Identification of distinct ciprofloxacin susceptibility in Acinetobacter spp. by detection of the gyrA gene mutation using real-time PCR

Ciprofloxacin-resistant (CipR) Acinetobacter spp. was associated with a mutation in quinolone resistance-determining region (QRDR) of gyrA gene from Ser83 to Leu83. A total of 54 Acinetobacter baumannii, 11 A. genospecies 3, and 17 A. genospecies 13TU clinical isolates were determined for ciprofloxacin susceptibility and their QRDR sequenced. Most of A. baumannii were CipR and had a mutated QRDR of gyrA gene, and all A. genospecies 3 and 13TU isolates were ciprofloxacin-susceptible (CipS) and had a wildtype QRDR of gyrA gene. A real-time PCR assay was developed to rapidly differentiate the CipR and CipS Acinetobacter spp. This assay was based on probe hybridization followed by melting temperature (T-m) analysis to discriminate the QRDR of gyrA gene. All CipR strains had a T-m at 47 degrees C, and most of CipS strains had a higher T-m at 51.5 degrees C. Four CipS A. genospecies 3 isolates with an A base at 264 nt of the gyrA gene had a T-m at 49.5 degrees C. The assay can rapidly and accurately identify the mutated QRDR of gyrA gene in CipR Acinetobacter spp., and potentially increase the rate of the appropriate therapy for Acinetobacter infections. (C) 2009 Elsevier Ltd. All rights reserved.