RESUMO
Resistance to carbapenems has become a problem due to Klebsiella pneumoniae (K. pneumoniae), harboring carbapenemases. Among them, there are isolates that are recognized as carbapenem-susceptible; however, these carbapenemase-producing strains with low meropenem minimal inhibitory concentrations (MICs) may pose a threat to public health. We aimed to investigate the impact of the ability to produce carbapenemases by a bacterial isolate on the effectiveness of meropenem in the hollow-fiber infection model. K. pneumoniae and Escherichia coli (E. coli) strains with equal meropenem MICs but differing in their ability to produce carbapenemases were used in pharmacodynamic simulations with meropenem. In addition to standard MIC determination, we assessed the MICs against tested strains at high inoculum density to test if the inoculum effect occurs. According to pharmacodynamic data, the carbapenemase-producing strains were characterized with a relatively decreased meropenem effectiveness compared to non-producers. Meanwhile, the effect of meropenem perfectly correlated with the meropenem exposure expressed as the DOSE/MIC ratio when high-inoculum (HI) MICs but not standard-inoculum (SI) MICs were used for regression analysis. It could be concluded that meropenem-susceptible carbapenemase-producing strains may not respond to meropenem therapy; the antibiotic inoculum effect (IE) may have a prognostic value to reveal the meropenem-susceptible Enterobacterales that harbor carbapenemase genes.
RESUMO
OXA-48 carbapenemases are frequently expressed by Klebsiella pneumoniae clinical isolates; they decrease the effectiveness of carbapenem therapy, particularly with meropenem. Among these isolates, meropenem-susceptible carbapenemase-producers may show decreased meropenem effectiveness. However, the probability of the emergence of resistance in susceptible carbapenemase-producing isolates and its dependence on specific K. pneumoniae meropenem MICs is not completely known. It is also not completely clear what resistance patterns will be exhibited by these bacteria exposed to meropenem, if they would follow the patterns of non-beta-lactamase-producing bacteria and other than beta-lactams antibiotics. These issues might be clarified if patterns of meropenem resistance related to the mutant selection window (MSW) hypothesis. To test the applicability of the MSW hypothesis to meropenem, OXA-48-carbapenemase-producing K. pneumoniae clinical isolates with MICs in a 64-fold range (from susceptible to resistant) were exposed to meropenem in a hollow-fiber infection model; epithelial lining fluid meropenem pharmacokinetics were simulated following administration of 2 grams every 8 hours in a 3-hour infusion. Strong bell-shaped relationships between the meropenem daily dose infused to the model as related to the specific isolate MIC and both the antimicrobial effect and the emergence of resistance were observed. The applicability of the MSW hypothesis to meropenem and carbapenemase producing K. pneumoniae was confirmed. Low meropenem efficacy indicates very careful prescribing of meropenem to treat K. pneumoniae infections when the causative isolate is confirmed as an OXA-48-carbapenemase producer.
Assuntos
Antibacterianos , Infecções por Klebsiella , Humanos , Meropeném/farmacologia , Antibacterianos/farmacologia , Klebsiella pneumoniae , Proteínas de Bactérias/genética , Proteínas de Bactérias/farmacologia , beta-Lactamases/genética , beta-Lactamases/farmacologia , Carbapenêmicos/farmacologia , Testes de Sensibilidade Microbiana , Infecções por Klebsiella/tratamento farmacológico , Infecções por Klebsiella/microbiologiaRESUMO
The development and implementation of diagnostic methods that allow rapid assessment of antibiotic activity against pathogenic microorganisms is an important step towards antibiotic therapy optimization and increase in the likelihood of successful treatment outcome. To determine whether fluorescence microscopy with acridine orange can be used for rapid assessment (≤8 h) of the meropenem activity against Klebsiella pneumoniae, six isolates including three OXA-48-carbapenemase-producers were exposed to meropenem at different levels of its concentration (0.5 × MIC, 1 × MIC, 8 or 16 µg/mL) and the changes in the viable counts within 24 h were evaluated using fluorescence microscopy and a control culture method. The approach was to capture the regrowth of bacteria as early as possible. Within the first 8 h fluorescence microscopy allowed to categorize 5 out of 6 K. pneumoniae strains by their meropenem susceptibility (based on the MIC breakpoint of 8 mg/L), but meropenem activity against three isolates, two of which were OXA-48-producers, could not be accurately determined at 8 h. The method proposed in our study requires improvement in terms of accelerating the bacterial growth and regrowth for early meropenem MIC determination. Volume-dependent elevation in meropenem MICs against OXA-48-producers was found and this phenomenon should be studied further.
