strains were tested for the presence of the uidA, stx1, and stx2 genes, and identification was confirmed by the automated PCR BAX system for E. coli O157:H7. Genes encoding Shiga-like toxins (stx) were found in eight (6.06%) of 132 strains previously isolated from minced beef; four (50%) of these strains yielded amplification products for both toxin genes (stx1 and stx2), and four (50%) yielded products only for the stx2 toxin. None of the strains analyzed were positive by PCR for the presence of the single base-pair mutation in the uidA gene from E. coli O157:H7; these results were confirmed by the BAX system analysis. A multiplex PCR assay was standardized for the three genes. Results from this study confirmed previous data about the low prevalence of E. coli O157:H7 and Shiga-like toxins in Colombia and is the first known report of the prevalence of non-O157 enterohemorrhagic E. coli in this country.

  10. Escherichia coli, cattle and the propagation of disease.

    PubMed

    Stein, Richard A; Katz, David E

    2017-03-01

    Several early models describing host-pathogen interaction have assumed that each individual host has approximately the same likelihood of becoming infected or of infecting others. More recently, a concept that has been increasingly emphasized in many studies is that for many infectious diseases, transmission is not homogeneous but highly skewed at the level of populations. In what became known as the '20/80 rule', about 20% of the hosts in a population were found to contribute to about 80% of the transmission potential. These heterogeneities have been described for the interaction between many microorganisms and their human or animal hosts. Several epidemiological studies have reported transmission heterogeneities for Escherichia coli by cattle, a phenomenon with far-reaching agricultural, medical and public health implications. Focusing on E. coli as a case study, this paper will describe super-spreading and super-shedding by cattle, review the main factors that shape these transmission heterogeneities and examine the interface with human health. Escherichia coli super-shedding and super-spreading by cattle are shaped by microorganism-specific, cattle-specific and environmental factors. Understanding the factors that shape heterogeneities in E. coli dispersion by cattle and the implications for human health represent key components that are critical for targeted infection control initiatives. © FEMS 2017. All rights reserved. For permissions, please e-mail: journals.permissions@oup.com.

  11. Gentamicin-Loaded Borate Bioactive Glass Eradicates Osteomyelitis Due to Escherichia coli in a Rabbit Model

    PubMed Central

    Xie, Zongping; Cui, Xu; Zhao, Cunju; Huang, Wenhai; Wang, Jianqiang

    2013-01-01

    The treatment of osteomyelitis induced by Gram-negative bacilli is rarely reported in the literature. This study established a rabbit tibia model of osteomyelitis induced by the Gram-negative bacillus Escherichia coli. Using this model, pellets composed of a chitosan-bonded mixture of borate bioactive glass and gentamicin were evaluated in vitro and in vivo for the treatment of osteomyelitis induced by Escherichia coli. Our results showed that the pellets in phosphate-buffered saline released gentamicin continuously over 26 days. Without the simultaneous use of a systemic antibiotic, the implantation of the gentamicin-loaded pellets into the osteomyelitis region of the tibia resulted in the eradication of 81.82% of infections, as determined by microbiological, histological and radiographic evaluation, and supported the ingrowth of new bone into the tibia defects after 6 weeks of implantation. The results indicate that the gentamicin-loaded borate bioactive glass implant, combining sustained drug release with the ability to support new bone formation, could provide a method for treating osteomyelitis induced by Gram-negative bacilli. PMID:23629702

  12. Gentamicin-loaded borate bioactive glass eradicates osteomyelitis due to Escherichia coli in a rabbit model.

    PubMed

    Xie, Zongping; Cui, Xu; Zhao, Cunju; Huang, Wenhai; Wang, Jianqiang; Zhang, Changqing

    2013-07-01

    The treatment of osteomyelitis induced by Gram-negative bacilli is rarely reported in the literature. This study established a rabbit tibia model of osteomyelitis induced by the Gram-negative bacillus Escherichia coli. Using this model, pellets composed of a chitosan-bonded mixture of borate bioactive glass and gentamicin were evaluated in vitro and in vivo for the treatment of osteomyelitis induced by Escherichia coli. Our results showed that the pellets in phosphate-buffered saline released gentamicin continuously over 26 days. Without the simultaneous use of a systemic antibiotic, the implantation of the gentamicin-loaded pellets into the osteomyelitis region of the tibia resulted in the eradication of 81.82% of infections, as determined by microbiological, histological and radiographic evaluation, and supported the ingrowth of new bone into the tibia defects after 6 weeks of implantation. The results indicate that the gentamicin-loaded borate bioactive glass implant, combining sustained drug release with the ability to support new bone formation, could provide a method for treating osteomyelitis induced by Gram-negative bacilli.

  13. [Escherichia coli, a pathogen under fire from the news].

    PubMed

    Cohen, R; Raymond, J; Gendrel, D; Bingen, E

    2012-11-01

    Escherichia coli is both a gastrointestinal tract commensal and a major pathogen. In recent years, E. coli is under fire from the news due to a better understanding of pathogenic factors, outbreaks of infections caused by enterohaemorrhagic strains, and last but not least, the worrying development of antibiotic resistance. Due to the absence of new compounds active against these strains, producing extended-spectrum ß-lactamases (ESBL) and frequently multiresistant to other antibiotics, their emergence will pose therapeutic problems for practitioners of all pediatric specialties. The gold standard treatment for severe infections due to ESBL-E. coli family is the penem class. The frequent use of penems promotes the emergence of strains resistant to carbapenems. Sparing carbapenems should be a clear objective for non life-threatening infections.

  14. Chemotaxis towards autoinducer 2 mediates autoaggregation in Escherichia coli

    PubMed Central

    Laganenka, Leanid; Colin, Remy; Sourjik, Victor

    2016-01-01

    Bacteria communicate by producing and sensing extracellular signal molecules called autoinducers. Such intercellular signalling, known as quorum sensing, allows bacteria to coordinate and synchronize behavioural responses at high cell densities. Autoinducer 2 (AI-2) is the only known quorum-sensing molecule produced by Escherichia coli but its physiological role remains elusive, although it is known to regulate biofilm formation and virulence in other bacterial species. Here we show that chemotaxis towards self-produced AI-2 can mediate collective behaviour—autoaggregation—of E. coli. Autoaggregation requires motility and is strongly enhanced by chemotaxis to AI-2 at physiological cell densities. These effects are observed regardless whether cell–cell interactions under particular growth conditions are mediated by the major E. coli adhesin (antigen 43) or by curli fibres. Furthermore, AI-2-dependent autoaggregation enhances bacterial stress resistance and promotes biofilm formation. PMID:27687245

  15. Engineering Escherichia coli K12 MG1655 to use starch.

    PubMed

    Rosales-Colunga, Luis Manuel; Martínez-Antonio, Agustino

    2014-05-21

    To attain a sustainable bioeconomy, fuel, or valuable product, production must use biomass as substrate. Starch is one of the most abundant biomass resources and is present as waste or as a food and agroindustry by-product. Unfortunately, Escherichia coli, one of the most widely used microorganisms in biotechnological processes, cannot use starch as a carbon source. We engineered an E. coli strain capable of using starch as a substrate. The genetic design employed the native capability of the bacterium to use maltodextrins as a carbon source plus expression and secretion of its endogenous α-amylase, AmyA, in an adapted background. Biomass production improved using 35% dissolved oxygen and pH 7.2 in a controlled bioreactor. The engineered E. coli strain can use starch from the milieu and open the possibility of optimize the process to use agroindustrial wastes to produce biofuels and other valuable chemicals.

