Fluoroquinolone resistance in Elizabethkingia bruuniana

Abstract

The Gram-negative bacterial genus Elizabethkingia displays natural multiple antimicrobial resistance and is an emerging pathogen. One of the therapeutic agents still effective in the treatment of the Elizabethkingia are the fluoroquinolones (e.g. ciprofloxacin). Even though these drugs are effective, the emergence of target gene mutation-mediated resistance has been reported in E. anophelis and E. meningoseptica. Fluroquinolones target DNA biosynthesis by inactivating the DNA gyrase and topoisomerase IV which are produced by the gyrAB and parCE genes. I now intend to determine the mechanisms by which Elizabethkingia species become resistant to the fluoroquinolone ciprofloxacin. I hypothesized that any fluoroquinolone-resistant mutants would exhibit compromised growth, elevated MICs, and mutations in the quinolone resistance-determining regions (QRDRs) of gyrAB and/or parCE. First, I isolated fluoroquinolone-resistant mutants via single-step selection in media containing ciprofloxacin. Standard minimum inhibitory concentration assays were performed to compare suspected mutants to the parent. I also performed checkerboard assays to assess potential synergistic interactions between ciprofloxacin and the cell wall active antibiotic vancomycin. Changes in the growth of the mutants were compared to the parent strains using standard growth curves. Finally, I utilized cloning and Sanger sequencing to determine the sequences of the QRDRs in gyrAB and parCE in the confirmed fluoroquinolone-resistant mutants of E. ursingii. I determined that fluoroquinolone-resistant mutants showed MICs elevated up to 32-fold compared to their parent strains. Checkerboard assays showed that the drug interaction was not synergistic. I demonstrated slowed growth in almost all fluoroquinolone-resistant mutants compared to their respective parent strains, suggesting a growth fitness cost from the acquisition of fluoroquinolone resistance mutations. Sanger sequencing of the QRDRs in E. bruuniana did not reveal the presence of mutations common to other fluoroquinolone-resistant Gram-negative ii bacteria. Further sequencing and experimentation are necessary to determine the mechanisms by which E. bruuniana acquires fluoroquinolone resistance.