AUTHOR=Ota Yusuke , Okada Reina , Takahashi Hideyuki , Saito Ryoichi 

TITLE=Molecular detection of fluoroquinolone-resistant Neisseria meningitidis by using mismatched PCR-restriction fragment length polymorphism technique

JOURNAL=Frontiers in Cellular and Infection Microbiology

VOLUME=12

YEAR=2022

URL=https://www.frontiersin.org/journals/cellular-and-infection-microbiology/articles/10.3389/fcimb.2022.911911

DOI=10.3389/fcimb.2022.911911

ISSN=2235-2988

ABSTRACT=<p>Ciprofloxacin (CIP) is a commonly used antibiotic for meningococcal chemoprophylaxis, and the mutations in the quinolone resistance-determining region of <italic>gyrA</italic> are associated with CIP-resistant <italic>Neisseria meningitidis</italic>. Here, we established a mismatched PCR-restriction fragment length polymorphism (RFLP) assay to detect a mutation at codon 91 of <italic>gyrA</italic>, followed by high-level CIP-resistant meningococci. We designed PCR-RFLP primers to detect the T91I mutation in <italic>gyrA</italic> by introducing an artificial <italic>Aci</italic>I cleavage site. This assay was performed using 26 <italic>N. meningitidis</italic> strains whose <italic>gyrA</italic> sequences have been characterized. The amplified 160 bp PCR product from <italic>gyrA</italic> was digested into three fragments (80, 66, and 14 bp) when there was no mutation, or two fragments (146 and 14 bp) when there was a mutation at codon 91. A correlation was observed between the mismatched PCR-RFLP assay and <italic>gyrA</italic> sequencing. This rapid, simple, and accurate assay has the potential to detect CIP-resistant <italic>N. meningitidis</italic> in clinical microbiology laboratories, contributing to the appropriate antibiotic selection for meningococcal chemoprophylaxis, will help maintain an effective treatment for close contacts of IMD patients, and prevent the spread of CIP-resistant <italic>N. meningitidis</italic>.</p>