scholarly journals Development of a Whole-Cell Assay for Peptidoglycan Biosynthesis Inhibitors

2002 ◽  
Vol 46 (4) ◽  
pp. 943-946 ◽  
Author(s):  
Maria D. F. S. Barbosa ◽  
Gaoyun Yang ◽  
Jie Fang ◽  
Michael G. Kurilla ◽  
David L. Pompliano
Keyword(s):  
Active Site ◽  
Antimicrobial Agents ◽  
Inhibitory Concentration ◽  
Direct Detection ◽  
Enzyme Inactivation ◽  
Penicillin G ◽  
Dependent Manner ◽  
Freezing And Thawing ◽  
E Coli ◽  
Peptidoglycan Biosynthesis

ABSTRACT Osmotically stabilized Escherichia coli cells subjected to freezing and thawing were utilized as the source of enzymes for a peptidoglycan pathway assay that can be used to simultaneously test all targets of the committed steps of cell wall biosynthesis. The use of 14C-labeled UDP-N-acetylglucosamine (UDP-GlcNAc) as a substrate allows the direct detection of cross-linked peptidoglycan formed. The assay was validated with known antibiotics. Fosfomycin was the strongest inhibitor of the pathway assay, with a 50% inhibitory concentration of 1 μM. Flavomycin, bacitracin, vancomycin, d-cycloserine, penicillin G, and ampicillin also inhibited formation of radiolabeled peptidoglycan by the E. coli cells. Screening of compounds identified two inhibitors of the pathway, Cpd1 and Cpd2. Subsequent tests with a biochemical assay utilizing purified enzyme implicated UDP-GlcNAc enolpyruvyl transferase (MurA) as the target of Cpd1. This compound inhibits the first enzyme of the pathway in a time-dependent manner. Moreover, enzyme inactivation is dependent on preincubation in the presence of UDP-GlcNAc, which forms a complex with MurA, exposing its active site. Cpd1 also displayed antimicrobial activity against a panel of microorganisms. The pathway assay used in conjunction with assays for individual enzymes provides an efficient means of detecting and characterizing novel antimicrobial agents.

Effectiveness of ZnO Nano Particles against the Foodborne Microbial Pathogens E. coli and S. aureus

2020 ◽  
Vol 15 (2) ◽  
pp. 87-94
Keyword(s):  
Antimicrobial Agents ◽  
Inhibitory Concentration ◽  
Antimicrobial Effect ◽  
Nano Particles ◽  
Microbial Pathogens ◽  
Gram Positive ◽  
Gram Negative ◽  
E Coli ◽  
Antimicrobial Susceptibility Tests ◽  
Susceptibility Tests

In this work, various concentrations of ZnO nano particles, prepared by the coprecipitation method with a size range of 47-68 nm, have been investigated as antimicrobial agents. Dilution antimicrobial susceptibility tests were carried out on two kinds of microbes (Gram-positive Staphylococcus aureus and Gram-negative Escherichia coli) according to the standard method recommended by Clinical and Laboratory Standards Institute, CLSI-2015-M07-A10. The results showed that the antimicrobial effect is larger, the higher the concentration of ZnO nano particles in solution. It was also found that Gram-positive microbes are more sensitive to ZnO nano particles when compared with the Gram-negative ones. The minimum inhibitory concentration (MIC) for E. coli was found to be 50 mg/mL while that for S. aureus was 25 mg/mL. The minimum bactericidal concentration (MBC) was 1600 mg/mL for E. coli and 800 mg/mL for S. aureus.

Synthesis and antimicrobial activity of aryldiazenyl/arylhydrazono pyrazoles

2021 ◽  
Vol 45 (11-12) ◽  
pp. 1093-1099
Author(s):  
Abdulrhman Alsayari ◽  
Yahya I Asiri ◽  
Abdullatif Bin Muhsinah ◽  
Mohd. Zaheen Hassan
Keyword(s):  
Active Site ◽  
Phenyl Ring ◽  
Antimicrobial Agents ◽  
Inhibitory Concentration ◽  
Docking Studies ◽  
Considerable Effect ◽  
Design Synthesis ◽  
Structure Activity ◽  
Antimicrobial Evaluation