  16. Quantitative method for enumeration of enterotoxigenic Escherichia coli.

    PubMed Central

    Calderon, R L; Levin, M A

    1981-01-01

    A rapid method was developed to quantify toxigenic Escherichia coli, using a membrane filter procedure. After filtration of samples, the membrane filter was first incubated on a medium selective for E. coli (24 h, 44 degrees C) and then transferred to tryptic soy agar (3%; 6 h, 37 degrees C). To assay for labile toxin-producing colonies, the filter was then transferred to a monolayer of Y-1 cells, the E. coli colonies were marked on the bottom of the petri dish, and the filter was removed after 15 min. The monolayer was observed for a positive rounding effect after a 15- to 24-h incubation. The method has an upper limit of detecting 30 toxigenic colonies per plate and can detect as few as one toxigenic colony per plate. A preliminary screening for these enterotoxigenic strains in polluted waters and known positive fecal samples was performed, and positive results were obtained with fecal samples only. PMID:7007415

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

    SciTech Connect

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

    1988-06-01

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

  18. Functions of the gene products of Escherichia coli.

    PubMed Central

    Riley, M

    1993-01-01

    A list of currently identified gene products of Escherichia coli is given, together with a bibliography that provides pointers to the literature on each gene product. A scheme to categorize cellular functions is used to classify the gene products of E. coli so far identified. A count shows that the numbers of genes concerned with small-molecule metabolism are on the same order as the numbers concerned with macromolecule biosynthesis and degradation. One large category is the category of tRNAs and their synthetases. Another is the category of transport elements. The categories of cell structure and cellular processes other than metabolism are smaller. Other subjects discussed are the occurrence in the E. coli genome of redundant pairs and groups of genes of identical or closely similar function, as well as variation in the degree of density of genetic information in different parts of the genome. PMID:7508076

  19. Lipid domains and mechanical plasticity of Escherichia coli lipid monolayers.

    PubMed

    López-Montero, Iván; Arriaga, Laura R; Rivas, Germán; Vélez, Marisela; Monroy, Francisco

    2010-01-01

    Lipid monolayers can be laterally dilated under the action of the barriers in a Langmuir trough thus allowing for measurements of their mechanical response. We study the stress response of Escherichia coli polar lipid extract and POPC against oscillatory deformations stressed up to a 20% of the initial area. For E. coli monolayers a nonlinear regime described by a series of Fourier harmonics of the excitation mode is found beyond a critical strain (u(C) approximately 1%). In contrast, the mechanical response of POPC monolayers is found linear upon much larger deformations. For E. coli monolayers the stress-strain plot reflects stress softening (plastic-like) behaviour whilst POPC behaves as a linear elastic body. No viscous delay with respect to the applied strain is detected in both systems, as expected for high fluid materials. The presence of phase coexistence domains as lipid reservoirs to facilitate lateral diffusion is claimed as a plausible mechanism underlying the observed mechanical plasticity.

  20. Inhibition of Injured Escherichia coli by Several Selective Agents 1

    PubMed Central

    Scheusner, D. L.; Busta, F. F.; Speck, M. L.

    1971-01-01

    A population of Escherichia coli ML30 cells was exposed to a quaternary ammonium compound, and injury to the cells was measured by a comparison of counts on Trypticase Soy Agar and Violet Red Bile Agar. Substantial injury could not be detected with a minimal medium. The ingredients of Violet Red Bile Agar were tested against damaged cells. The bile salts mixture alone in the medium prevented as many injured cells from growing as did any combination of the selective agents and inhibited as many injured bacteria as were inhibited by Violet Red Bile Agar itself. These dyes and salts were similarly assayed in minimal agar, and comparable results were obtained. Individual bile salts and other potential selective agents were added to the minimal medium, and the media were tested for inhibition of injured E. coli. Sodium deoxycholate was the bile salt most inhibitory to damaged E. coli cells. PMID:4924998

  1. Engineering Escherichia coli K12 MG1655 to use starch

    PubMed Central

    2014-01-01

    Background To attain a sustainable bioeconomy, fuel, or valuable product, production must use biomass as substrate. Starch is one of the most abundant biomass resources and is present as waste or as a food and agroindustry by-product. Unfortunately, Escherichia coli, one of the most widely used microorganisms in biotechnological processes, cannot use starch as a carbon source. Results We engineered an E. coli strain capable of using starch as a substrate. The genetic design employed the native capability of the bacterium to use maltodextrins as a carbon source plus expression and secretion of its endogenous α-amylase, AmyA, in an adapted background. Biomass production improved using 35% dissolved oxygen and pH 7.2 in a controlled bioreactor. Conclusion The engineered E. coli strain can use starch from the milieu and open the possibility of optimize the process to use agroindustrial wastes to produce biofuels and other valuable chemicals. PMID:24886307

  2. Incidence of Escherichia coli in Black Walnut Meats

    PubMed Central

    Meyer, Melvin T.; Vaughn, Reese H.

    1969-01-01

    Examination of commercially shelled black walnut meats showed inconsistent numbers of total aerobic bacteria, coliforms, and Escherichia coli; variation occurred among different meat sizes and within each meat size. The incidence of E. coli on meats of commercially hulled black walnuts depended on the physical condition of the nuts. Apparently tightly sealed ones contained only a few or none, whereas those with visibly separated sutures and spoiled meats yielded the most. This contamination was in part correlated to a hulling operation. Large numbers of E. coli on the husk of the walnuts contaminated the hulling water, subsequently also contaminating the meats by way of separated sutures. Chlorination of the hulling wash water was ineffective. Attempts were made to decontaminate the walnut meats without subsequent deleterious changes in flavor or texture. A treatment in coconut oil at 100 C followed by removal of excess surface oil by centrifugation was best. PMID:4905608

  3. Crystal structure of Escherichia coli YidC, a membrane protein chaperone and insertase.

    PubMed

    Kumazaki, Kaoru; Kishimoto, Toshiki; Furukawa, Arata; Mori, Hiroyuki; Tanaka, Yoshiki; Dohmae, Naoshi; Ishitani, Ryuichiro; Tsukazaki, Tomoya; Nureki, Osamu

    2014-12-03

    Bacterial YidC, an evolutionally conserved membrane protein, functions as a membrane protein chaperone in cooperation with the Sec translocon and as an independent insertase for membrane proteins. In Gram-negative bacteria, the transmembrane and periplasmic regions of YidC interact with the Sec proteins, forming a multi-protein complex for Sec-dependent membrane protein integration. Here, we report the crystal structure of full-length Escherichia coli YidC. The structure reveals that a hydrophilic groove, formed by five transmembrane helices, is a conserved structural feature of YidC, as compared to the previous YidC structure from Bacillus halodurans, which lacks a periplasmic domain. Structural mapping of the substrate- or Sec protein-contact sites suggested the importance of the groove for the YidC functions as a chaperone and an insertase, and provided structural insight into the multi-protein complex.

  4. Expression of thermostable β-xylosidase in Escherichia coli for use in saccharification of plant biomass.

    PubMed

    Aftab, Muhammad N; Zafar, Asma; Awan, Ali R

    2017-01-31

    The present work is aimed to evaluate the saccharification potential of a thermostable β-xylosidase cloned from Bacillus licheniformis into Escherichia coli for production of bioethanol from plant biomass. Recombinant β-xylosidase enzyme possesses the ability of bioconversion of plant biomass like wheat straw, rice straw and sugarcane bagass. By using this approach, plant biomass that mainly constitute cellulose can be converted to reducing sugars that could then be easily converted to bioethanol by simple fermentation process. The production of bioethanol will help to overcome energy requirements due to depleting fossil fuels and will also help to protect environment by reducing greenhouse gas emission. In the end, future directions are briefly mentioned that can be utilized to reduce the cost and increase the yield of biofuels.

