Influence of different antibiotic groups on the development of mutational resistance to colistin among Klebsiella pneumoniae

Abstract

Objective. To determine the concentration of colistin, preventing the selection of colistin-resistant mutants of K. pneumoniae, and to evaluate the effect of antibiotics of different groups on the development of mutational resistance to colistin. Materials and Methods. Minimum inhibitory concentrations (MIC) of colistin were determined for 88 K. pneumoniae strains by the method of serial microdilutions in broth, and carbapenemase genes were detected. The selection of colistin-resistant subpopulations was performed on cation-adjusted MüllerHinton agar (MHA) with the addition of 16 mg/l colistin. Mutant prevention concentration (MPC) of colistin is determined on MHA containing 0, 1, 2, 4, 8, 16, 32, 64 and 128 mg/l of colistin. Also, MPCs of colistin were determined in the presence of a fixed concentration of the second antibiotic: clarithromycin (2 mg/l), azithromycin (2 mg/l), rifampicin (1 mg/l), clindamycin (0.5 mg/l), meropenem (8 mg/l), linezolid (2 mg/l), amikacin (1 mg/l), vancomycin (2 mg/l), doxycycline (2 mg/l). Results. All strains remained susceptible to colistin (colistin MIC 0.06–1.0 mg/l). Resistance to meropenem (MIC > 8 mg/l) was detected in 48 strains (54.5%), 46 of them were carbapenemase producers: KPC – 6 strains (6.8%), OXA-48 – 26 strains (29.5%), NDM – 14 strains (15.9%). Growth of colonies on MHA with 16 mg/l of colistin was found for 96.6% of the strains. The frequency of mutational resistance occurrence ranged from 6 × 10-9 to 10-6 (median: 2 × 10-7). The mutational nature of colistin resistance was confirmed for 36.4% of the strains. The MPC values of colistin were in the range of 16–256 mg/l; (MPC50 32 mg/l, MPC90 256 mg/l) and significantly (32–1024 times) exceeded the MIC values. In the presence of 1 mg/l of rifampicin, the MPC of colistin decreased 4–64 times (MPC50 4 mg/l, MPC90 4 mg/l). In the presence of 2 mg/l of doxycycline, MPC of colistin decreased 2–64 times for all strains (MPC50 8 mg/l, MPC90 16 mg/l). The presence of linezolid (2 mg/l) and vancomycin (2 mg/l) did not significantly change MPC of colistin. Meropenem at a concentration of 8 mg/l had no significant effect on colistin MPC for carbapenemase-producing K. pneumoniae strains. None of the antibiotics lowered the MPC50 of colistin to its clinically achievable serum concentrations. Conclusions. A high frequency of formation of mutational resistance to colistin in K. pneumoniae was revealed. The MPC values of colistin are outside the range of clinically achievable serum concentrations and may decrease in the presence of other antibiotics.

Keywords

• klebsiella pneumoniae
• colistin
• rifampicin
• mutational resistance
• antibiotic combinations