Pharmacodynamics of colistin resistance in carbapenemase-producing Klebsiella pneumoniae: the double-edged sword of heteroresistance and adaptive resistance.

Introduction. The presence of heteroresistant subpopulations and the development of resistance during drug exposure (adaptive resistance) limits colistin's efficacy against carbapenemase-producing Klebsiella pneumoniae (CP-Kp) isolates.Hypothesis/Gap statement. The pharmacokinetic/pharmacodynamic (PK/PD) characteristics of both types of colistin resistance against CP-Kp are unknown.Aim. We therefore studied the PK/PD characteristics of colistin resistance in an in vitro PK/PD model simulating clinical colistin exposures.Methods. Two K. pneumoniae clinical isolates, one non-CP-Kp and one CP-Kp, with colistin MICs of 0.5-1 mg l-1 at a final inoculum of 107 c.f.u. ml-1 were used in an in vitro PK/PD dialysis/diffusion closed model simulating 4.5 MU q12h and 3 MU q8h clinical dosing regimens. Heteroresistant (HRS, bacteria with stable high-level resistance present before drug exposure) and adaptive resistant (ARS, bacteria with reversible low-level resistance emerging after drug exposure) subpopulations were measured and optimal PK/PD targets for reducing both ARS and HRS were determined. Cumulative fractional response (CFR) was calculated with Monte Carlo simulation for 9 MU q24h, 4.5 MU q12h and 3 MU q8h clinical dosing regimens.Results. A 2-5 log10c.f.u. ml-1 decrease of the total bacterial population was observed within the first 2 h of exposure, followed by regrowth at 12 h. Colistin exposure was positively and negatively correlated with HRS and ARS 24-0 h c.f.u. ml-1 changes, respectively. An optimal PK/PD (~0.5log10 increase) target of 35 fAUC/MIC (the ratio of the area under the unbound concentration-time curve to the MIC) was found for reducing both HRS and ARS of high-level resistance (MIC >16 mg l-1). The 4.5 MU q12h regimen had slightly higher CFR (74 %) compared to the other dosing regimens.Conclusions. High colistin exposures reduced high-level adaptive resistance at the expense of selection of heteroresistant subpopulations.

Triple combination of meropenem, colistin and tigecycline was bactericidal in a dynamic model despite mere additive interactions in chequerboard assays against carbapenemase-producing Klebsiella pneumoniae isolates.

Background: Combination schemes are commonly used for the treatment of infections due to carbapenemase-producing Klebsiella pneumoniae (CP-Kp). We therefore investigated the in vitro effectiveness of double and triple combinations of meropenem, colistin and tigecycline against CP-Kp isolates with different resistance mechanisms in a static broth microdilution model and a pharmacokinetic-pharmacodynamic model. Methods: One WT isolate and seven CP-Kp isolates with different carbapenem resistance mechanisms and increasing MICs of meropenem (4-512 mg/L), colistin (0.5-32 mg/L) and tigecycline (0.25-4 mg/L) were tested with a 3D chequerboard microdilution method. Combinations were then assessed in an in vitro pharmacokinetic-pharmacodynamic model simulating 50 and 100 mg of tigecycline q12h as a 1 h infusion, 4.5 million units of colistin q12h as a 1 h infusion and 1 g of meropenem q8h as 1 and 0.5 h infusions for 2 days. Results: In the chequerboard assay, interactions within the triple combination were mainly additive with a median (range) fractional inhibitory index of 0.66 (0.22-1.26). In the dynamic model, meropenem alone was bactericidal against isolates with MICs up to 4 mg/L, whereas bactericidal activity was found with the double combination meropenem + colistin and the triple combination meropenem + colistin + tigecycline against CP-Kp isolates with meropenem MICs of 16 and 256 mg/L, respectively. A high dose (100 mg) of tigecycline and a prolonged infusion (1 h) of meropenem increased the efficacy of the triple combination. Conclusions: Despite the merely additive interactions in the chequerboard assay, the triple combination of meropenem, tigecycline and colistin was bactericidal in the dynamic model against highly resistant CP-Kp isolates. This effect was more pronounced if prolonged infusion of meropenem and high tigecycline dosing were used.