  5. Nutrient-dependent methylation of a membrane-associated protein of Escherichia coli

    SciTech Connect

    Young, C.C.; Alvarez, J.D.; Bernlohr, R.W. )

    1990-09-01

    Starvation of a mid-log-phase culture of Escherichia coli B/r for nitrogen, phosphate, or carbon resulted in methylation of a membrane-associated protein of about 43,000 daltons (P-43) in the presence of chloramphenicol and (methyl-3H)methionine. The in vivo methylation reaction occurred with a doubling time of 2 to 5 min and was followed by a slower demethylation process. Addition of the missing nutrient to a starving culture immediately prevented further methylation of P-43. P-43 methylation is not related to the methylated chemotaxis proteins because P-43 is methylated in response to a different spectrum of nutrients and because P-43 is methylated on lysine residues. The characteristics of P-43 are similar to those of a methylated protein previously described in Bacillus subtilis and B. licheniformis and are consistent with the proposal that methylation of this protein functions in nutrient sensing.

  6. Lambda Red recombinase-mediated integration of the high molecular weight DNA into the Escherichia coli chromosome.

    PubMed

    Juhas, Mario; Ajioka, James W

    2016-10-05

    Escherichia coli K-12 is a frequently used host for a number of synthetic biology and biotechnology applications and chassis for the development of the minimal cell factories. Novel approaches for integrating high molecular weight DNA into the E. coli chromosome would therefore greatly facilitate engineering efforts in this bacterium. We developed a reliable and flexible lambda Red recombinase-based system, which utilizes overlapping DNA fragments for integration of the high molecular weight DNA into the E. coli chromosome. Our chromosomal integration strategy can be used to integrate high molecular weight DNA of variable length into any non-essential locus in the E. coli chromosome. Using this approach we integrated 15 kb DNA encoding sucrose catabolism and lactose metabolism and transport operons into the fliK locus of the flagellar region 3b in the E. coli K12 MG1655 chromosome. Furthermore, with this system we integrated 50 kb of Bacillus subtilis 168 DNA into two target sites in the E. coli K12 MG1655 chromosome. The chromosomal integrations into the fliK locus occurred with high efficiency, inhibited motility, and did not have a negative effect on the growth of E. coli. In addition to the rational design of synthetic biology devices, our high molecular weight DNA chromosomal integration system will facilitate metabolic and genome-scale engineering of E. coli.

  7. Biosynthesis of isoprene in Escherichia coli via methylerythritol phosphate (MEP) pathway.

    PubMed

    Zhao, Yaru; Yang, Jianming; Qin, Bo; Li, Yonghao; Sun, Yuanzhang; Su, Sizheng; Xian, Mo

    2011-06-01

    Isoprene is an aviation fuel of high quality and an important polymer building block in the synthetic chemistry industry. In light of high oil prices, sustained availability, and environmental concerns, isoprene from renewable materials is contemplated as a substitute for petroleum-based product. Escherichia coli with advantages over other wild microorganisms, is considered as a powerful host for biofuels and chemicals. Here, we constructed a synthetic pathway of isoprene in E. coli by introducing an isoprene synthase (ispS) gene from Populus nigra, which catalyzes the conversion of dimethylallyl diphosphate (DMAPP) to isoprene. To improve the isoprene production, we overexpressed the native 1-deoxy-D: -xylulose-5-phosphate (DXP) synthase gene (dxs) and DXP reductoisomerase gene (dxr) in E. coli, which catalyzed the first step and the second step of MEP pathway, respectively. The fed-batch fermentation results showed that overexpression of DXS is helpful for the improvement of isoprene production. Surprisingly, heterologous expression of dxs and dxr from Bacillus subtilis in the E. coli expressing ispS resulted in a 2.3-fold enhancement of isoprene production (from 94 to 314 mg/L). The promising results showed that dxs and dxr from B. subtilis functioned more efficiently on the enhancement of isoprene production than native ones. This could be caused by the consequence of great difference in protein structures of the two original DXSs. It could be practical to produce isoprene in E. coli via MEP pathway through metabolic engineering. This work provides an alternative way for production of isoprene by engineered E. coli via MEP pathway through metabolic engineering.

  8. Inhibition of Escherichia coli in cultivated cattle manure.

    PubMed

    Weinberg, Z G; Szakacs, G; Chen, Y; Pinto, R; Bernstein, S; Konya, B; Sela Saldinger, S

    2014-05-01

    A common practice on Israeli dairy barns comprises daily cultivation of the manure. Cultivation is a mechanical process used to break up and till the manure bedding and it results in a drier and aerated bedding and cleaner cows, which consequently reduces the incidence of mastitis. Cultivation was associated with a shorter survival of Escherichia coli in cultivated manure as compared with noncultivated manure. The objective of the current study was to elucidate the mechanism responsible for the shorter survival duration of E. coli in the cultivated manure. We hypothesized that microorganisms that are antagonistic to E. coli, developing in the cultivated manure, are responsible for this phenomenon. A cow manure derived E. coli strain expressing the green fluorescence protein and antibiotic resistance markers was used to inoculate cow manure in 1.5-L jars. Manure treatments included cultivated and noncultivated manure. Half the jars of each cultivation treatment were autoclave sterilized at 121°C for 1 h on 3 successive days to eliminate from the manure antagonistic microorganisms. Each cultivation-sterilization treatment was performed in triplicate jars. Following sterilization, E. coli numbers in the cultivated and noncultivated manure were comparable, while in the nonsterilized manure the numbers were lower in the cultivated compared with the noncultivated manure. Several fungi isolated from the cultivated manure samples displayed inhibition effect on the tagged E. coli. Antagonistic fungi were also isolated from large-scale cultivated manure samples collected on several dairy farms in Israel. These findings support the notion that manure cultivation might facilitate the development of microorganisms that are antagonistic to E. coli, thus contributing to the general hygiene of the cattle. Identifying the mechanisms by which the antagonistic fungi affect the survival of E. coli in manure could be exploited for improvement of the animal health and for limiting the

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

    PubMed

    Vahjen, W; Cuisiniere, T; Zentek, J

    2017-10-03

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

  10. Effect of tannins on the in viro growth of Escherichia coli O157:H7 and in vivo growth of generic Escherichia coli excreted from steers

    USDA-ARS?s Scientific Manuscript database

    The effect of commercially available chestnut and mimosa tannins in vitro (experiment 1) or in vivo (experiment 2) on the growth or recovery of Escherichia coli O157:H7 or generic fecal E. coli was evaluated. In experiment 1, the mean growth rate of E. coli O157:H7, determined via the measurement o...

  11. Effective medicinal plants against enterohaemorrhagic Escherichia coli O157:H7.

    PubMed

    Voravuthikunchai, Supayang; Lortheeranuwat, Amornrat; Jeeju, Wanpen; Sririrak, Trechada; Phongpaichit, Souwalak; Supawita, Thanomjit

    2004-09-01

    The stimulating effect of subinhibitory concentrations of antibiotics on the production of verocytotoxin (VT) by enterohaemorrhagic Escherichia coli (EHEC) O157:H7 has been claimed. The purpose of this study was to find an alternative, but bioactive medicine for the treatment of this organism. Fifty-eight preparations of aqueous and ethanolic extracts of 38 medicinal plant species commonly used in Thailand to cure gastrointestinal infections were tested for their antibacterial activity against different strains of Escherichia coli, including 6 strains of Escherichia coli O157:H7, Escherichia coli O26:H11, Escherichia coli O111:NM, Escherichia coli O22; 5 strains of Escherichia coli isolated from bovine; and Escherichia coli ATCC 25922. Inhibition of growth was primarily tested by the paper disc agar diffusion method. Among the medicinal plants tested, only 8 species (21.05%) exhibited antimicrobial activity against Escherichia coli O157:H7. Acacia catechu, Holarrhena antidysenterica, Peltophorum pterocarpum, Psidium guajava, Punica granatum, Quercus infectoria, Uncaria gambir, and Walsura robusta demonstrated antibacterial activity with inhibition zones ranging from 7 to 17 mm. The greatest inhibition zone against Escherichia coli O157:H7 (RIMD 05091083) was produced from the ethanolic extract of Quercus infectoria. Minimal inhibitory concentration (MIC) and minimal bactericidal concentration (MBC) were determined by the agar microdilution method and agar dilution method in petri dishes with millipore filter. Both aqueous and ethanolic extracts of Quercus infectoria and aqueous extract of Punica granatum were highly effective against Escherichia coli O157:H7 with the best MIC and MBC values of 0.09, 0.78, and 0.19, 0.39 mg/ml, respectively. These plant species may provide alternative but bioactive medicines for the treatment of Escherichia coli O157:H7 infection.