We report the design, synthesis, and in vitro antimicrobial evaluation of functionalized pyrazoles containing a hydrazono/diazenyl moiety. Among these newly synthesized derivatives, 4-[2-(4-chlorophenyl)hydrazono]-5-methyl-2-[2-(naphthalen-2-yloxy)acetyl]-2,4-dihydro-3 H-pyrazol-3-one is a promising antimicrobial agent against Staphylococcus aureus (minimum inhibitory concentration 0.19 μg mL−1). Structure–activity relationship studies reveal that the electronic environment on the distal phenyl ring has a considerable effect on the antimicrobial potential of the hybrid analogues. Molecular docking studies into the active site of S. aureus dihydrofolate reductase also prove the usefulness of hybridizing a pyrazole moiety with azo and hydrazo groups in the design of new antimicrobial agents.

scholarly journals Adaptation of Esherichia coli to Antiobiotic Cycling via Single Nucleotide Polymorphisms

2017 ◽  
Vol 14 (1) ◽  
Author(s):  
Samuel Hager ◽  
Ellen Jensen ◽  
Timothy Johnson ◽  
David Mitchell
Keyword(s):  
Escherichia Coli ◽  
Minimum Inhibitory Concentration ◽  
Nalidixic Acid ◽  
Inhibitory Concentration ◽  
Acid Resistance ◽  
Penicillin G ◽  
Single Nucleotide ◽  
E Coli ◽  
Esherichia Coli ◽  
Nalidixic Acid Resistance

Bacteria are quick to adapt and evolve, especially under the effects of selective pressures from chemical antibiotics. In addition, bacteria may develop resistance to antibiotics from multiple classes simultaneously, making their eradication from the human body particularly challenging. This study aims to demonstrate that bacterial multiple-drug resistance can be developed and retained in a laboratory setting. Escherichia coli B was grown in tryptic soy broth in the presence of a small, increasing concentration of streptomycin. This exposure resulted in a strain of E. coli, which had an increased minimum inhibitory concentration (MIC) towards streptomycin, or “resistance.” This resistant strain was then grown in like manner in nalidixic acid and then penicillin G. The result was a strain that became resistant to streptomycin and nalidixic acid, and increasingly resistant to nalidixic acid after penicillin G exposure. Additionally, the bacteria retained resistance to streptomycin and nalidixic acid even after exposure to those chemicals ceased. Genome sequencing and comparison to E. coli B reference strain REL606 revealed the emergence of point mutations with each exposure to an antibiotic. Of particular interest is a mutation associated with the appearance of nalidixic acid resistance. Base pair 4,553,488 was changed from adenine to guanine, resulting in a change from aspartate to glycine in the protein helicase. Previous studies have not indicated mutations to this locus as nalidixic acid resistance conferring. Thus, this mutation may be a novel mutation conferring E. coli B nalidixic acid resistance. Since the region of the mutated helicase is functionally undefined, a mechanism is not apparent. Further research needs to be done to confirm this hypothesis and illuminate a mechanism. KEYWORDS: Bacteria; Escherichia coli; Evolution; Antibiotic Resistance; Nalidixic Acid; Streptomycin; Point Mutation; Single-nucleotide Polymorphism; Helicase; Minimum Inhibitory Concentration

scholarly journals Antibacterial effect of silver nanoparticles on antibiotic resistant E. coli O157:H7 isolated from some dairy products

2020 ◽  
Vol 23 (4) ◽  
pp. 432-442
Author(s):  
W. Elsherif ◽  
D. Ali
Keyword(s):  
Food Safety ◽  
Dairy Products ◽  
Food Packaging ◽  
Antimicrobial Agents ◽  
Bactericidal Effect ◽  
Health Concern ◽  
Penicillin G ◽  
Economic Losses ◽  
Ag Nps ◽  
E Coli

