Impact of Different Carbapenems and Regimens of Administration on Resistance Emergence for Three Isogenic Pseudomonas aeruginosa Strains with Differing Mechanisms of Resistance

ABSTRACT We compared drugs (imipenem and doripenem), doses (500 mg and 1 g), and infusion times (0.5 and 1.0 [imipenem], 1.0 and 4.0 h [doripenem]) in our hollow-fiber model, examining cell kill and resistance suppression for three isogenic strains of Pseudomonas aeruginosa PAO1. The experiments ran for 10 days. Serial samples were taken for total organism and resistant subpopulation counts. Drug concentrations were determined by high-pressure liquid chromatography-tandem mass spectrometry (LC/MS/MS). Free time above the MIC (time > MIC) was calculated using ADAPT II. Time to resistance emergence was examined with Cox modeling. Cell kill and resistance emergence differences were explained, in the main, by differences in potency (MIC) between doripenem and imipenem. Prolonged infusion increased free drug time > MIC and improved cell kill. For resistance suppression, the 1-g, 4-h infusion was able to completely suppress resistance for the full period of observation for the wild-type isolate. For the mutants, control was ultimately lost, but in all cases, this was the best regimen. Doripenem gave longer free time > MIC than imipenem and, therefore, better cell kill and resistance suppression. For the wild-type organism, the 1-g, 4-h infusion regimen is preferred. For organisms with resistance mutations, larger doses or addition of a second drug should be studied.

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The Combination of Meropenem and Levofloxacin Is Synergistic with Respect to both Pseudomonas aeruginosa Kill Rate and Resistance Suppression

Antimicrobial Agents and Chemotherapy ◽

10.1128/aac.00065-10 ◽

2010 ◽

Vol 54 (6) ◽

pp. 2646-2654 ◽

Cited By ~ 46

Author(s):

Arnold Louie ◽

Caroline Grasso ◽

Nadzeya Bahniuk ◽

Brian Van Scoy ◽

David L. Brown ◽

...

Keyword(s):

Pseudomonas Aeruginosa ◽

Combination Therapy ◽

Infection Model ◽

Combination Regimen ◽

Time Data ◽

Kaplan Meier ◽

Drug Concentrations ◽

Cell Kill ◽

The Impact

ABSTRACT New approaches are needed for the treatment of Pseudomonas aeruginosa infections. All available single agents are suboptimal, especially for resistance suppression. Classical β-lactam/aminoglycoside combinations are not used often enough at least in part because of concern for nephrotoxicity. We evaluated the combination of meropenem and levofloxacin against the P. aeruginosa PAO1 wild type and its isogenic MexAB pump-overexpressed mutant. The drugs were studied using an in vitro hollow-fiber pharmacodynamic infection model. There were 16 different regimens evaluated for both isolates. Both total population and resistant subpopulations were quantified. Drug concentrations were measured by liquid chromatography-tandem mass spectrometry (LC-MS-MS). The impact of monotherapy versus that of combination therapy for attainment of a 3-log cell kill and for resistance suppression was examined using Kaplan-Meier analysis. Drug exposures were calculated by fitting the concentration-time data using the ADAPT II package of programs. For both isolates, monotherapy allowed resistance emergence with all but the largest exposure or with all exposures. In contrast, there was no resistance emergence with any combination regimen. Kaplan-Meier analysis showed significant differences in time to attainment of a 3-log cell kill as well as time to resistance emergence for monotherapy and combination therapy for both isolates, in favor of the combination regimens. Determination of the pharmacodynamic indices associated with resistance suppression demonstrated a 2- to 3-fold reduction with the use of combinations. Combination therapy with meropenem and levofloxacin provides a significantly faster time to attain a 3-log cell kill and significantly better resistance suppression than does either monotherapy. This combination should be evaluated in a clinical trial.