  12. Soil solarization reduces Escherichia coli O157:H7 and total Escherichia coli on cattle feedlot pen surfaces.

    PubMed

    Berry, Elaine D; Wells, James E

    2012-01-01

    Feedlot pen soil is a source for transmission of Escherichia coli O157:H7, and therefore a target for preharvest strategies to reduce this pathogen in cattle. The objective of this study was to determine the ability of soil solarization to reduce E. coli O157:H7 in feedlot surface material (FSM). A feedlot pen was identified in which naturally occurring E. coli O157:H7 was prevalent and evenly distributed in the FSM. Forty plots 3 by 3 m were randomly assigned such that five plots of each of the solarization times of 0, 1, 2, 3, 4, 6, 8, and 10 weeks were examined. Temperature loggers were placed 7.5 cm below the surface of each plot, and plots to be solarized were covered with clear 6-mil polyethylene. At each sampling time, five FSM samples were collected from each of five solarized and five unsolarized plots. E. coli concentrations and E. coli O157:H7 presence by immunomagnetic separation and plating were determined for each FSM sample. Initial percentages of E. coli O157:H7-positive samples in control and solarized FSM were 84 and 80%, respectively, and did not differ (P > 0.05). E. coli O157:H7 was no longer detectable by 8 weeks of solarization, but was still detected in unsolarized FSM at 10 weeks. The average initial concentration of E. coli in FSM was 5.56 log CFU/g and did not differ between treatments (P > 0.05). There was a 2.0-log decrease of E. coli after 1 week of solarization, and a >3.0-log reduction of E. coli by week 6 of solarization (P, 0.05). E. coli levels remained unchanged in unsolarized FSM (P > 0.05). Daily peak FSM temperatures were on average 8.7°C higher for solarized FSM compared with unsolarized FSM, and reached temperatures as high as 57°C. Because soil solarization reduces E. coli O157:H7, this technique may be useful for reduction of persistence and transmission of this pathogen in cattle production, in addition to remediation of E. coli O157:H7-contaminated soil used to grow food crops.

  13. 77 FR 26725 - Changes to FSIS Traceback, Recall Procedures for Escherichia coli O157:H7 Positive Raw Beef...

    Federal Register 2010, 2011, 2012, 2013, 2014

    2012-05-07

    ... Food Safety and Inspection Service Changes to FSIS Traceback, Recall Procedures for Escherichia coli... find raw ground beef presumptive positive for Escherichia coli (E. coli) O157:H7. This methodology will... Escherichia coli O157:H7'' and requested comments on these documents. FSIS also held a public meeting...

  14. Synergistic effect of lipopeptide biosurfactant with antibiotics against Escherichia coli CFT073 biofilm.

    PubMed

    Rivardo, Fabrizio; Martinotti, Maria Giovanna; Turner, Raymond Joseph; Ceri, Howard

    2011-04-01

    Biofilms are microcolonies of microbes adherent to biotic and abiotic surfaces, often responsible for chronic infections and medical device contamination. Escherichia coli is one of the prevalent pathogens involved in uropathogenic infections and contamination of catheters. A biosurfactant produced by Bacillus licheniformis V9T14 was tested alone and in association with various antibiotics against a mature 24-h uropathogenic E. coli CFT073 biofilm. Biofilm was grown on polystyrene pegs of a Calgary Biofilm Device, providing a tool to evaluate the efficacy of antimicrobial agents. Antibiotics tested were ampicillin, cefazolin, ceftriaxone, ciprofloxacin, piperacillin, tobramycin and trimethoprim/sulfamethoxazole (19:1). Biosurfactant alone at the concentrations tested was not able to remove the adherent cells of the pre-formed biofilm. However, the difference between the effect of antibiotic alone and in combination with the biosurfactant was significant and exceeded 1log(10) (90%) reduction in most cases. Results of this study indicate that V9T14 biosurfactant in association with antibiotics leads to a synergistic increase in the efficacy of antibiotics in biofilm killing, and in some combinations leads to total eradication of E. coli CFT073 biofilm. Copyright © 2011 Elsevier B.V. and the International Society of Chemotherapy. All rights reserved.

  15. A modified pathway for the production of acetone in Escherichia coli.

    PubMed

    May, Antje; Fischer, Ralf-Jörg; Maria Thum, Simone; Schaffer, Steffen; Verseck, Stefan; Dürre, Peter; Bahl, Hubert

    2013-01-01

    A modified synthetic acetone operon was constructed. It consists of two genes from Clostridium acetobutylicum (thlA coding for thiolase and adc coding for acetoacetate decarboxylase) and one from Bacillus subtilis or Haemophilus influenzae (teII(srf) or ybgC, respectively, for thioesterase). Expression of this operon in Escherichia coli resulted in the production of acetone starting from the common metabolite acetyl-CoA via acetoacetyl-CoA and acetoacetate. The thioesterases do not need a CoA acceptor for acetoacetyl-CoA hydrolysis. Thus, in contrast to the classic acetone pathway of Clostridium acetobutylicum and related microorganisms which employ a CoA transferase, the new pathway is acetate independent. The genetic background of the host strains was crucial. Only E. coli strains HB101 and WL3 were able to produce acetone via the modified plasmid based pathway, up to 64mM and 42mM in 5-ml cultures, respectively. Using glucose fed-batch cultures the concentration could be increased up to 122mM acetone with HB101 carrying the recombinant plasmid pUC19ayt (thioesterase from H. influenzae). The formation of acetone led to a decreased acetate production by E. coli.

  16. Triphenyltin degradation and proteomic response by an engineered Escherichia coli expressing cytochrome P450 enzyme.

    PubMed

    Yi, Wenying; Yang, Kunliang; Ye, Jinshao; Long, Yan; Ke, Jing; Ou, Huase

    2017-03-01

    Although triphenyltin (TPT) degradation pathway has been determined, information about the enzyme and protein networks involved was severely limited. To this end, a cytochrome P450 hydroxylase (CYP450) gene from Bacillus thuringiensis was cloned and expressed in Escherichia coli BL21 (DE3), namely E. coli pET32a-CYP450, whose dosage at 1gL(-1) could degrade 54.6% TPT at 1mgL(-1) within 6 d through attacking the carbon-tin bonds of TPT by CYP450. Sequence analysis verified that the CYP450 gene had a 1214bp open reading frame, encoding a protein with 404 amino acids. Proteomic analysis determined that 60 proteins were significantly differentially regulated expression in E. coli pET32a-CYP450 after TPT degradation. The up-regulated proteins enriched in a network related to transport, cell division, biosynthesis of amino acids and secondary metabolites, and microbial metabolism in diverse environments. The current findings demonstrated for the first time that P450 received electrons transferring from NADH could effectively cleave carbon-metal bonds.