Food safety is a worldwide health goal so foodborne diseases are a main health concern. A total 150 of dairy products samples (locally made yoghurt, ice cream and Talaga cheese) (50 for each type) were examined for E.coli O157:H7 detection and PCR confirmation using fliCH7 gene. E. coli O157:H7 was detected at 18%, 4%, 8% respectively, in samples. The isolates showed broad antibiotic resistance against vancomycin (84.6%), penicillin G (76.9%), cloxacillin (69.2%) and tetracycline (61.5%). Because of increasing number of microorganisms that are resistant to multiple antibiotics causing continuing economic losses in dairy manufacturing, there is an urgent need for development of alternative, cost-effective, and efficient antimicrobial agents to overcome antimicrobial resistance. Here, silver nanoparticle (AgNPs) solution was prepared, identified by transmission electron microscopy (TEM) with an average size 26.5 nm and examined for bactericidal activity against E. coli O157:H7 by using well diffusion assay. The mean inhibition zones of 25 and 50 µg/ml concentrations of Ag-NPs were 15.0±1.2 and 20.9±1.4 mm, respectively. In addition, the statistical analysis showed highly significant differences in the bactericidal effect of different Ag-NPs concentrations on E. coli O157:H7 strains. Bacterial sensitivity to nanoparticles is a key factor in manufacture, so nanoparticles were considered suitable for long life application in food packaging and food safety.

Proteome Analysis of Virulence Factor Regulated by Autoinducer-2–like Activity in Escherichia coli O157:H7

2007 ◽  
Vol 70 (2) ◽  
pp. 300-307 ◽  
Author(s):  
YOUNGHOON KIM ◽  
SANGNAM OH ◽  
EUN YOUNG AHN ◽  
JEE-YOUNG IMM ◽  
SEJONG OH ◽  
...  
Keyword(s):  
Escherichia Coli ◽  
Pathogenic Bacteria ◽  
Antimicrobial Agents ◽  
Proteome Analysis ◽  
Protein Translation ◽  
Escherichia Coli O157 ◽  
Dependent Manner ◽  
Two Dimensional Gel Electrophoresis ◽  
E Coli ◽  
Autoinducer 2

Many pathogenic bacteria, including Escherichia coli O157:H7, can control gene expression in a cell density–dependent manner by producing small signaling molecules (autoinducers) in a process known as quorum sensing. In this study, the effects of the autoinducer-2–like activity on the expression of proteins, including virulence factors, in E. coli O157:H7 were characterized by proteomic analysis. Compared with the control, E. coli O157:H7 strains in the presence of autoinducer-2–like activity exhibited elevated virulence by more rapidly forming cell aggregates on epithelial cells and rapidly killing the nematode Caenorhabditis elegans, the surrogate host. Two-dimensional gel electrophoresis revealed 18 proteins that were upregulated by autoinducer-2–like activity and 4 proteins that were down-regulated. These proteins were further characterized by matrix-assisted laser desorption-ionization time-of-flight mass spectrometry and are involved in the metabolic process, adaptation and protection, cell motility, secretion, envelope biogenesis, and protein translation. These results indicate that the newly identified proteins are associated with the control of virulence in E. coli O157:H7 and that these proteins can be potential targets for the development of antibiotics and other antimicrobial agents.

Phytochemical study: molecular docking of eugenol derivatives as antioxidant and antimicrobial agents

2022 ◽  
Vol 19 ◽  
Author(s):  
Achraf Abdou ◽  
Sabrine Idouaarame ◽  
Mohammed Salah ◽  
Nabil Nor ◽  
Soukaina Zahm, ◽  
...  
Keyword(s):  
Molecular Docking ◽  
Active Site ◽  
Natural Compound ◽  
Antimicrobial Agents ◽  
Aromatic Plants ◽  
Phytochemical Study ◽  
E Coli ◽  
Biological Interest ◽  
Antioxidant Agent ◽  
And Staphylococcus Aureus

Abstract: Eugenol (4-allyl-2-methoxyphenol) is a natural phenolic compound present in certain aromatic plants; however, it is generally extracted from essential oil of Eugenia caryophyllata (Syzygiumaromaticum) (L.) Merr. and L.M. Perry. This bioactive natural compound has generated considerable biological interest with well-known antimicrobial and antioxidant actions. The authors have aimed to the evaluations of eugenol derivatives and their as antimicrobial and antioxidant agent with the aid of molecular dynamic simulation. The starting material was extracted from cloves using hydrodistillation. Two eugenol derivatives, acetyleugenol (4-allyl-2-methoxyphenylacetate) and epoxyeugenol (4-allyl-2-methoxyphenol) were prepared and tested against two strains Escherichia coli (E. Coli) and Staphylococcus aureus (S. Aureus). The results have revealed that the three compounds (Eugenol, acetyleugenol and epoxyeugenol) possess important potentials of inhibition against E. coli and S. Aureus. The antioxidant activity of eugenol derivatives was evaluated by the reaction with DPPH (1,1-diphenyl-2-picrylhydrazyl), showed that the epoxyeugenol was the most active compound. The molecular docking scores of three compounds and the amino acids in the active site pockets of the selected proteins of the two bacteria have approved and explain the biological experimental outcomes.