Exploring colistin pharmacodynamics against Klebsiella pneumoniae: a need to revise current susceptibility breakpoints.

Objectives: Because the pharmacokinetic/pharmacodynamic (PK/PD) characteristics of colistin against Enterobacteriaceae are not well explored, we studied the activity of colistin against K. pneumoniae in an in vitro PK/PD model simulating different dosing regimens. Methods: Three clinical isolates of K. pneumoniae with MICs of 0.5, 1 and 4 mg/L were tested in an in vitro PK/PD model following a dose-fractionation design over a period of 24 h. A high and low inoculum of 107 and 104 cfu/mL with and without a heteroresistant subpopulation, respectively, were used. PK/PD indices associated with colistin activity were explored and Monte Carlo analysis was performed in order to determine the PTA for achieving a bactericidal effect (2 log kill). Results: The fAUC/MIC (R2 = 0.64-0.68) followed by fCmax/MIC (R2 = 0.55-0.63) best described colistin's 24 h log10 cfu/mL reduction for both low and high inocula. Dosing regimens with fCmax/MIC ≥6 were always associated with a bactericidal effect (P = 0.0025). However, at clinically achievable concentrations, usually below fCmax/MIC = 6, an fAUC/MIC ≥25 was more predictive of a bactericidal effect. Using a dosing regimen of 9 MU/day, the PTA for this pharmacodynamic target was 100%, 5%-70% and 0%, for isolates with MICs of ≤0.5, 1 and ≥2 mg/L, respectively. Dosing regimens that aim for a trough level of 1 mg/L achieve coverage of strains up to 0.5 mg/L (target trough/MIC = 2 mg/L). Conclusions: Characterization of the pharmacodynamics of colistin against Enterobacteriaceae in an in vitro model of infection indicates that a revision of current susceptibility breakpoints is needed. Therapeutic drug monitoring of colistin to achieve pharmacodynamic targets in individual patients is highly recommended.

Impact of bacterial load on pharmacodynamics and susceptibility breakpoints for tigecycline and Klebsiella pneumoniae.

OBJECTIVES: In the absence of other therapeutic options, tigecycline is used to treat bloodstream infections and pneumonia caused by carbapenemase-producing Klebsiella pneumoniae (CP-Kp). In this study, the standard and high tigecycline dosing regimens were simulated and tested against different inocula of CP-Kp isolates in an in vitro pharmacokinetic (PK)/pharmacodynamic (PD) model. METHODS: Four susceptible isolates (EUCAST MICs of 0.125-1 mg/L) and two intermediately susceptible CP-Kp clinical isolates (MICs of 2 mg/L) were tested at three different inocula (107, 105 and 103 cfu/mL), simulating tigecycline serum and lung fCmax concentrations of 0.15 and 1.5 mg/L, respectively, of 50 mg tigecycline every 12 h for 48 h. The exposure-effect relationships were described and the probability of target attainment was calculated for each inoculum in order to determine PK/PD susceptibility breakpoints. RESULTS: No cfu reduction was observed at serum concentrations. At lung concentrations and low inocula, a bacteriostatic and killing effect was found for isolates with MICs of 0.25 and 0.125 mg/L, respectively. The fAUC0-24/MIC (tAUC0-24/MIC) associated with half-maximal activity was 16 (150) with 103 cfu/mL, 28 (239) with 105 cfu/mL and 79 (590) with 107 cfu/mL. A PK/PD susceptibility breakpoint of ≤0.06 and ≤0.125 mg/L for bacteraemia with ≤101 cfu/mL and ≤0.25 and ≤0.5 mg/L for pneumonia with ≤103 cfu/g was determined for the standard tigecycline dose of 50 mg and the higher dose of 100 mg, respectively. CONCLUSIONS: Tigecycline monotherapy with either 50 or 100 mg would not be sufficient for most patients with bacteraemia, though the higher dose of 100 mg could be effective for patients with pneumonia with low bacterial load.