  17. Multidrug-resistant Escherichia coli soft tissue infection investigated with bacterial whole genome sequencing

    PubMed Central

    Buchanan, Ruaridh; Stoesser, Nicole; Crook, Derrick; Bowler, Ian C J W

    2014-01-01

    A 45-year-old man with dilated cardiomyopathy presented with acute leg pain and erythema suggestive of necrotising fasciitis. Initial surgical exploration revealed no necrosis and treatment for a soft tissue infection was started. Blood and tissue cultures unexpectedly grew a Gram-negative bacillus, subsequently identified by an automated broth microdilution phenotyping system as an extended-spectrum β-lactamase producing Escherichia coli. The patient was treated with a 3-week course of antibiotics (ertapenem followed by ciprofloxacin) and debridement for small areas of necrosis, followed by skin grafting. The presence of E. coli triggered investigation of both host and pathogen. The patient was found to have previously undiagnosed liver disease, a risk factor for E. coli soft tissue infection. Whole genome sequencing of isolates from all specimens confirmed they were clonal, of sequence type ST131 and associated with a likely plasmid-associated AmpC (CMY-2), several other resistance genes and a number of virulence factors. PMID:25331151

  18. A Moonlighting Enzyme Links Escherichia coli Cell Size with Central Metabolism

    PubMed Central

    Hill, Norbert S.; Buske, Paul J.; Shi, Yue; Levin, Petra Anne

    2013-01-01

    Growth rate and nutrient availability are the primary determinants of size in single-celled organisms: rapidly growing Escherichia coli cells are more than twice as large as their slow growing counterparts. Here we report the identification of the glucosyltransferase OpgH as a nutrient-dependent regulator of E. coli cell size. During growth under nutrient-rich conditions, OpgH localizes to the nascent septal site, where it antagonizes assembly of the tubulin-like cell division protein FtsZ, delaying division and increasing cell size. Biochemical analysis is consistent with OpgH sequestering FtsZ from growing polymers. OpgH is functionally analogous to UgtP, a Bacillus subtilis glucosyltransferase that inhibits cell division in a growth rate-dependent fashion. In a striking example of convergent evolution, OpgH and UgtP share no homology, have distinct enzymatic activities, and appear to inhibit FtsZ assembly through different mechanisms. Comparative analysis of E. coli and B. subtilis reveals conserved aspects of growth rate regulation and cell size control that are likely to be broadly applicable. These include the conservation of uridine diphosphate glucose as a proxy for nutrient status and the use of moonlighting enzymes to couple growth rate-dependent phenomena to central metabolism. PMID:23935518

  19. Discovery of Fur binding site clusters in Escherichia coli by information theory models

    PubMed Central

    Chen, Zehua; Lewis, Karen A.; Shultzaberger, Ryan K.; Lyakhov, Ilya G.; Zheng, Ming; Doan, Bernard; Storz, Gisela; Schneider, Thomas D.

    2007-01-01

    Fur is a DNA binding protein that represses bacterial iron uptake systems. Eleven footprinted Escherichia coli Fur binding sites were used to create an initial information theory model of Fur binding, which was then refined by adding 13 experimentally confirmed sites. When the refined model was scanned across all available footprinted sequences, sequence walkers, which are visual depictions of predicted binding sites, frequently appeared in clusters that fit the footprints (∼83% coverage). This indicated that the model can accurately predict Fur binding. Within the clusters, individual walkers were separated from their neighbors by exactly 3 or 6 bases, consistent with models in which Fur dimers bind on different faces of the DNA helix. When the E. coli genome was scanned, we found 363 unique clusters, which includes all known Fur-repressed genes that are involved in iron metabolism. In contrast, only a few of the known Fur-activated genes have predicted Fur binding sites at their promoters. These observations suggest that Fur is either a direct repressor or an indirect activator. The Pseudomonas aeruginosa and Bacillus subtilis Fur models are highly similar to the E. coli Fur model, suggesting that the Fur–DNA recognition mechanism may be conserved for even distantly related bacteria. PMID:17921503

  20. A moonlighting enzyme links Escherichia coli cell size with central metabolism.

    PubMed

    Hill, Norbert S; Buske, Paul J; Shi, Yue; Levin, Petra Anne

    2013-01-01

    Growth rate and nutrient availability are the primary determinants of size in single-celled organisms: rapidly growing Escherichia coli cells are more than twice as large as their slow growing counterparts. Here we report the identification of the glucosyltransferase OpgH as a nutrient-dependent regulator of E. coli cell size. During growth under nutrient-rich conditions, OpgH localizes to the nascent septal site, where it antagonizes assembly of the tubulin-like cell division protein FtsZ, delaying division and increasing cell size. Biochemical analysis is consistent with OpgH sequestering FtsZ from growing polymers. OpgH is functionally analogous to UgtP, a Bacillus subtilis glucosyltransferase that inhibits cell division in a growth rate-dependent fashion. In a striking example of convergent evolution, OpgH and UgtP share no homology, have distinct enzymatic activities, and appear to inhibit FtsZ assembly through different mechanisms. Comparative analysis of E. coli and B. subtilis reveals conserved aspects of growth rate regulation and cell size control that are likely to be broadly applicable. These include the conservation of uridine diphosphate glucose as a proxy for nutrient status and the use of moonlighting enzymes to couple growth rate-dependent phenomena to central metabolism.

  1. Salmonella typhimurium intercepts Escherichia coli signaling to enhance antibiotic tolerance

    PubMed Central

    Vega, Nicole M.; Allison, Kyle R.; Samuels, Amanda N.; Klempner, Mark S.; Collins, James J.

    2013-01-01

    Bacterial communication plays an important role in many population-based phenotypes and interspecies interactions, including those in host environments. These interspecies interactions may prove critical to some infectious diseases, and it follows that communication between pathogenic bacteria and commensal bacteria is a subject of growing interest. Recent studies have shown that Escherichia coli uses the signaling molecule indole to increase antibiotic tolerance throughout its population. Here, we show that the intestinal pathogen Salmonella typhimurium increases its antibiotic tolerance in response to indole, even though S. typhimurium does not natively produce indole. Increased antibiotic tolerance can be induced in S. typhimurium by both exogenous indole added to clonal S. typhimurium populations and indole produced by E. coli in mixed-microbial communities. Our data show that indole-induced tolerance in S. typhimurium is mediated primarily by the oxidative stress response and, to a lesser extent, by the phage shock response, which were previously shown to mediate indole-induced tolerance in E. coli. Further, we find that indole signaling by E. coli induces S. typhimurium antibiotic tolerance in a Caenorhabditis elegans model for gastrointestinal infection. These results suggest that the intestinal pathogen S. typhimurium can intercept indole signaling from the commensal bacterium E. coli to enhance its antibiotic tolerance in the host intestine. PMID:23946425

  2. Pulsed-Plasma Disinfection of Water Containing Escherichia coli

    NASA Astrophysics Data System (ADS)

    Satoh, Kohki; MacGregor, Scott J.; Anderson, John G.; Woolsey, Gerry A.; Fouracre, R. Anthony

    2007-03-01

    The disinfection of water containing the microorganism, Escherichia coli (E. coli) by exposure to a pulsed-discharge plasma generated above the water using a multineedle electrode (plasma-exposure treatment), and by sparging the off-gas of the pulsed plasma into the water (off-gas-sparging treatment), is performed in the ambient gases of air, oxygen, and nitrogen. For the off-gas-sparging treatment, bactericidal action is observed only when oxygen is used as the ambient gas, and ozone is found to generate the bactericidal action. For the plasma-exposure treatment, the density of E. coli bacteria decreases exponentially with plasma-exposure time for all the ambient gases. It may be concluded that the main contributors to E. coli inactivation are particle species produced by the pulsed plasma. For the ambient gases of air and nitrogen, the influence of acidification of the water in the system, as a result of pulsed-plasma exposure, may also contribute to the decay of E. coli density.

