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

Lesley R Varughese,Ravinder Mehra,Vikas Beniwal,Soniya Goyal,Meenakshi Rajpoot,Vinod Chhokar,Muna Anjum

doi:10.1371/journal.pone.0190729

Lesley R Varughese, Ravinder Mehra + Show 5 more

Open Access

https://doi.org/10.1371/journal.pone.0190729

Copy DOI

Abstract

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.

Highlights

Quinolones comprise broad-spectrum antibiotics successfully used over the years for the treatment of many infections
Ciprofloxacin, levofloxacin, norfloxacin and ofloxacin (Cipla Ltd, Mumbai) were the quinolones used in this study
Antimicrobial susceptibility testing was performed on Muller Hinton Agar by agar dilution method to determine the minimum inhibitory concentration (MIC) at varying concentrations of quinolone antibiotics as per guidelines [14]

Summary

Introduction

Quinolones comprise broad-spectrum antibiotics successfully used over the years for the treatment of many infections. The greater potency and versatility of these drugs paved the way for effective therapy of many diseases like urinary tract infections, osteomyelitis, pneumonia and gastrointestinal diseases. Their extensive use and misuse has led to the emergence and spread of resistance among bacteria [1, 2]. DNA gyrase and topoisomerase IV (type II topoisomerases) are the prime targets of quinolones. DNA gyrase, a tetramer of two A and two B subunits, encoded by the gyrA and gyrB genes respectively, is responsible for uncoiling the intertwined circles of double-stranded.

Methods

Results

Conclusion