‘Supermutators’ found amongst highly levofloxacin-resistant <i>E. coli</i> isolates: a rapid protocol for the detection of mutation sites

Zayed, Ahmed A.; Essam, Tamer; Hashem, Abdelgawad; El–Tayeb, O. M.

doi:10.1038/emi.2015.4

Cited by 19 publications

(20 citation statements)

References 32 publications

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“…Another genetic alteration commonly seen was a frame shift at codon 29 of AcrR regulator protein (V29fs). In the same codon, V29G was reported by others [55]. Additionally, we detected other SNPs that might be significant, but have not been reported previously in the literature (S3 Table).…”

Section: Mechanisms Of Carbapenem Resistancesupporting

confidence: 84%

“…For the rest of the strains, we reported high frequency of missense mutations in porins (OmpC and OmpF), while only one was seen in regulatory OmpR protein in one strain (OM150) (S3 Table). Regarding efflux pump, we reported already known missense mutations in AcrR (T5N), MarR (G103S, Y137H, K62R, S3N), SoxR (T38S, G74R), and SoxS (A12S) to induce overexpression of the AcrAB-TolC efflux pump and contribute to fluoroquinolones and tigecycline resistance in E. coli [50][51][52][53][54][55]. Also, we saw a A111T missense mutation in SoxR, which has unclear clinical significance, as it has been seen previously in isolates with normal expression of soxS mRNA [52,55].…”

Section: Mechanisms Of Carbapenem Resistancementioning

confidence: 99%

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Clinical and molecular characteristics of carbapenem non-susceptible Escherichia coli: A nationwide survey from Oman

et al. 2020

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The prevalence of carbapenem-resistant Enterobacterales (CRE) in the Arabian Peninsula is predicted to be high, as suggested from published case reports. Of particular concern, is carbapenem-resistant E. coli (CR-EC), due to the importance of this species as a community pathogen. Herein, we conducted a comprehensive molecular characterization of putative CR-EC strains from Oman. We aim to establish a baseline for future molecular monitoring. We performed whole-genome sequencing (WGS) for 35 putative CR-EC. Isolates were obtained from patients at multiple centers in 2015. Genetic relatedness was investigated using several typing approaches such as MLST, SNP calling, phylogroup and CRISPR typing. Maxiuium likelihood SNP-tree was performed by RAxML after variant calling and removal of recombination regions with Snippy and Gubbins, respectively. Resistance genes, plasmid replicon types, virulence genes, and prophage were also characterised. The online databases CGE, CRISPRcasFinder, Phaster and EnteroBase were used for the in silico analyses. Screening for mutations in genes regulating the expression of porins and efflux pump as well as mutations lead to fluoroquinolones resistance were performed with CLC Genomics Workbench. The genetic diversity suggests a polyclonal population structure with 21 sequence types (ST), of which ST38 being the most prevalent (11%). SNPs analysis revealed possible transmission episodes. Whereas, CRISPR typing helped to spot outlier strains belonged to phylogroups other than B2 which was CRISPRfree. The virulent phylogroups B2 and D were detected in 4 and 9 isolates, respectively. In some strains bacteriophages acted as vectors for virulence genes. Regarding resistance to β-lactam, 22 were carbapenemase producers, 3 carbapenem non-susceptible but carbapenemase-negative, 9 resistant to expanded-spectrum cephalosporins, and one isolate with susceptibility to cephalosporins and carbapenems. Thirteen out of the 22 (59%) carbapenemase-producing isolates were NDM and 7 (23%) were OXA-48-like which mirrors the situation in Indian subcontinent. Two isolates co-produced NDM and OXA-48-like enzymes. In total, 80% (28/35) were CTX-M-15 producers and 23% (8/35) featured AmpC. The high-risk subclones ST131-H30Rx/C2, ST410-H24RxC and ST1193-H64RxC were detected, the

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Section: Mechanisms Of Carbapenem Resistancesupporting

confidence: 84%

Section: Mechanisms Of Carbapenem Resistancementioning

confidence: 99%

Clinical and molecular characteristics of carbapenem non-susceptible Escherichia coli: A nationwide survey from Oman

et al. 2020

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“…Norfloxacin MICs for most of the double mutants were greater than 1024mg/L and ciprofloxacin MICs 256mg/L. Double mutations in gyrase are thought to reduce the affinity of the gyrase-DNA complex for quinolones [ 21 ]. Sequencing of QRDR region by Namboodiri et al [ 22 ] observed that isolates with an additional gyrA substitution Asp87→Asn were resistant to both ciprofloxacin as well as nalidixic acid.…”