  3. Recent Advances in Understanding Enteric Pathogenic Escherichia coli

    PubMed Central

    Croxen, Matthew A.; Law, Robyn J.; Scholz, Roland; Keeney, Kristie M.; Wlodarska, Marta

    2013-01-01

    SUMMARY Although Escherichia coli can be an innocuous resident of the gastrointestinal tract, it also has the pathogenic capacity to cause significant diarrheal and extraintestinal diseases. Pathogenic variants of E. coli (pathovars or pathotypes) cause much morbidity and mortality worldwide. Consequently, pathogenic E. coli is widely studied in humans, animals, food, and the environment. While there are many common features that these pathotypes employ to colonize the intestinal mucosa and cause disease, the course, onset, and complications vary significantly. Outbreaks are common in developed and developing countries, and they sometimes have fatal consequences. Many of these pathotypes are a major public health concern as they have low infectious doses and are transmitted through ubiquitous mediums, including food and water. The seriousness of pathogenic E. coli is exemplified by dedicated national and international surveillance programs that monitor and track outbreaks; unfortunately, this surveillance is often lacking in developing countries. While not all pathotypes carry the same public health profile, they all carry an enormous potential to cause disease and continue to present challenges to human health. This comprehensive review highlights recent advances in our understanding of the intestinal pathotypes of E. coli. PMID:24092857

  4. Selective detection of Escherichia coli DNA using fluorescent carbon spindles.

    PubMed

    Roy, Anurag; Chatterjee, Sabyasachi; Pramanik, Srikrishna; Devi, Parukuttyamma Sujatha; Suresh Kumar, Gopinatha

    2016-04-28

    We investigate the interaction of hydrophilic blue emitting carbon spindles with various deoxyribonucleic acids (DNA) having different base pair compositions, such as Herring testes (HT), calf thymus (CT), Escherichia coli (EC) and Micrococcus lysodeikticus (ML) DNA, to understand the mode of interaction. Interestingly, the fluorescent carbon spindles selectively interacted with E. coli DNA resulting in enhanced fluorescence of the former. Interaction of the same carbon with other DNAs exhibited insignificant changes in fluorescence. In addition, in the presence of EC DNA, the D band in the Raman spectrum attributed to the defect state completely disappeared, resulting in enhanced crystallinity. Microscopy images confirmed the wrapping of DNA on the carbon spindles leading to the assembly of spindles in the form of flowers. Dissociation of double-stranded DNA occurred upon interaction with carbon spindles, resulting in selective E. coli DNA interaction. The carbon spindles also exhibited a similar fluorescence enhancement upon treating with E. coli bacteria. These results confirm the possibility of E. coli detection in water and other liquid foods using such fluorescent carbon.

  5. Unusual "flesh-eating" strains of Escherichia coli.

    PubMed

    Shaked, Hila; Samra, Zmira; Paul, Michal; Madar-Shapiro, Liora; Cohen, Jonathan; Pitlik, Silvio; Bishara, Jihad

    2012-12-01

    Monomicrobial necrotizing fasciitis (type II) is typically caused by group A streptococcus alone or in combination with Staphylococcus aureus. Escherichia coli has been isolated from polymicrobial or Fournier's gangrene but has rarely been reported in monomicrobial necrotizing fasciitis. We describe the clinical characteristics and outcomes of seven cases of monomicrobial E. coli necrotizing fasciitis and/or severe soft tissue infection diagnosed at a single institution during an 18-month period. Four isolates from three patients and two isolates from two patients with type I polymicrobial severe soft tissue infection (controls) were assayed by the randomly amplified polymorphic DNA (RAPD) analysis for fingerprinting and PCR amplification of primers in order to detect cytotoxic necrotizing factor 1 and 2 (cnf1 and cnf2) genes. All patients had some type of immune suppression. The limb was the most commonly involved organ. In all cases, E. coli was isolated as a monomicrobial pathogen from blood, fascia, or both. All patients died during hospitalization, three within the first 48 h. The RAPD amplification assay showed a high degree of genetic diversity among the "flesh-eating" strains and controls. The cnf1 toxin gene was identified in two out of three cases, but not in the controls. cnf2 was not detected in any of the patients. E. coli may be responsible for life-threatening necrotizing fasciitis. Further research is needed to reveal relevant risk factors, reservoirs, and modes of transmission of cnf1 E. coli.

  6. Nonthermal atmospheric argon plasma jet effects on Escherichia coli biomacromolecules.

    PubMed

    Hosseinzadeh Colagar, Abasalt; Memariani, Hamed; Sohbatzadeh, Farshad; Valinataj Omran, Azadeh

    2013-12-01

    Nonthermal atmospheric plasma jet, a promising technology based on ionized gas at low temperatures, can be applied for disinfection of contaminated surfaces. In this study, Escherichia coli cells and their macromolecules were exposed to the nonthermal atmospheric argon plasma jet for different time durations. Total protein, genomic DNA, and malondialdehyde (MDA) levels of E. coli were assessed by sodium dodecyl sulfate-polyacrylamide gel electrophoresis and silver staining; agarose gel electrophoresis; and measurement of absorbance at 534 nm, respectively. After exposure, the spectroscopic results of liquid samples indicated that the survival reduction of E. coli can reach to 100 % in an exposure time of 600 s. Moreover, inactivation zones of E. coli, DNA degradation, and MDA levels were significantly increased. Additionally, banding patterns of total protein were changed and amino acid concentrations increased following ninhydrin test. The experimental results suggest that the nonthermal plasma could serve as an effective instrument for both sterilizing E. coli and degrading macromolecules from the surface of the objects being sterilized.

  7. Paper-based ELISA to rapidly detect Escherichia coli.

    PubMed

    Shih, Cheng-Min; Chang, Chia-Ling; Hsu, Min-Yen; Lin, Jyun-Yu; Kuan, Chen-Meng; Wang, Hsi-Kai; Huang, Chun-Te; Chung, Mu-Chi; Huang, Kui-Chou; Hsu, Cheng-En; Wang, Chun-Yuan; Shen, Ying-Cheng; Cheng, Chao-Min

    2015-12-01

    Escherichia coli is a generic indicator of fecal contamination, and certain serotypes cause food- and water-borne illness such as O157:H7. In the clinic, detection of bacteriuria, which is often due to E. coli, is critical before certain surgical procedures or in cases of nosocomial infection to prevent further adverse events such as postoperative infection or sepsis. In low- and middle-income countries, where insufficient equipment and facilities preclude modern methods of detection, a simple, low-cost diagnostic device to detect E. coli in water and in the clinic will have significant impact. We have developed a simple paper-based colorimetric platform to detect E. coli contamination in 5h. On this platform, the mean color intensity for samples with 10(5)cells/mL is 0.118±0.002 (n=4), and 0.0145±0.003 (P<0.01⁎⁎) for uncontaminated samples. This technique is less time-consuming, easier to perform, and less expensive than conventional methods. Thus, paper-based ELISA is an innovative point-of-care diagnostic tool to rapidly detect E. coli, and possibly other pathogens when customized as appropriate, especially in areas that lack advanced clinical equipment.

  8. Fumarate-Mediated Persistence of Escherichia coli against Antibiotics

    PubMed Central

    Kim, Jun-Seob; Cho, Da-Hyeong; Heo, Paul; Jung, Suk-Chae; Park, Myungseo; Oh, Eun-Joong; Sung, Jaeyun; Kim, Pan-Jun; Lee, Suk-Chan; Lee, Dae-Hee; Lee, Sarah; Lee, Choong Hwan; Shin, Dongwoo

    2016-01-01

    Bacterial persisters are a small fraction of quiescent cells that survive in the presence of lethal concentrations of antibiotics. They can regrow to give rise to a new population that has the same vulnerability to the antibiotics as did the parental population. Although formation of bacterial persisters in the presence of various antibiotics has been documented, the molecular mechanisms by which these persisters tolerate the antibiotics are still controversial. We found that amplification of the fumarate reductase operon (FRD) in Escherichia coli led to a higher frequency of persister formation. The persister frequency of E. coli was increased when the cells contained elevated levels of intracellular fumarate. Genetic perturbations of the electron transport chain (ETC), a metabolite supplementation assay, and even the toxin-antitoxin-related hipA7 mutation indicated that surplus fumarate markedly elevated the E. coli persister frequency. An E. coli strain lacking succinate dehydrogenase (SDH), thereby showing a lower intracellular fumarate concentration, was killed ∼1,000-fold more effectively than the wild-type strain in the stationary phase. It appears that SDH and FRD represent a paired system that gives rise to and maintains E. coli persisters by producing and utilizing fumarate, respectively. PMID:26810657