Distinguishing On-Target versus Off-Target Activity in Early Antibacterial Drug Discovery Using a Macromolecular Synthesis Assay

2013 ◽  
Vol 18 (9) ◽  
pp. 1018-1026 ◽  
Author(s):  
Mark L. Cunningham ◽  
Bryan P. Kwan ◽  
Kirk J. Nelson ◽  
Daniel C. Bensen ◽  
Karen J. Shaw
Keyword(s):  
Antimicrobial Agents ◽  
Inhibitory Concentration ◽  
Antibacterial Drug ◽  
Macromolecular Synthesis ◽  
Topoisomerase Iv ◽  
Structure Based Drug Design ◽  
E Coli ◽  
Drug Classes ◽  
Pathway Inhibition ◽  
Target Activity

The macromolecular synthesis assay was optimized in both S. aureus and E. coli imp and used to define patterns of inhibition of DNA, RNA, protein, and cell wall biosynthesis of several drug classes. The concentration of drug required to elicit pathway inhibition differed among the antimicrobial agents tested, with inhibition detected at concentrations significantly below the minimum inhibitory concentration (MIC) for tedizolid; within 4-fold of the MIC for ciprofloxacin, cefepime, vancomycin, tetracycline, and chloramphenicol; and significantly above the MIC for rifampicin and kanamycin. In a DNA gyrase/topoisomerase IV structure-based drug design optimization program, the assay rapidly identified undesirable off-target activity within certain chemotypes, altering the course of the program to focus on the series that maintained on-target activity.

Improved Agar Diffusion Method for Detecting Residual Antimicrobial Agents

2001 ◽  
Vol 64 (3) ◽  
pp. 361-366 ◽  
Author(s):  
CHIN-EN TSAI ◽  
FUSAO KONDO
Keyword(s):  
Antimicrobial Agents ◽  
Micrococcus Luteus ◽  
Oxolinic Acid ◽  
Photobacterium Phosphoreum ◽  
Penicillin G ◽  
Diffusion Method ◽  
E Coli ◽  
Assay Medium ◽  
Agar Diffusion ◽  
Agar Diffusion Method

The improved agar diffusion method for determination of residual antimicrobial agents was investigated, and the sensitivities of various combinations of test organisms and assay media were determined using 7 organisms, 5 media, and 31 antimicrobial agents. Bacillus stearothermophilus and synthetic assay medium (SAM) showed the greatest sensitivity for screening penicillins (penicillin G and ampicillin). The combination of Bacillus subtilis and minimum medium (MM) was the most sensitive for tetracyclines (oxytetracycline and chlortetracycline), B. stearothermophilus and SAM or Micrococcus luteus and Mueller-Hinton agar (MHA) for detecting tylosin and erythromycin, B. subtilis and MHA for aminoglycosides (streptomycin, kanamycin, gentamicin, and dihydrostreptomycin), B. stearothermophilus and SAM for polyethers (salinomycin and lasalocid), and B. subtilis and MM or Clostridium perfringens and GAM for polypeptides (thiopeptin, enramycin, virginiamycin, and bacitracin). However, gram-negative bacterium Escherichia coli ATCC 27166 and MM were better for screening for colistin and polymixin-B. For detecting the synthetic drugs tested, the best combination was B. subtilis and MM for sulfonamides, E. coli 27166 and MM for quinolones (oxolinic acid and nalidixic acid), B. subtilis and MM for furans (furazolidone), and the bioluminescent bacterium Photobacterium phosphoreum and luminescence assay medium for chloramphenicol and oxolinic acid. The results showed that the use of four assay plates, B. stearothermophilus and SAM, B. subtilis and MM, M. luteus and MHA, and E. coli 27166 and MM, was superior to the currently available techniques for screening for residual antimicrobial agents in edible animal tissues.

