A Novel Mismatched PCR-Restriction Fragment Length Polymorphism Assay for Rapid Detection of gyrA and parC Mutations Associated With Fluoroquinolone Resistance in Acinetobacter baumannii

• DOI: 10.3343/alm.2020.40.1.27
• Publication Date: 2019-08-21T07:23:56Z
• Authors: Naoki Kakuta; Ryuichi Nakano; Akiyo Nakano; Yuki Suzuki; Ayako Tanouchi; Takashi Masui; Saori Horiuchi; Shiro Endo; Risako Kakuta; Yasuo Ono; Hisakazu Yano
• Keywords: Acinetobacter baumannii

Mutations in the quinolone resistance-determining regions (QRDRs) of Acinetobacter baumannii DNA gyrase (gyrA) and topoisomerase IV (parC) are linked to fluoroquinolone (FQ) resistance. We developed a mismatched PCR-restriction fragment length polymorphism (RFLP) assay detect mutations gyrA parC QRDRs associated with FQ resistance A. baumannii.Based on conserved sequences baumanniigyrA parC, two primer sets were designed for PCR-RFLP (codons 83 87) 80 84) by introducing an artificial restriction enzyme cleavage site into PCR products. This was evaluated using 58 strains 37 other that have been identified RNA polymerase β-subunit gene sequence analysis.PCR amplification successful all strains. In 11 -susceptible strains, products digested selected enzymes at containing 84). from 47 FQ-resistant not mutation. As non-baumannii although obtained 28 no product any strain.This specifically amplified detected