Molecular characterization of moxifloxacin resistance from Canadian Clostridium difficile clinical isolates.

Fluoroquinolone resistance in Clostridium difficile has been implicated in recent outbreaks of C. difficile infection. The purpose of this report was to characterize the molecular mechanism conferring resistance to moxifloxacin among C. difficile clinical isolates. Eighty-four C. difficile clinical isolates (collected as part of the Canadian Nosocomial Infection Surveillance Program) were evaluated in the current study. Pulsed-field gel electrophoresis was used to type the isolates. Susceptibility testing was performed using Clinical and Laboratory Standards Institute agar dilution methods. The quinolone resistance-determining region of both gyrA and gyrB was amplified using polymerase chain reaction and sequenced for each isolate. The proportion of isolates studied by the North American pulsed-field (NAP) type was as follows: NAP1 (47.6%), NAP2 (20.2%), NAP3 (5.9%), NAP4 (4.8%), NAP5 (2.4%), NAP6 (3.6%), and other patterns (15.5%). All isolates were resistant to ciprofloxacin. Among moxifloxacin-susceptible isolates (MIC < or =2 microg/mL), no amino acid substitutions were detected in either GyrA or GyrB. Three distinct amino acid substitutions were observed among the 3 isolates that had a moxifloxacin MIC of 8 microg/mL (GyrA Asp71 to Val, GyrB Asp426 to Asn, or Glu466 to Val). Isolates with a moxifloxacin MIC of 16 or 32 microg/mL (moderate-level resistance) all had a single identical amino acid substitution in GyrA (Thr82 to Ile). For isolates with a moxifloxacin MIC of > or =64 microg/mL (high-level resistance), this Thr82 to Ile substitution in GyrA was accompanied by at least 1 other amino acid substitution in either GyrA (Asp71 to Glu, Pro116 to Ala, or Ala118 to Ser) or GyrB (Ser366 to Ala, Asp426 to Asn, Asp426 to Val, or Leu444 to Phe) in all but 1 case. Moderate-level moxifloxacin resistance was associated with a single substitution in GyrA. High-level moxifloxacin resistance was associated with this GyrA substitution plus at least 1 other substitution in GyrA or GyrB.