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  • 51.
    Olofsson, Sara K
    et al.
    Uppsala University, Disciplinary Domain of Medicine and Pharmacy, Faculty of Medicine, Department of Medical Sciences.
    Marcusson, Linda L.
    Uppsala University, Disciplinary Domain of Science and Technology, Biology, Department of Cell and Molecular Biology.
    Strömbäck, Ann
    Uppsala University, Disciplinary Domain of Medicine and Pharmacy, Faculty of Medicine, Department of Medical Sciences.
    Hughes, Diarmaid
    Uppsala University, Disciplinary Domain of Science and Technology, Biology, Department of Cell and Molecular Biology.
    Cars, Otto
    Uppsala University, Disciplinary Domain of Medicine and Pharmacy, Faculty of Medicine, Department of Medical Sciences, Infectious Diseases.
    Dose-related selection of fluoroquinolone-resistant Escherichia coli2007In: Journal of Antimicrobial Chemotherapy, ISSN 0305-7453, E-ISSN 1460-2091, Vol. 60, no 4, p. 795-801Article in journal (Refereed)
    Abstract [en]

    Objectives: To investigate the effects of clinically used doses of norfloxacin,ciprofloxacin and moxifloxacin on survival and selection inEscherichia coli populations containing fluoroquinolone-resistantsubpopulations and to measure the value of the pharmacodynamicindex AUC/mutant prevention concentration (MPC) that preventsthe growth of pre-existing resistant mutants.

    Methods: Mixed cultures of susceptible wild-type and isogenic single(gyrA S83L) or double (gyrA S83L, marR) fluoroquinolone-resistantmutants were exposed to fluoroquinolones for 24 h in an in vitrokinetic model. Antibiotic concentrations modelled pharmacokineticsattained with clinical doses.

    Results: All tested doses eradicated the susceptible wild-type strain.Norfloxacin 200 mg administered twice daily selected for bothsingle and double mutants. Ciprofloxacin 250 mg administeredtwice daily eradicated the single mutant, but not the doublemutant. For that, 750 mg administered twice daily was required.Moxifloxacin 400 mg once daily eliminated the single mutant,but did not completely remove the double mutant. The MPC ofciprofloxacin was determined and based on those dose simulationsthat eradicated mutant subpopulations, an AUC/MPCwild-type of35 prevented selection of the single mutant, whereas an AUC/MPCsinglemutant of 14 (equivalent to an AUC/MPCwild-type of 105) preventedselection of the double mutant.

    Conclusions: All tested clinical dosing regimens were effective in eradicatingsusceptible bacteria, but ciprofloxacin 750 mg twice daily wasthe only dose that prevented the selection of single- and double-resistantE. coli mutants. Thus, among approved fluoroquinolone dosingregimens, some are significantly more effective than othersin exceeding the mutant selection window and preventing theenrichment of resistant mutants.

  • 52.
    Papareddy, Praveen
    et al.
    Lund Univ, Dept Clin Sci, Div Dermatol & Venereol, Lund, Sweden.;Lund Univ, Dept Clin Sci, Div Infect Med, Lund, Sweden..
    Kasetty, Gopinath
    Lund Univ, Dept Clin Sci, Div Dermatol & Venereol, Lund, Sweden.;Lund Univ, Dept Clin Sci, Respiratory Med & Allergol, Lund, Sweden..
    Kalle, Martina
    Lund Univ, Dept Clin Sci, Div Dermatol & Venereol, Lund, Sweden..
    Bhongir, Ravi K. V.
    Lund Univ, Dept Clin Sci, Div Dermatol & Venereol, Lund, Sweden.;Lund Univ, Dept Clin Sci, Respiratory Med & Allergol, Lund, Sweden..
    Morgelin, Matthias
    Lund Univ, Dept Clin Sci, Div Infect Med, Lund, Sweden..
    Schmidtchen, Artur
    Lund Univ, Dept Clin Sci, Div Dermatol & Venereol, Lund, Sweden.;Nanyang Technol Univ, LKCMed, Dermatol, Singapore 639798, Singapore..
    Malmsten, Martin
    Uppsala University, Disciplinary Domain of Medicine and Pharmacy, Faculty of Pharmacy, Department of Pharmacy.
    NLF20: an antimicrobial peptide with therapeutic potential against invasive Pseudomonas aeruginosa infection2016In: Journal of Antimicrobial Chemotherapy, ISSN 0305-7453, E-ISSN 1460-2091, Vol. 71, no 1, p. 170-180Article in journal (Refereed)
    Abstract [en]

    Objectives:Increasing resistance to antibiotics makes antimicrobial peptides interesting as novel therapeutics. Here, we report on studies of the peptide NLF20 (NLFRKLTHRLFRRNFGYTLR), corresponding to an epitope of the D helix of heparin cofactor II (HCII), a plasma protein mediating bacterial clearance. Methods: Peptide effects were evaluated by a combination of in vitro and in vivo methods, including antibacterial, anti-inflammatory and cytotoxicity assays, fluorescence and electron microscopy, and experimental models of endotoxin shock and Pseudomonas aeruginosa sepsis. Results: The results showed that NLF20 displayed potent antimicrobial effects against the Gram-negative bacteria Escherichia coli and P. aeruginosa, the Gram-positive Bacillus subtilis and Staphylococcus aureus and the fungi Candida albicans and Candida parapsilosis. Importantly, this antimicrobial effect was retained in human blood, particularly for P. aeruginosa. Fluorescence and electron microscopy studies showed that the peptide exerted membrane-breaking effects. In an animal model of P. aeruginosa sepsis, NLF20 reduced bacterial levels, resulting in improved survival. Reduced mortality was also observed in experimental animal models of endotoxin shock, which was paralleled with modulated IFN-gamma, IL-10 and coagulation responses. Conclusions: Together, these results indicate that functional epitopes of HCII may have therapeutic potential against bacterial infection.

