High resolution characterisation of E. coli proliferation profiles in livestock faeces

Bacterial cytokinesis is mediated by the Z-ring, which is formed by the prokaryotic tubulin homolog FtsZ. Recent data indicate that the Z-ring is composed of small patches of FtsZ protofilaments that travel around the bacterial cell by treadmilling. Treadmilling involves a switch from a relaxed (R) state, favored for monomers, to a tense (T) conformation, which is favored upon association into filaments. The R conformation has been observed in numerous monomeric FtsZ crystal structures and the T conformation in Staphylococcus aureus FtsZ crystallized as assembled filaments. However, while Escherichia coli has served as a main model system for the study of the Z-ring and the associated divisome, a structure has not yet been reported for E. coli FtsZ. To address this gap, structures were determined of the E. coli FtsZ mutant FtsZ(L178E) with GDP and GTP bound to 1.35 and 1.40 Å resolution, respectively. The E. coli FtsZ(L178E) structures both crystallized as straight filaments with subunits in the R conformation. These high-resolution structures can be employed to facilitate experimental cell-division studies and their interpretation in E. coli.

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Predictive Fluorescent Amplified-Fragment Length Polymorphism Analysis of Escherichia coli: High-Resolution Typing Method with Phylogenetic Significance

Journal of Clinical Microbiology ◽

10.1128/jcm.37.5.1274-1279.1999 ◽

1999 ◽

Vol 37 (5) ◽

pp. 1274-1279 ◽

Cited By ~ 42

Author(s):

Catherine Arnold ◽

Lou Metherell ◽

Geraldine Willshaw ◽

Anthony Maggs ◽

John Stanley

Keyword(s):

Escherichia Coli ◽

High Resolution ◽

Length Polymorphism ◽

Fragment Length ◽

Fragment Length Polymorphism ◽

Amplified Fragment Length Polymorphism ◽

Polymorphism Analysis ◽

Enzyme Electrophoresis ◽

Typing Method ◽

E Coli

The fluorescent amplified-fragment length polymorphism (FAFLP) assay potentially amplifies a unique set of genome fragments from each bacterial clone. It uses stringently hybridizing primers which carry a fluorescent label. Precise fragment sizing is achieved by the inclusion of an internal size standard in every lane. Therefore, a unique genotype identifier(s) can be found in the form of fragments of precise size or sizes, and these can be generated reproducibly. In order to evaluate the potential of FAFLP as an epidemiological typing method with a valid phylogenetic basis, we applied it to 87 strains ofEscherichia coli. These comprised the EcoR collection, which has previously been classified by multilocus enzyme electrophoresis (MLEE) and which represents the genetic diversity of the species E. coli, plus 15 strains of the clinically important serogroup O157. FAFLP with an unlabelled nonselectiveEcoRI primer (Eco+0) and a labelled selectiveMseI primer (Mse+TA) gave strain-specific profiles. Fragments of identical sizes (in base pairs) were assumed to be identical, and the genetic distances between the strains were calculated. A phylogenetic tree derived from measure of distance correlated closely with the MLEE groupings of the EcoR collection and placed the verocytotoxin-producing O157 strains on an outlier branch. Our data indicate that FAFLP is suitable for epidemiological investigation of E. coli infection, providing well-defined and reproducible identifiers of genotype for each strain. Since FAFLP objectively samples the whole genome, each strain or isolate can be assigned a place within the broad context of the whole species and can also be subjected to a high-resolution comparison with closely related strains to investigate epidemiological clonality.

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Interfacial nanomechanical heterogeneity of the E. coli biofilm matrix

Nanoscale ◽

10.1039/d0nr03646c ◽

2020 ◽

Vol 12 (32) ◽

pp. 16819-16830

Author(s):

Christian Titus Kreis ◽

Ruby May A. Sullan

Keyword(s):

High Resolution ◽

Quantitative Imaging ◽

E Coli ◽

High Resolution Structure ◽

Biofilm Matrix ◽

Resolution Structure

Quantitative imaging correlates high-resolution structure and nanomechanics of the biofilm interface.