  9. The Escherichia coli Proteome: Past, Present, and Future Prospects†

    PubMed Central

    Han, Mee-Jung; Lee, Sang Yup

    2006-01-01

    Proteomics has emerged as an indispensable methodology for large-scale protein analysis in functional genomics. The Escherichia coli proteome has been extensively studied and is well defined in terms of biochemical, biological, and biotechnological data. Even before the entire E. coli proteome was fully elucidated, the largest available data set had been integrated to decipher regulatory circuits and metabolic pathways, providing valuable insights into global cellular physiology and the development of metabolic and cellular engineering strategies. With the recent advent of advanced proteomic technologies, the E. coli proteome has been used for the validation of new technologies and methodologies such as sample prefractionation, protein enrichment, two-dimensional gel electrophoresis, protein detection, mass spectrometry (MS), combinatorial assays with n-dimensional chromatographies and MS, and image analysis software. These important technologies will not only provide a great amount of additional information on the E. coli proteome but also synergistically contribute to other proteomic studies. Here, we review the past development and current status of E. coli proteome research in terms of its biological, biotechnological, and methodological significance and suggest future prospects. PMID:16760308

  10. The Genetic Basis of Escherichia coli Pathoadaptation to Macrophages

    PubMed Central

    Miskinyte, Migla; Sousa, Ana; Ramiro, Ricardo S.; de Sousa, Jorge A. Moura; Kotlinowski, Jerzy; Caramalho, Iris; Magalhães, Sara; Soares, Miguel P.; Gordo, Isabel

    2013-01-01

    Antagonistic interactions are likely important driving forces of the evolutionary process underlying bacterial genome complexity and diversity. We hypothesized that the ability of evolved bacteria to escape specific components of host innate immunity, such as phagocytosis and killing by macrophages (MΦ), is a critical trait relevant in the acquisition of bacterial virulence. Here, we used a combination of experimental evolution, phenotypic characterization, genome sequencing and mathematical modeling to address how fast, and through how many adaptive steps, a commensal Escherichia coli (E. coli) acquire this virulence trait. We show that when maintained in vitro under the selective pressure of host MΦ commensal E. coli can evolve, in less than 500 generations, virulent clones that escape phagocytosis and MΦ killing in vitro, while increasing their pathogenicity in vivo, as assessed in mice. This pathoadaptive process is driven by a mechanism involving the insertion of a single transposable element into the promoter region of the E. coli yrfF gene. Moreover, transposition of the IS186 element into the promoter of Lon gene, encoding an ATP-dependent serine protease, is likely to accelerate this pathoadaptive process. Competition between clones carrying distinct beneficial mutations dominates the dynamics of the pathoadaptive process, as suggested from a mathematical model, which reproduces the observed experimental dynamics of E. coli evolution towards virulence. In conclusion, we reveal a molecular mechanism explaining how a specific component of host innate immunity can modulate microbial evolution towards pathogenicity. PMID:24348252

  11. Antibiotic Resistance in Urinary Isolates of Escherichia coli

    PubMed Central

    Abduzaimovic, Amila; Aljicevic, Mufida; Rebic, Velma; Vranic, Sabina Mahmutovic; Abduzaimovic, Kadrija; Sestic, Sabina

    2016-01-01

    Objectives: The aim of this study was to examine the presence of antimicrobial resistance / susceptibility strains of Escherichia coli in inpatients and outpatients. Materials and methods: It is a retrospective study carried out at the Department of Microbiology, Parasitology and Virology Faculty of Medicine, University of Sarajevo. In cooperation with the Microbiological laboratory of the Cantonal Hospital Zenica and the Microbiological laboratory of the General Hospital Tesanj, 3863 urine samples were processed in the period from March 1st to March 31st 2016. Results: Our study showed that E. coli had the highest antimicrobial resistance to trimethoprim / sulfamethoxazole (38.61%), followed by amoxicillin / clavulanic acid (19.62%), ciprofloxacin (9.49%), gentamicin (8.86%), cephalexin (8.23%), nitrofurantoin (8.23%), cefuroxime (7.52%), ceftazidime (6.33%), cefuroxime (89.87%), amikacin (4.43%). Conclusions: The isolated strains of E. coli showed the highest resistance to trimethoprim / sulfamethoxazole and amoxicillin / clavulanic acid. The isolated strains of E. coli showed the greatest susceptibility to amikacin and ceftazidime. Gender distribution of positive E. coli isolates showed statistically significant differences in favor of females. PMID:28144190

  12. Antibiotic Resistance of Escherichia coli Serotypes from Cochin Estuary

    PubMed Central

    Sukumaran, Divya P.; Durairaj, Srinivasan; Abdulla, Mohamed Hatha

    2012-01-01

    This study aimed at detecting the prevalence of antibiotic-resistant serotypes of Escherichia coli in Cochin estuary, India. E. coli strains were isolated during the period January 2010–December 2011 from five different stations set at Cochin estuary. Water samples from five different stations in Cochin estuary were collected on a monthly basis for a period of two years. Isolates were serotyped, antibiogram-phenotyped for twelve antimicrobial agents, and genotyped by polymerase chain reaction for uid gene that codes for β-D-glucuronidase. These E. coli strains from Cochin estuary were tested against twelve antibiotics to determine the prevalence of multiple antibiotic resistance among them. The results revealed that more than 53.33% of the isolates were multiple antibiotic resistant. Thirteen isolates showed resistance to sulphonamides and two of them contained the sul 1 gene. Class 1 integrons were detected in two E. coli strains which were resistant to more than seven antibiotics. In the present study, O serotyping, antibiotic sensitivity, and polymerase chain reaction were employed with the purpose of establishing the present distribution of multiple antibiotic-resistant serotypes, associated with E. coli isolated from different parts of Cochin estuary. PMID:23008708

  13. Escherichia coli exports cyclic AMP via TolC.

    PubMed

    Hantke, Klaus; Winkler, Karin; Schultz, Joachim E

    2011-03-01

    In Escherichia coli more than 180 genes are regulated by the cyclic AMP (cAMP)-cAMP receptor protein (CRP) complex. However, more than 90% of cAMP that is made by intracellular adenylyl cyclases is found in the culture medium. How is cAMP exported from E. coli? In a tolC mutant, 0.03 mM IPTG (isopropyl-β-d-thiogalactopyranoside) was sufficient to induce β-galactosidase compared to 0.1 mM IPTG in the parent strain. In a cya mutant unable to produce cAMP about 1 mM extracellular cAMP was required to induce β-galactosidase, whereas in a cya tolC mutant 0.1 mM cAMP was sufficient. When cAMP in E. coli cya was generated intracellularly by a recombinant, weakly active adenylyl cyclase from Corynebacterium glutamicum, the critical level of cAMP necessary for induction of maltose degradation was only achieved in a tolC mutant and not in the parent strain. Deletion of a putative cAMP phosphodiesterase of E. coli, CpdA, resulted in a slightly similar, yet more diffuse phenotype. The data demonstrate that export of cAMP via TolC is a most efficient way of E. coli to lower high concentrations of cAMP in the cell and maintain its sensitivity in changing metabolic environments.