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Antibiotic Resistance Evolution Is Contingent on the Quorum-Sensing Response in Pseudomonas aeruginosa

Molecular Biology and Evolution ◽

10.1093/molbev/msz144 ◽

2019 ◽

Vol 36 (10) ◽

pp. 2238-2251 ◽

Cited By ~ 9

Author(s):

Sara Hernando-Amado ◽

Fernando Sanz-García ◽

José Luis Martínez

Keyword(s):

Pseudomonas Aeruginosa ◽

Antibiotic Resistance ◽

Quorum Sensing ◽

Wild Type ◽

Resistance Mutations ◽

Epistatic Interactions ◽

Adaptive Mutations ◽

Resistance Evolution ◽

Evolutionary Trajectories

Abstract Different works have explored independently the evolution toward antibiotic resistance and the role of eco-adaptive mutations in the adaptation to a new habitat (as the infected host) of bacterial pathogens. However, knowledge about the connection between both processes is still limited. We address this issue by comparing the evolutionary trajectories toward antibiotic resistance of a Pseudomonas aeruginosa lasR defective mutant and its parental wild-type strain, when growing in presence of two ribosome-targeting antibiotics. Quorum-sensing lasR defective mutants are selected in P. aeruginosa populations causing chronic infections. Further, we observed they are also selected in vitro as a first adaptation for growing in culture medium. By using experimental evolution and whole-genome sequencing, we found that the evolutionary trajectories of P. aeruginosa in presence of these antibiotics are different in lasR defective and in wild-type backgrounds, both at the phenotypic and the genotypic levels. Recreation of a set of mutants in both genomic backgrounds (either wild type or lasR defective) allowed us to determine the existence of negative epistatic interactions between lasR and antibiotic resistance determinants. These epistatic interactions could lead to mutual contingency in the evolution of antibiotic resistance when P. aeruginosa colonizes a new habitat in presence of antibiotics. If lasR mutants are selected first, this would constraint antibiotic resistance evolution. Conversely, when resistance mutations (at least those studied in the present work) are selected, lasR mutants may not be selected in presence of antibiotics. These results underlie the importance of contingency and epistatic interactions in modulating antibiotic resistance evolution.

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In vitro analysis of human immunodeficiency virus type 1 resistance to nevirapine and fitness determination of resistant variants

Journal of General Virology ◽

10.1099/0022-1317-83-1-93 ◽

2002 ◽

Vol 83 (1) ◽

pp. 93-101 ◽

Cited By ~ 37

Author(s):

Maria Dolores Iglesias-Ussel ◽

Concepción Casado ◽

Eloísa Yuste ◽

Isabel Olivares ◽

Cecilio López-Galíndez

Keyword(s):

Fold Increase ◽

Resistant Mutant ◽

Wild Type ◽

Resistance Mutations ◽

Phenotypic Resistance ◽

Drug Concentrations ◽

Immunodeficiency Virus ◽

Vitro Analysis

Nevirapine-resistant variants were generated by serial passages in MT-2 cells in the presence of increasing drug concentrations. In passage 5, mutations V106A, Y181C and G190A were detected in the global population, associated with a 100-fold susceptibility decrease. Sequence analysis of biological clones obtained from passage 5 and subsequent passages showed that single mutants, detected in first passages, were progressively replaced in passage 15 by double mutants, correlating with a 500-fold increase in phenotypic resistance. Fitness determination of single mutants confirmed that, in the presence of nevirapine, every variant was more fit than wild-type with a fitness order Y181C>V106A>G190A>wild-type. Unexpectedly, in the absence of the drug, the Y181C resistant mutant was more fit than wild-type, with a fitness gradient Y181C>wild-type >G106A⩾V190A. Using a molecular clone in which the Y181C mutation was introduced by in vitro mutagenesis, the greater fitness of the Y181C mutant was confirmed in new competition cultures. These data exemplify the role of resistance mutations on virus phenotype but also on virus evolution leading, occasionally, to resistant variants fitter than the wild-type in the absence of the drug.

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Optimization of Meropenem Minimum Concentration/MIC Ratio To Suppress In Vitro Resistance of Pseudomonas aeruginosa

Antimicrobial Agents and Chemotherapy ◽

10.1128/aac.49.12.4920-4927.2005 ◽

2005 ◽

Vol 49 (12) ◽

pp. 4920-4927 ◽

Cited By ~ 113

Author(s):

Vincent H. Tam ◽

Amy N. Schilling ◽

Shadi Neshat ◽

Keith Poole ◽

David A. Melnick ◽

...