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High Resolution Melt Assays to Detect and Identify Vibrio parahaemolyticus, Bacillus cereus, Escherichia coli, and Clostridioides difficile Bacteria

Microorganisms ◽

10.3390/microorganisms8040561 ◽

2020 ◽

Vol 8 (4) ◽

pp. 561

Author(s):

Allison C. Bender ◽

Jessica A. Faulkner ◽

Katherine Tulimieri ◽

Thomas H. Boise ◽

Kelly M. Elkins

Keyword(s):

Escherichia Coli ◽

High Resolution ◽

Bacillus Cereus ◽

Vibrio Parahaemolyticus ◽

Bacterial Species ◽

Human Pathogens ◽

High Resolution Melt ◽

E Coli ◽

Clostridioides Difficile ◽

Sensitivity Specificity

Over one hundred bacterial species have been determined to comprise the human microbiota in a healthy individual. Bacteria including Escherichia coli, Bacillus cereus, Clostridioides difficile, and Vibrio parahaemolyticus are found inside of the human body and B. cereus and E. coli are also found on the skin. These bacteria can act as human pathogens upon ingestion of contaminated food or water, if they enter an open wound, or antibiotics, and environment or stress can alter the microbiome. In this study, we present new polymerase chain reaction (PCR) high-resolution melt (HRM) assays to detect and identify the above microorganisms. Amplified DNA from C. difficile, E. coli, B. cereus, and V. parahaemolyticus melted at 80.37 ± 0.45 °C, 82.15 ± 0.37 °C, 84.43 ± 0.50 °C, and 86.74 ± 0.65 °C, respectively. A triplex PCR assay was developed to simultaneously detect and identify E. coli, B. cereus, and V. parahaemolyticus, and cultured microorganisms were successfully amplified, detected, and identified. The assays demonstrated sensitivity, specificity, reproducibility, and robustness in testing.

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Insights into the improved macrolide inhibitory activity from the high-resolution cryo-EM structure of dirithromycin bound to the E. coli 70S ribosome

RNA ◽

10.1261/rna.073817.119 ◽

2020 ◽

Vol 26 (6) ◽

pp. 715-723 ◽

Cited By ~ 5

Author(s):

Evgeny B. Pichkur ◽

Alena Paleskava ◽

Andrey G. Tereshchenkov ◽

Pavel Kasatsky ◽

Ekaterina S. Komarova ◽

...

Keyword(s):

High Resolution ◽

Inhibitory Activity ◽

E Coli ◽

70S Ribosome

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Transcription-Factor-Mediated DNA Looping Probed by High-Resolution, Single-Molecule Imaging in Live E. coli Cells

PLoS Biology ◽

10.1371/journal.pbio.1001591 ◽

2013 ◽

Vol 11 (6) ◽

pp. e1001591 ◽

Cited By ~ 44

Author(s):

Zach Hensel ◽

Xiaoli Weng ◽

Arvin Cesar Lagda ◽

Jie Xiao

Keyword(s):

Transcription Factor ◽

High Resolution ◽

Single Molecule ◽

Dna Looping ◽

Single Molecule Imaging ◽

E Coli ◽

Resolution Single

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High-resolution cryo-EM structures of the E. coli hemolysin ClyA oligomers

10.1101/558338 ◽

2019 ◽

Author(s):

Wei Peng ◽

Marcela de Souza Santos ◽

Yang Li ◽

Diana R. Tomchick ◽

Kim Orth

Keyword(s):

High Resolution ◽

Structural Model ◽

Complex Structure ◽

Transition Process ◽

Target Cells ◽

E Coli ◽

Interaction Mode ◽

Major Branch ◽

Pore Complexes ◽

Pore Forming Proteins

SummaryPore-forming proteins (PFPs) represent a functionally important protein family, that are found in organisms from viruses to humans. As a major branch of PFPs, bacteria pore-forming toxins (PFTs) permeabilize membranes and usually cause the death of target cells. E. coli hemolysin ClyA is the first member with the pore complex structure solved among α-PFTs, employing α-helices as transmembrane elements. ClyA is proposed to form pores composed of various numbers of protomers. With high-resolution cryo-EM structures, we observe that ClyA pore complexes can exist as newly confirmed oligomers of a tridecamer and a tetradecamer, at estimated resolutions of 3.2 Å and 4.3 Å, respectively. The 2.8 Å cryo-EM structure of a dodecamer dramatically improves the existing structural model. Structural analysis indicates that protomers from distinct oligomers resemble each other and neighboring protomers adopt a conserved interaction mode. We also show a stabilized intermediate state of ClyA during the transition process from soluble monomers to pore complexes. Unexpectedly, even without the formation of mature pore complexes, ClyA can permeabilize membranes and allow leakage of particles less than ∼400 Daltons. In addition, we are the first to show that ClyA forms pore complexes in the presence of cholesterol within artificial liposomes. These findings provide new mechanistic insights into the dynamic process of pore assembly for the prototypical α-PFT ClyA.

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