Pyrosequencing of the DNA gyrase gene in Neisseria species: effective indicator of ciprofloxacin resistance in Neisseria gonorrhoeae

Abstract

The quinolone resistance determining region (QRDR) of the gyrA gene in ciprofloxacin-susceptible strains (n=53) and strains of Neisseria spp. with reduced susceptibility (n=70) was determined by the pyrosequencing method. Results showed that the QRDR of the gyrA gene is an effective molecular indicator of resistance to ciprofloxacin in Neisseria gonorrhoeae, and presumably in Neisseria meningitidis, but not in all other Neisseria spp. This sequence was not unique for N. gonorrhoeae and seems unsuitable for species verification of N. gonorrhoeae. However, whether it is also possible to use this region for verification depends on the specificity of the primary screening method used.

Combined Real-Time PCR and Pyrosequencing Strategy for Objective, Sensitive, Specific, and High-Throughput Identification of Reduced Susceptibility to Penicillins in Neisseria meningitidis

“…By using real-time PCR and pyrosequencing for examination of the two Pen i -specific sites, rapid sequence data that are objective, sensitive, specific, portable for comparison between laboratories, and reproducible for detection of reduced susceptibility to ␤-lactam antibiotics in N. meningitidis isolates are generated. Pyrosequencing has also previously been beneficially used for detection of antibiotic resistance in bacteria (4,5,9,10). Pyrosequencing is accurate and reproducible and enables reading from the first base after the sequencing primer.…”

DNA Pyrosequencing-Based Bacterial Pathogen Identification in a Pediatric Hospital Setting

“…Since its inception in the mid-1990s, DNA pyrosequencing assays have been developed for diverse applications, including genotyping, single nucleotide polymorphism detection, and microorganism identification (21). Pyrosequencing has been used to detect point mutations in antiviral or antimicrobial resistance genes as a strategy for molecular resistance testing (12,20,34). Pyrosequencing has been applied to organism identification by combining short-stretch DNA sequencing with signature matching in the well-characterized phylogenetic target, the 16S rRNA gene (14,30), in addition to in a variety of target genes in bacteria (9,13,24,32).…”