Keyword(s):

Pseudomonas Aeruginosa ◽

Infection Model ◽

Minimum Concentration ◽

Extended Duration ◽

Serial Samples ◽

The Relationship ◽

Isogenic Strains ◽

Vitro Data ◽

In Vitro Data

ABSTRACT Suppression of resistance in a dense Pseudomonas aeruginosa population has previously been shown with optimized quinolone exposures. However, the relevance to β-lactams is unknown. We investigated the bactericidal activity of meropenem and its propensity to suppress P. aeruginosa resistance in an in vitro hollow-fiber infection model (HFIM). Two isogenic strains of P. aeruginosa (wild type and an AmpC stably derepressed mutant [MIC = 1 mg/liter]) were used. An HFIM inoculated with approximately 1 × 108 CFU/ml of bacteria was subjected to various meropenem exposures. Maintenance doses were given every 8 h to simulate the maximum concentration achieved after a 1-g dose in all regimens, but escalating unbound minimum concentrations (C mi ns) were simulated with different clearances. Serial samples were obtained over 5 days to quantify the meropenem concentrations, the total bacterial population, and subpopulations with reduced susceptibilities to meropenem (>3× the MIC). For both strains, a significant bacterial burden reduction was seen with all regimens at 24 h. Regrowth was apparent after 3 days, with the C min/MIC ratio being ≤1.7 (time above the MIC, 100%). Selective amplification of subpopulations with reduced susceptibilities to meropenem was suppressed with a C min/MIC of ≥6.2 or by adding tobramycin to meropenem (C min/MIC = 1.7). Investigations that were longer than 24 h and that used high inocula may be necessary to fully evaluate the relationship between drug exposures and the likelihood of resistance suppression. These results suggest that the C min/MIC of meropenem can be optimized to suppress the emergence of non-plasmid-mediated P. aeruginosa resistance. Our in vitro data support the use of an extended duration of meropenem infusion for the treatment of severe nosocomial infections in combination with an aminoglycoside.

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Evaluation of Ceftolozane-Tazobactam in Combination with Meropenem against Pseudomonas aeruginosa Sequence Type 175 in a Hollow-Fiber Infection Model

Antimicrobial Agents and Chemotherapy ◽

10.1128/aac.00026-18 ◽

2018 ◽

Vol 62 (5) ◽

Cited By ~ 8

Author(s):

M. Montero ◽

Brian D. VanScoy ◽

Carla López-Causapé ◽

Haley Conde ◽

Jonathan Adams ◽

...

Keyword(s):

Pseudomonas Aeruginosa ◽

Hollow Fiber ◽

Sequence Type ◽

University Hospital ◽

Infection Model ◽

Content Type ◽

Clinical Exposure ◽

Density Reduction ◽

Drug Concentrations ◽

Serial Samples

ABSTRACT The aim of this study was to investigate the efficacy of ceftolozane-tazobactam in combination with meropenem against an extensively drug-resistant (XDR) Pseudomonas aeruginosa high-risk clone, sequence type 175, isolated in a Spanish university hospital. A 14-day hollow-fiber infection model was used to simulate clinical exposure of the two drug regimens alone and in combination, and serial samples were collected to determine drug concentrations and CFU counts. The untreated control failed, as did each study regimen when administered alone. However, when ceftolozane-tazobactam was administered in combination with meropenem, there was a >4-log 10 CFU/ml bacterial density reduction and suppression of resistance for the duration of the study. These data suggest that ceftolozane-tazobactam plus meropenem may be a useful combination for treating XDR P. aeruginosa .

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Faculty Opinions recommendation of Biofilm formation by Pseudomonas aeruginosa wild type, flagella and type IV pili mutants.

Faculty Opinions – Post-Publication Peer Review of the Biomedical Literature ◽

10.3410/f.1011135.192160 ◽

2003 ◽

Author(s):

Leo Eberl

Keyword(s):

Pseudomonas Aeruginosa ◽

Biofilm Formation ◽

Type Iv Pili ◽

Wild Type ◽

Type Iv

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The effects of lestaurtinib (CEP701) and PKC412 on primary AML blasts: the induction of cytotoxicity varies with dependence on FLT3 signaling in both FLT3-mutated and wild-type cases

Blood ◽

10.1182/blood-2006-04-015487 ◽

2006 ◽

Vol 108 (10) ◽

pp. 3494-3503 ◽

Cited By ~ 88

Author(s):

Steven Knapper ◽

Kenneth I. Mills ◽

Amanda F. Gilkes ◽

Steve J. Austin ◽

Val Walsh ◽

...