  14. Effects of Escherichia coli hemolysin on endothelial cell function.

    PubMed Central

    Suttorp, N; Flöer, B; Schnittler, H; Seeger, W; Bhakdi, S

    1990-01-01

    Escherichia coli hemolysin is considered an important virulence factor in extraintestinal E. coli infections. The present study demonstrates that cultured pulmonary artery endothelial cells are susceptible to attack by low concentrations of E. coli hemolysin (greater than or equal to 0.05 hemolytic units/ml; greater than or equal to 5 ng/ml). Sublytic amounts of hemolysin increased the permeability of endothelial cell monolayers in a time- and dose-dependent manner. The hydraulic conductivity increased approximately 30-fold and the reflection coefficient for large molecules dropped from 0.71 to less than 0.05, indicating a toxin-induced loss of endothelial barrier function. The alterations of endothelial monolayer permeability were accompanied by cell retraction and interendothelial gap formation. In addition, E. coli hemolysin stimulated prostacyclin synthesis in endothelial cells. This effect was strictly dependent on the presence of extracellular Ca2+ but not of Mg2+. An enhanced passive influx of 45Ca2+ and 3H-sucrose but not of tritiated inulin and dextran was noted in toxin-treated cells, indicating that small transmembrane pores comparable to those detected in rabbit erythrocytes had been generated in endothelial cell membranes. These pores may act as nonphysiologic Ca2+ gates, thereby initiating different Ca2+-dependent cellular processes. We conclude that endothelial cells are highly susceptible to E. coli hemolysin and that two major endothelial cell functions are altered by very low concentrations of hemolysin. Images PMID:2121650

  15. Escherichia coli β-Lactamases: What Really Matters

    PubMed Central

    Bajaj, Priyanka; Singh, Nambram S.; Virdi, Jugsharan S.

    2016-01-01

    Escherichia coli strains belonging to diverse pathotypes have increasingly been recognized as a major public health concern. The β-lactam antibiotics have been used successfully to treat infections caused by pathogenic E. coli. However, currently, the utility of β-lactams is being challenged severely by a large number of hydrolytic enzymes – the β-lactamases expressed by bacteria. The menace is further compounded by the highly flexible genome of E. coli, and propensity of resistance dissemination through horizontal gene transfer and clonal spread. Successful management of infections caused by such resistant strains requires an understanding of the diversity of β-lactamases, their unambiguous detection, and molecular mechanisms underlying their expression and spread with regard to the most relevant information about individual bacterial species. Thus, this review comprises first such effort in this direction for E. coli, a bacterial species known to be associated with production of diverse classes of β-lactamases. The review also highlights the role of commensal E. coli as a potential but under-estimated reservoir of β-lactamases-encoding genes. PMID:27065978

  16. Escherichia coli ST131, an Intriguing Clonal Group

    PubMed Central

    Bertrand, Xavier; Madec, Jean-Yves

    2014-01-01

    SUMMARY In 2008, a previously unknown Escherichia coli clonal group, sequence type 131 (ST131), was identified on three continents. Today, ST131 is the predominant E. coli lineage among extraintestinal pathogenic E. coli (ExPEC) isolates worldwide. Retrospective studies have suggested that it may originally have risen to prominence as early as 2003. Unlike other classical group B2 ExPEC isolates, ST131 isolates are commonly reported to produce extended-spectrum β-lactamases, such as CTX-M-15, and almost all are resistant to fluoroquinolones. Moreover, ST131 E. coli isolates are considered to be truly pathogenic, due to the spectrum of infections they cause in both community and hospital settings and the large number of virulence-associated genes they contain. ST131 isolates therefore seem to contradict the widely held view that high levels of antimicrobial resistance are necessarily associated with a fitness cost leading to a decrease in pathogenesis. Six years after the first description of E. coli ST131, this review outlines the principal traits of ST131 clonal group isolates, based on the growing body of published data, and highlights what is currently known and what we need to find out to provide public health authorities with better information to help combat ST131. PMID:24982321

  17. EFFECT OF LYSERGIC ACID DIETHYLAMIDE ON Escherichia coli, STRAIN B/r(lambda).

    DTIC Science & Technology

    LYSERGIC ACIDS , *ESCHERICHIA COLI), GROWTH(PHYSIOLOGY), CHROMOSOMES, DAMAGE, DOSAGE, PURINE ALKALOIDS, ULTRAVIOLET RADIATION, DEOXYRIBONUCLEIC ACIDS , INHIBITION, HALLUCINOGENS, CHEMICAL WARFARE AGENTS, BIOASSAY

  18. Diarrhea, bacteremia and multiorgan dysfunction due to an extraintestinal pathogenic Escherichia coli strain with enteropathogenic E. coli genes.

    PubMed

    Kessler, Robert; Nisa, Shahista; Hazen, Tracy H; Horneman, Amy; Amoroso, Anthony; Rasko, David A; Donnenberg, Michael S

    2015-11-01

    A 55-year-old man with well-controlled HIV had severe diarrhea for 3 weeks and developed multiorgan dysfunction and bacteremia due to Escherichia coli. The genome of the patient's isolate had features characteristic of extraintestinal pathogenic E. coli and genes distantly related to those defining enteropathogenic E. coli.

  19. Role of enteroaggregative Escherichia coli virulence factors in uropathogenesis.

    PubMed

    Boll, Erik J; Struve, Carsten; Boisen, Nadia; Olesen, Bente; Stahlhut, Steen G; Krogfelt, Karen A

    2013-04-01

    A multiresistant clonal Escherichia coli O78:H10 strain qualifying molecularly as enteroaggregative Escherichia coli (EAEC) was recently shown to be the cause of a community-acquired outbreak of urinary tract infection (UTI) in greater Copenhagen, Denmark, in 1991. This marks the first time EAEC has been associated with an extraintestinal disease outbreak. Importantly, the outbreak isolates were recovered from the urine of patients with symptomatic UTI, strongly implying urovirulence. Here, we sought to determine the uropathogenic properties of the Copenhagen outbreak strain and whether these properties are conferred by the EAEC-specific virulence factors. We demonstrated that through expression of aggregative adherence fimbriae, the principal adhesins of EAEC, the outbreak strain exhibited pronouncedly increased adherence to human bladder epithelial cells compared to prototype uropathogenic strains. Moreover, the strain was able to produce distinct biofilms on abiotic surfaces, including urethral catheters. These findings suggest that EAEC-specific virulence factors increase uropathogenicity and may have played a significant role in the ability of the strain to cause a community-acquired outbreak of UTI. Thus, inclusion of EAEC-specific virulence factors is warranted in future detection and characterization of uropathogenic E. coli.

  20. Metabolic engineering of Escherichia coli for the production of cinnamaldehyde.

    PubMed

    Bang, Hyun Bae; Lee, Yoon Hyeok; Kim, Sun Chang; Sung, Chang Keun; Jeong, Ki Jun

    2016-01-19

    Plant parasitic nematodes are harmful to agricultural crops and plants, and may cause severe yield losses. Cinnamaldehyde, a volatile, yellow liquid commonly used as a flavoring or food additive, is increasingly becoming a popular natural nematicide because of its high nematicidal activity and, there is a high demand for the development of a biological platform to produce cinnamaldehyde. We engineered Escherichia coli as an eco-friendly biological platform for the production of cinnamaldehyde. In E. coli, cinnamaldehyde can be synthesized from intracellular L-phenylalanine, which requires the activities of three enzymes: phenylalanine-ammonia lyase (PAL), 4-coumarate:CoA ligase (4CL), and cinnamoyl-CoA reductase (CCR). For the efficient production of cinnamaldehyde in E. coli, we first examined the activities of enzymes from different sources and a gene expression system for the selected enzymes was constructed. Next, the metabolic pathway for L-phenylalanine biosynthesis was engineered to increase the intracellular pool of L-phenylalanine, which is a main precursor of cinnamaldehyde. Finally, we tried to produce cinnamaldehyde with the engineered E. coli. According to this result, cinnamaldehyde production as high as 75 mg/L could be achieved, which was about 35-fold higher compared with that in the parental E. coli W3110 harboring a plasmid for cinnamaldehyde biosynthesis. We also confirmed that cinnamaldehyde produced by our engineered E. coli had a nematicidal activity similar to the activity of commercial cinnamaldehyde by nematicidal assays against Bursaphelenchus xylophilus. As a potential natural pesticide, cinnamaldehyde was successfully produced in E. coli by construction of the biosynthesis pathway and, its production titer was also significantly increased by engineering the metabolic pathway of L-phenylalanine.