Comparison of Short Versus Prolonged Infusion of Standard Dose of Meropenem Against Carbapenemase-Producing Klebsiella pneumoniae Isolates in Different Patient Groups: A Pharmacokinetic-Pharmacodynamic Approach.

Dose optimization is required to increase carbapenem's efficacy against carbapenemase-producing isolates. Four clinical Klebsiella pneumoniae isolates were used: one susceptible to meropenem with minimum inhibitory concentration (MIC) 0.031 mg/L and 3 verona integron-borne metallo bete-lactamase-1-producing isolates with MICs 8, 16, and 128 mg/L. The human pharmacokinetics of short (0.5-h) and prolonged (3-h) infusion regimens of 1 g meropenem every 8 h were simulated in an in vitro pharmacokinetic-pharmacodynamic model. Time-kill curves were constructed for each isolate and dosing regimen, and the %T > MIC associated with maximal bactericidal activity was estimated. The percentage of pharmacodynamic target attainment for isolates with different MICs was calculated for 350 ICU, surgical, and internal medicine patients. The isolates with MIC ≤8 mg/L were killed with both dosing regimens. The %T > MIC corresponding to maximal bactericidal activity was ∼40%. The percentages of target attainment were >90%, 61%-83%, 23%-33%, and <3% with the short infusion regimen and >90%, 98%-99%, 55%-79%, and <5% with the prolonged infusion regimen for isolates with MIC ≤2, 4, 8, and ≥16 mg/L, respectively. The lowest target attainment rates were observed for the ICU patients and the highest for internal medicine patients. The prolonged infusion regimen was more effective than the short infusion regimen against isolates with MIC 4-8 mg/L.

Pharmacokinetic-pharmacodynamic modelling of meropenem against VIM-producing Klebsiella pneumoniae isolates: clinical implications.

VIM-producing Klebsiella pneumoniae isolates are usually associated with high MICs to carbapenems. Preclinical studies investigating the pharmacokinetic-pharmacodynamic (PK-PD) characteristics of carbapenems against these isolates are lacking. The in vitro antibacterial activity of meropenem against one WT and three VIM-producing K. pneumoniae clinical isolates (median MICs 0.031, 8, 16 and 128 mg l- 1) was studied in a dialysis-diffusion PK-PD model and verified in a thigh infection neutropenic animal model by testing selected strains and exposures. The in vitro PK-PD target associated with bactericidal activity was estimated and the target attainment for different dosing regimens was calculated with Monte Carlo analysis. The in vitro model was correlated with the in vivo data, with log10CFU/ml reduction of < 1 for the VIM-producing (MIC 16 mg l- 1) and >2 for the WT (MIC 0.031 mg l- 1) isolates, with %f T >MIC 25 and 100%, respectively. The in vitro bactericidal activity for all isolates was associated with 40 % f T>MIC and attained in >90% of cases with the standard 1 g q8 0.5 h infusion dosing regimen only for isolates with MICs up to 1 mg l- 1. For isolates with MICs of 2-8 mg l- 1, prolonged infusion regimens (4 h infusion q8 or 2 h infusion q4) of standard (1 g) and higher (2 g) doses or continuous infusion regimens (3-6 g) are required. For isolates with a MIC of 16 mg l- 1 the unconventional dosing regimen of 2 g as 2 h infusion q4 or 12 g continuous infusion will be required. Prolonged and continuous infusion regimens of meropenem may increase efficacy against VIM-producing K. pneumoniae isolates.

Evaluation of the "Dip Effect" Phenomenon in Antifungal Susceptibility Testing of Candida spp. against Echinocandins by Use of Gradient Concentration Strips.

The "dip effect" phenomenon complicates antifungal susceptibility testing with gradient concentration strips. Of 60 Candida isolates tested with the three echinocandins, this phenomenon was observed only for caspofungin with most (>90%) Candida albicans, Candida glabrata, and Candida tropicalis isolates and for isolates with CLSI MICs of ≤0.25 mg/liter. In order to facilitate MIC determination, a practical approach was developed using the inhibition zones at 32, 8, 2, and 1 mg/liter, increasing the agreement with the CLSI method >86%.