Keyword(s):

Myeloid Leukemia ◽

Wild Type ◽

Mutation Status ◽

Activating Mutations ◽

Drug Concentrations ◽

Flt3 Inhibitors ◽

Show Evidence ◽

Flt3 Expression ◽

Molecular Therapeutic ◽

Cytotoxic Responses

Abstract The receptor tyrosine kinase FLT3 is a promising molecular therapeutic target in acute myeloid leukemia (AML). Activating mutations of FLT3 are present in approximately one-third of patients, while many nonmutants show evidence of FLT3 activation, which appears to play a significant role in leukemogenesis. We studied the effects of lestaurtinib (CEP701) and PKC412, 2 small molecule inhibitors of FLT3, on 65 diagnostic AML blast samples. Both agents induced concentration-dependent cytotoxicity in most cases, although responses to PKC412 required higher drug concentrations. Cytotoxic responses were highly heterogeneous and were only weakly associated with FLT3 mutation status and FLT3 expression. Importantly, lestaurtinib induced cytotoxicity in a synergistic fashion with cytarabine, particularly in FLT3 mutant samples. Both lestaurtinib and PKC412 caused inhibition of FLT3 phosphorylation in all samples. Translation of FLT3 inhibition into cytotoxicity was influenced by the degree of residual FLT3 phosphorylation remaining and correlated with deactivation of STAT5 and MAP kinase. FLT3 mutant and wild-type cases both varied considerably in their dependence on FLT3 signaling for survival. These findings support the continued clinical assessment of FLT3 inhibitors in combination with cytotoxic chemotherapy: Entry to future clinical trials should include FLT3 wild-type patients and should remain unrestricted by FLT3 expression level.

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Biofilm formation by Pseudomonas aeruginosa wild type, flagella and type IV pili mutants

Molecular Microbiology ◽

10.1046/j.1365-2958.2003.03525.x ◽

2003 ◽

Vol 48 (6) ◽

pp. 1511-1524 ◽

Cited By ~ 581

Author(s):

Mikkel Klausen ◽

Arne Heydorn ◽

Paula Ragas ◽

Lotte Lambertsen ◽

Anders Aaes-Jørgensen ◽

...

Keyword(s):

Pseudomonas Aeruginosa ◽

Biofilm Formation ◽

Type Iv Pili ◽

Wild Type ◽

Type Iv

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ParB deficiency in Pseudomonas aeruginosa destabilizes the partner protein ParA and affects a variety of physiological parameters

Microbiology ◽

10.1099/mic.0.024661-0 ◽

2009 ◽

Vol 155 (4) ◽

pp. 1080-1092 ◽

Cited By ~ 28

Author(s):

A. A. Bartosik ◽

J. Mierzejewska ◽

C. M. Thomas ◽

G. Jagura-Burdzy

Keyword(s):

Pseudomonas Aeruginosa ◽

Fluorescence Microscopy ◽

Bacterial Growth ◽

Complete Loss ◽

Physiological Parameters ◽

Wild Type ◽

Partial Removal ◽

Partner Protein ◽

The One ◽

Mutant Phenotypes

Deletions leading to complete or partial removal of ParB were introduced into the Pseudomonas aeruginosa chromosome. Fluorescence microscopy of fixed cells showed that ParB mutants lacking the C-terminal domain or HTH motif formed multiple, less intense foci scattered irregularly, in contrast to the one to four ParB foci per cell symmetrically distributed in wild-type P. aeruginosa. All parB mutations affected both bacterial growth and swarming and swimming motilities, and increased the production of anucleate cells. Similar effects were observed after inactivation of parA of P. aeruginosa. As complete loss of ParA destabilized its partner ParB it was unclear deficiency of which protein is responsible for the mutant phenotypes. Analysis of four parB mutants showed that complete loss of ParB destabilized ParA whereas three mutants that retained the N-terminal 90 aa of ParB did not. As all four parB mutants demonstrate the same defects it can be concluded that either ParB, or ParA and ParB in combination, plays an important role in nucleoid distribution, growth and motility in P. aeruginosa.

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