scholarly journals Novel Strategy for Biofilm Inhibition by Using Small Molecules Targeting Molecular Chaperone DnaK

2014 ◽  
Vol 59 (1) ◽  
pp. 633-641 ◽  
Author(s):  
Ken-ichi Arita-Morioka ◽  
Kunitoshi Yamanaka ◽  
Yoshimitsu Mizunoe ◽  
Teru Ogura ◽  
Shinya Sugimoto
Keyword(s):  
Molecular Chaperone ◽  
Biofilm Formation ◽  
Antimicrobial Agents ◽  
Dependent Manner ◽  
Cellular Functions ◽  
Biofilm Inhibition ◽  
Content Type ◽  
E Coli ◽  
Chaperone Dnak ◽  
Biofilm Development

ABSTRACTBiofilms are complex communities of microorganisms that attach to surfaces and are embedded in a self-produced extracellular matrix. Since these cells acquire increased tolerance against antimicrobial agents and host immune systems, biofilm-associated infectious diseases tend to become chronic. We show here that the molecular chaperone DnaK is important for biofilm formation and that chemical inhibition of DnaK cellular functions is effective in preventing biofilm development. Genetic, microbial, and microscopic analyses revealed that deletion of thednaKgene markedly reduced the production of the extracellular functional amyloid curli, which contributes to the robustness ofEscherichia colibiofilms. We tested the ability of DnaK inhibitors myricetin (Myr), telmisartan, pancuronium bromide, and zafirlukast to prevent biofilm formation ofE. coli. Only Myr, a flavonol widely distributed in plants, inhibited biofilm formation in a concentration-dependent manner (50% inhibitory concentration [IC50] = 46.2 μM); however, it did not affect growth. Transmission electron microscopy demonstrated that Myr inhibited the production of curli. Phenotypic analyses of thermosensitivity, cell division, intracellular level of RNA polymerase sigma factor RpoH, and vulnerability to vancomycin revealed that Myr altered the phenotype ofE. coliwild-type cells to make them resemble those of the isogenicdnaKdeletion mutant, indicating that Myr inhibits cellular functions of DnaK. These findings provide insights into the significance of DnaK in curli-dependent biofilm formation and indicate that DnaK is an ideal target for antibiofilm drugs.

scholarly journals Application of LC-MS/MS with ion mobility for chemical analysis of propolis extracts with antimicrobial potential

2021 ◽  
pp. 86-86
Author(s):  
Ivana Sofrenic ◽  
Jovana Ljujic ◽  
Katarina Simic ◽  
Stefan Ivanovic ◽  
Jovana Stankovic-Jeremic ◽  
...  
Keyword(s):  
Quorum Sensing ◽  
Ion Mobility ◽  
Antimicrobial Agents ◽  
Inhibitory Concentration ◽  
Film Formation ◽  
Complex Mixture ◽  
Bacterial Strains ◽  
E Coli ◽  
Ethanol Extracts ◽  
The Given

The objective of this study was to test four-dimensional LC-ESI-MS/MS chromatography in analysis of complex mixture such as ethanol extracts of different propolis samples. In total more than 1200 picks were identified and only for 185 literature conformation was found. The given data represent the result of tentative identification, and summarized results are given in the text. Comparing the samples, from different altitudes, 96 components were detected as characteristic in high altitude samples and 18 in samples collected at low altitudes. Antimicrobial activity of ethanol extracts of propolis (EEP) and propylene glycol extracts of propolis (PGEP) were carried out on S. aureus, B. cereus, M. flavus, L. monocytogenes, P. aeruginosa, S. typhimurium, E. coli and E. cloacae bacterial strains and compared with broad-spectrum antibiotics, streptomycin and ampicillin. Anti-quorum sensing activity was performed on P. aeruginosa by testing the effect of representative propolis extracts on bio-film formation, twitching and motility activity and production of pyocyanin. We demonstrated that the majority of explored propolis extracts have greater or equal minimal inhibitory concentration and minimum bactericidal concentration values compared to antibiotics, independently of the solvent used for the extraction. The samples collected from the highest altitude emerged as least active antimicrobial agents but with the greatest potential as anti-quorum sensing agents.

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