Section: Resultsmentioning

confidence: 99%

Analytical profiling of mutations in quinolone resistance determining region of gyrA gene among UPEC

et al. 2018

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Mutations in gyrA are the primary cause of quinolone resistance encountered in gram-negative clinical isolates. The prospect of this work was to analyze the role of gyrA mutations in eliciting high quinolone resistance in uropathogenic E.coli (UPEC) through molecular docking studies. Quinolone susceptibility testing of 18 E.coli strains isolated from UTI patients revealed unusually high resistance level to all the quinolones used; especially norfloxacin and ciprofloxacin. The QRDR of gyrA was amplified and sequenced. Mutations identified in gyrA of E.coli included Ser83Leu, Asp87Asn and Ala93Gly/Glu. Contrasting previous reports, we found Ser83Leu substitution in sensitive strains. Strains with S83L, D87N and A93E (A15 and A26) demonstrated norfloxacin MICs ≥1024mg/L which could be proof that Asp87Asn is necessary for resistance phenotype. Resistance to levofloxacin was comparatively lower in all the isolates. Docking of 4 quinolones (ciprofloxacin, ofloxacin, levofloxacin and norfloxacin) to normal and mutated E.coli gyrase A protein demonstrated lower binding energies for the latter, with significant displacement of norfloxacin in the mutated GyrA complex and least displacement in case of levofloxacin.

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“…Many clinical isolates of E . coli cells highly resistant to fluoroquinolones have been isolated and the mutations that support such a high-level resistance have been identified [ 14 , 23 – 26 ]. It is reasonable to imagine that in clinical settings there are a large variety of pathways to high-level resistance, not necessarily similar to what we have devised here.…”

Section: Resultsmentioning

confidence: 99%

A Transporter Interactome Is Essential for the Acquisition of Antimicrobial Resistance to Antibiotics

Shuster¹,

Steiner‐Mordoch²,

Cudkowicz³

et al. 2016

PLoS ONE

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Awareness of the problem of antimicrobial resistance (AMR) has escalated and drug-resistant infections are named among the most urgent problems facing clinicians today. Our experiments here identify a transporter interactome and portray its essential function in acquisition of antimicrobial resistance. By exposing E. coli cells to consecutive increasing concentrations of the fluoroquinolone norfloxacin we generated in the laboratory highly resistant strains that carry multiple mutations, most of them identical to those identified in clinical isolates. With this experimental paradigm, we show that the MDTs function in a coordinated mode to provide an essential first-line defense mechanism, preventing the drug reaching lethal concentrations, until a number of stable efficient alterations occur that allow survival. Single-component efflux transporters remove the toxic compounds from the cytoplasm to the periplasmic space where TolC-dependent transporters expel them from the cell. We postulate a close interaction between the two types of transporters to prevent rapid leak of the hydrophobic substrates back into the cell. The findings change the prevalent concept that in Gram-negative bacteria a single multidrug transporter, AcrAB-TolC type, is responsible for the resistance. The concept of a functional interactome, the process of identification of its members, the elucidation of the nature of the interactions and its role in cell physiology will change the existing paradigms in the field. We anticipate that our work will have an impact on the present strategy searching for inhibitors of AcrAB-TolC as adjuvants of existing antibiotics and provide novel targets for this urgent undertaking.

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‘Supermutators’ found amongst highly levofloxacin-resistant E. coli isolates: a rapid protocol for the detection of mutation sites

Cited by 19 publications

References 32 publications

Clinical and molecular characteristics of carbapenem non-susceptible Escherichia coli: A nationwide survey from Oman

Clinical and molecular characteristics of carbapenem non-susceptible Escherichia coli: A nationwide survey from Oman

Analytical profiling of mutations in quinolone resistance determining region of gyrA gene among UPEC

A Transporter Interactome Is Essential for the Acquisition of Antimicrobial Resistance to Antibiotics

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