RESUMO
The minimal inhibitory concentration (MIC) is conventionally used to define in vitro levels of susceptibility or resistance of a specific bacterial strain to an antibiotic and to predict its clinical efficacy. Along with MIC, other measures of bacteria resistance exist: the MIC determined at high bacterial inocula (MICHI) that allow the estimation of the occurrence of inoculum effect (IE) and the mutant prevention concentration, MPC. Together, MIC, MICHI and MPC represent the bacterial "resistance profile". In this paper, we provide a comprehensive analysis of such profiles of K. pneumoniae strains that differ by meropenem susceptibility, ability to produce carbapenemases and specific carbapenemase types. In addition, we have analyzed inter-relations between the MIC, MICHI and MPC for each tested K. pneumoniae strain. Low IE probability was detected with carbapenemase-non-producing K. pneumoniae, and high IE probability was detected with those that were carbapenemase-producing. MICs did not correlate with the MPCs; significant correlation was observed between the MICHIs and the MPCs, indicating that these bacteria/antibiotic characteristics display similar resistance properties of a given bacterial strain. To determine the possible resistance-related risk due to a given K. pneumoniae strain, we propose determining the MICHI. This can more or less predict the MPC value of the particular strain.
RESUMO
The inoculum effect (IE) is a well-known phenomenon with beta-lactams. At the same time, the IE has not been extensively studied with carbapenem/carbapenemase inhibitor combinations. The antibiotic-to-inhibitor concentration ratio used in susceptibility testing can influence the in vitro activity of the combination. To explore the role of these factors, imipenem/relebactam and doripenem/relebactam MICs were estimated against six Klebsiella pneumoniae carbapenemase (KPC)-producing Klebsiella pneumoniae strains at standard inocula (SI) and high inocula (HI) by two methods: with a fixed relebactam concentration and with a fixed, pharmacokinetic-based carbapenem-to-relebactam concentration ratio. The combination MICs at HI, compared to SI, increased with most of the tested strains. However, the IE occurred with only two K. pneumoniae strains regardless of the MIC testing method. The relationship between the MICs at SI and the respective inoculum-induced MIC changes was observed when the MICs were estimated at pharmacokinetic-based carbapenem-to-relebactam concentration ratios. Thus, (1) IE was observed with both carbapenem/relebactam combinations regardless of the MIC testing method; however, IE was not observed frequently among tested K. pneumoniae strains. (2) At HI, carbapenem/relebactam combination MICs increased to levels associated with carbapenem resistance. (3) Combination MICs determined at pharmacokinetic-based carbapenem-to-inhibitor concentration ratios predict susceptibility elevations at HI in KPC-producing K. pneumoniae.
RESUMO
OBJECTIVES: To explore whether linezolid/daptomycin combinations can restrict Staphylococcus aureus resistance and if this restriction is associated with changes in the mutant prevention concentrations (MPCs) of the antibiotics in combination, the enrichment of resistant mutants was studied in an in vitro dynamic model. METHODS: Two MRSA strains, vancomycin-intermediate resistant ATCC 700699 and vancomycin-susceptible 2061 (both susceptible to linezolid and daptomycin), and their linezolid-resistant mutants selected by passaging on antibiotic-containing medium were used in the study. MPCs of antibiotics in combination were determined at a linezolid-to-daptomycin concentration ratio (1:2) that corresponds to the ratio of 24 h AUCs (AUC24s) actually used in the pharmacokinetic simulations. Each S. aureus strain was supplemented with respective linezolid-resistant mutants (mutation frequency 10-8) and treated with twice-daily linezolid and once-daily daptomycin, alone and in combination, simulated at therapeutic and sub-therapeutic AUC24s. RESULTS: Numbers of linezolid-resistant mutants increased at therapeutic and sub-therapeutic AUC24s, whereas daptomycin-resistant mutants were enriched only at sub-therapeutic AUC24 in single drug treatments. Linezolid/daptomycin combinations prevented the enrichment of linezolid-resistant S. aureus and restricted the enrichment of daptomycin-resistant mutants. The pronounced anti-mutant effects of the combinations were attributed to lengthening the time above MPC of both linezolid and daptomycin as their MPCs were lowered. CONCLUSIONS: The present study suggests that (i) the inhibition of S. aureus resistant mutants using linezolid/daptomycin combinations can be predicted by MPCs determined at pharmacokinetically derived antibiotic concentration ratios and (ii) T>MPC is a reliable predictor of the anti-mutant efficacy of antibiotic combinations as studied using in vitro dynamic models.
