Impact of Ciprofloxacin Exposure on Staphylococcus aureus Genomic Alterations Linked with Emergence of Rifampin Resistance

Abstract

ABSTRACTIntensive use of antimicrobial agents in health care settings not only leads to the selection of multiresistant nosocomial isolates ofStaphylococcus aureusbut may also promote endogenous, resistance-conferring mutations in bacterial genes that encode drug targets. We evaluated the spectrum of rifampin resistance-conferring mutations in cultures of methicillin-susceptibleS. aureus(MSSA) or methicillin-resistantS. aureus(MRSA) strains exposedin vitroto sub-MICs of ciprofloxacin. Growth of ciprofloxacin-susceptible MRSA strain MRGR3 and ciprofloxacin-resistant MSSA strain RA1 (a NCTC 8325 derivative) in the presence of 1/2× or 1/4× MIC of ciprofloxacin led to higher frequencies of rifampin-resistant mutants on agar supplemented with rifampin (0.25 mg/liter) than under ciprofloxacin-free conditions. While rifampin-resistant mutants from ciprofloxacin-free cultures essentially showed single-amino-acid substitutions, a significant proportion of rifampin-resistant mutants from ciprofloxacin-exposed cultures displayed in-frame deletions or insertions in therpoBgene at several positions of the rifampin resistance cluster I. In-frame deletions or insertions were also recorded inrpoBcluster I of rifampin-resistant mutants from ciprofloxacin-exposed cultures ofmutSandmutLDNA repair mutants of ciprofloxacin-resistantS. aureusstrain RA1. Frequencies of rifampin-resistant mutants grown under ciprofloxacin-free medium were higher for mutant strains RA1mutS2and RA1mutL, but not RA1recA, than for their parent RA1. In conclusion, ciprofloxacin-mediated DNA damage inS. aureus, as exemplified by the wide diversity of deletions or insertions inrpoB, suggests the occurrence of major, quinolone-mediated disturbances in DNA fork progression and replication repair. Besides promoting antibiotic resistance, accumulation of unrepaired DNA replication errors, including insertions and deletions, may also contribute to potentially lethal mutations.