Use of an efflux pump inhibitor to determine the prevalence of efflux pump-mediated fluoroquinolone resistance and multidrug resistance in Pseudomonas aeruginosa

Abstract

Fluoroquinolone-resistance in Pseudomonas aeruginosa may be due to efflux pump overexpression (EPO) and/or target mutations. EPO can result in multidrug resistance (MDR) due to broad substrate specificity of the pumps. MC-04,124, an efflux pump inhibitor (EPI) shown to significantly potentiate activity of levofloxacin in P. aeruginosa, was used to examine the prevalence of EPO in clinical isolates. MICs were determined for ciprofloxacin, levofloxacin, moxifloxacin, and gatifloxacin with or without EPI and for other antipseudomonal agents by using broth microdilution against P. aeruginosa isolates from adults (n = 119) and children (n = 24). The prevalence of the EPO phenotype (>/=8-fold MIC decrease when tested with EPI) was compared among subgroups with different resistance profiles. The EPO phenotype was more prevalent among levofloxacin-resistant than levofloxacin-sensitive strains (61%, 48/79 versus 9%, 6/64). EPO was present in 60% of fluoroquinolone-resistant strains without cross-resistance, while it was present at variable frequencies among strains with cross-resistance to other agents: piperacillin-tazobactam (86%), ceftazidime (76%), cefepime (65%), imipenem (56%), gentamicin (55%), tobramycin (48%), and amikacin (27%). The magnitude of MIC decrease with an EPI paralleled the frequency of which the EPO phenotype was observed in different subgroups. EPI reduced the levofloxacin MIC by as much as 16-fold in eight strains for which MICs were 128 microg/ml. Efflux-mediated resistance appears to contribute significantly to fluoroquinolone resistance and MDR in P. aeruginosa. Our data support the fact that increased fluoroquinolone usage can negatively impact susceptibility of P. aeruginosa to multiple classes of antipseudomonal agents.

FIG. 1.

Frequency of the efflux pump-overexpressed phenotype (EP) in LvxR strains showing cross-resistance to non-FQ antipseudomonal agents. n, number of LvxR isolates showing cross-resistance to the respective agent. CAZ, ceftazidime; FEP, cefepime; TZP, piperacillin-tazobactam; GEN, gentamicin; TOB, tobramycin; AMK, amikacin.

FIG. 2.

Effect of EPI on fluoroquinolone MICs. All strains were tested over the same range of concentrations (0.5 to 8 μg/ml) for all four FQ agents. n, number of strains showing at least an 8-fold decrease in MIC with an EPI; an additional 28 isolates showed a ≥8-fold decrease in the LVX MIC when tested at a broader concentration range (0.25 to 256 μg/ml). P was ≤0.0002 for CIP versus MOX, LVX, or GAT. P was >0.05 for comparison amongst MOX, LVX, and GAT.