Assuntos
Daptomicina , Staphylococcus aureus Resistente à Meticilina , Antibacterianos/farmacologia , Daptomicina/farmacologia , Farmacorresistência Bacteriana , Linezolida/farmacologia , Testes de Sensibilidade Microbiana , Staphylococcus aureus/genéticaRESUMO
To explore if the time inside the mutant selection window (TMSW) is a reliable predictor of emergence of bacterial resistance to linezolid, mixed inocula of each of three methicillin-resistant Staphylococcus aureus strains (MIC of linezolid 2 µg ml-1) and their previously selected resistant mutants (MIC 8 µg ml-1) were exposed to linezolid pharmacokinetics using an in vitro dynamic model. In five-day treatments simulated over a wide range of the 24-h area under the concentration-time curve (AUC24) to the MIC ratio, mutants resistant to 4 × MIC of antibiotic were enriched in a TMSW-dependent manner. With each strain, TMSW relationships with the area under the bacterial mutant concentration-time curve (AUBCM) exhibited a hysteresis loop, with the upper portion corresponding to the time above the mutant prevention concentration (MPC; T>MPC) of 0 and the lower portion-to the T>MPC > 0. Using AUBCM related to the maximal value observed with a given strain (normalized AUBCM) at T>MPC > 0, a strain-independent sigmoid relationship was established between AUBCM and TMSW, as well as T>MPC (r2 0.99 for both). AUC24/MIC and AUC24/MPC relationships with normalized AUBCM for combined data on the three studied S. aureus strains were bell-shaped (r2 0.85 and 0.80, respectively). These findings suggest that TMSW at T>MPC > 0, T>MPC, AUC24/MIC and AUC24/MPC are useful bacterial strain-independent predictors of the emergence of staphylococcal resistance to linezolid.
Assuntos
Antibacterianos/farmacologia , Farmacorresistência Bacteriana/genética , Linezolida/farmacologia , Staphylococcus aureus Resistente à Meticilina/efeitos dos fármacos , Mutação , Algoritmos , Área Sob a Curva , Cinética , Testes de Sensibilidade Microbiana , Seleção GenéticaRESUMO
To explore the relationship between pharmacokinetic variables and enterococcal resistance to linezolid, a vancomycin-resistant strain whose mutant prevention concentration (MPC) exceeded the MIC by two fold was selected among six clinical isolates of Enterococcus faecium. The selected strain was exposed to simulated pharmacokinetics of twice-daily linezolid for five days. Mutants resistant to 2 × MIC of the antibiotic were enriched at ratios of the 24-h area under the concentration-time curve (AUC24) to the MIC of 15 and 30 h but not at 60 and 120 h. These observations could be explained by the different times when antibiotic concentrations exceed the MPC (T>MPC): 0 to 14, 63 and 100% of the dosing interval. Using the area under the bacterial mutant concentration-time curve (AUBCM) determined in this study and in previous work with other E. faecium strains (MPC/MIC 4), a strain-independent T>MPC relationship with mutant enrichment was established.
Assuntos
Enterococcus faecium/efeitos dos fármacos , Linezolida/farmacologia , Antibacterianos/farmacologia , Área Sob a Curva , Farmacorresistência Bacteriana/efeitos dos fármacos , Infecções por Bactérias Gram-Positivas/tratamento farmacológico , Humanos , Linezolida/farmacocinética , Testes de Sensibilidade Microbiana/métodos , Resistência a Vancomicina/efeitos dos fármacosRESUMO
OBJECTIVES: To test the mutant selection window (MSW) hypothesis applied to linezolid-exposed Staphylococcus aureus and to delineate the concentration-resistance relationship, a mixed inoculum of linezolid-susceptible S. aureus cells and linezolid-resistant mutants (RMs) was exposed to linezolid multiple dosing. METHODS: Three S. aureus strains (MIC of linezolid 2 mg/L), S. aureus 479, S. aureus 688 and S. aureus ATCC 700699, and their RMs (MIC 8 mg/L) selected by passaging on antibiotic-containing media were used in the study. RMs of S. aureus 479 and S. aureus ATCC 700699 contained a G2576T replacement (Escherichia coli numbering) in one of the copies of the 23S rRNA gene, which had been reported in clinically isolated mutants. Five-day treatments with twice-daily linezolid were simulated over a 32-fold range of the 24 h AUC (AUC24) to the MIC ratio. RESULTS: A pronounced enrichment of mutants resistant to 2×, 4× and 8× MIC was observed at AUC24/MIC ratios of 30 and 60 when linezolid concentrations were within the MSW for more than half of the dosing interval for each strain. The number of viable mutant cells decreased significantly at the simulated AUC24/MIC ratio of 120, while the AUC24/MIC ratio of 240 completely prevented mutant enrichment in vitro. Bell-shaped AUBCM-AUC24/MIC and AUBCM-AUC24/MPC relationships (r2 0.91 and 0.79, respectively) were strain independent. CONCLUSIONS: The bell-shaped pattern of AUC24/MIC and AUC24/MPC relationships with S. aureus resistance to linezolid is consistent with the MSW hypothesis. 'Antimutant' AUC24/MIC ratios were predicted based on the AUC24/MIC relationship with AUBCM.