  • 53.
    Pietsch, Franziska
    et al.
    Uppsala University, Disciplinary Domain of Medicine and Pharmacy, Faculty of Medicine, Department of Medical Biochemistry and Microbiology.
    Bergman, Jessica M.
    Uppsala University, Disciplinary Domain of Medicine and Pharmacy, Faculty of Medicine, Department of Medical Biochemistry and Microbiology.
    Brandis, Gerrit
    Uppsala University, Disciplinary Domain of Medicine and Pharmacy, Faculty of Medicine, Department of Medical Biochemistry and Microbiology.
    Marcusson, Linda L.
    Uppsala University, Disciplinary Domain of Medicine and Pharmacy, Faculty of Medicine, Department of Medical Biochemistry and Microbiology.
    Zorzet, Anna
    Uppsala University, Disciplinary Domain of Medicine and Pharmacy, Faculty of Medicine, Department of Medical Biochemistry and Microbiology.
    Huseby, Douglas L.
    Uppsala University, Disciplinary Domain of Medicine and Pharmacy, Faculty of Medicine, Department of Medical Biochemistry and Microbiology.
    Hughes, Diarmaid
    Uppsala University, Disciplinary Domain of Medicine and Pharmacy, Faculty of Medicine, Department of Medical Biochemistry and Microbiology.
    Ciprofloxacin selects for RNA polymerase mutations with pleiotropic antibiotic resistance effects2017In: Journal of Antimicrobial Chemotherapy, ISSN 0305-7453, E-ISSN 1460-2091, Vol. 72, no 1, p. 75-84Article in journal (Refereed)
    Abstract [en]

    Objectives: Resistance to the fluoroquinolone drug ciprofloxacin is commonly linked to mutations that alter the drug target or increase drug efflux via the major AcrAB-TolC transporter. Very little is known about other mutations that might also reduce susceptibility to ciprofloxacin. We discovered that an Escherichia coli strain experimentally evolved for resistance to ciprofloxacin had acquired a mutation in rpoB, the gene coding for the beta-subunit of RNA polymerase. The aim of this work was to determine whether this mutation, and other mutations in rpoB, contribute to ciprofloxacin resistance and, if so, by which mechanism. Methods: Independent lineages of E. coli were evolved in the presence of ciprofloxacin and clones from endpoint cultures were screened for mutations in rpoB. Ciprofloxacin-selected rpoB mutations were identified and characterized in terms of effects on susceptibility and mode of action. Results: Mutations in rpoB were selected at a high frequency in 3 out of 10 evolved lineages, in each case arising after the occurrence of mutations affecting topoisomerases and drug efflux. All ciprofloxacin-selected rpoB mutations had a high fitness cost in the absence of drug, but conferred a competitive advantage in the presence of ciprofloxacin. RNA sequencing and quantitative RT-PCR analysis showed that expression of mdtK, encoding a multidrug efflux transporter, was significantly increased by the ciprofloxacin-selected rpoB mutations. The susceptibility phenotype was shown to depend on the presence of an active mdtK and a mutant rpoB allele. Conclusions: These data identify mutations in RNA polymerase as novel contributors to the evolution of resistance to ciprofloxacin and show that the phenotype is mediated by increased MdtK-dependent drug efflux.

  • 54. Poulsen, Hanna O.
    et al.
    Johansson, Anders
    Granholm, Susanne
    Kahlmeter, Gunnar
    Uppsala University, Disciplinary Domain of Medicine and Pharmacy, Faculty of Medicine, Department of Medical Sciences, Clinical Bacteriology.
    Sundqvist, Martin
    Uppsala University, Disciplinary Domain of Medicine and Pharmacy, Faculty of Medicine, Department of Medical Sciences, Infectious Diseases.
    High genetic diversity of nitrofurantoin- or mecillinam-resistant Escherichia coli indicates low propensity for clonal spread2013In: Journal of Antimicrobial Chemotherapy, ISSN 0305-7453, E-ISSN 1460-2091, Vol. 68, no 9, p. 1974-1977Article in journal (Refereed)
    Abstract [en]

    The empirical treatment with trimethoprim or ciprofloxacin of urinary tract infections (UTIs) is now questioned, partly due to the global expansion of a few resistant clonal groups of Escherichia coli. In this study we investigated the clonal structure of 34 strains of E. coli (collected from non-pregnant women aged 1865 years with uncomplicated UTIs in Europe and Canada) resistant to either of two other common treatment alternatives for uncomplicated UTIs, mecillinam or nitrofurantoin, using multilocus sequence typing (MLST). The 34 isolates were, despite high levels of multiresistance, distributed all over the E. coli genetic diversity spectrum with little association of antibiotic resistance to specific clonal groups. The results of this study indicate a low probability of a future clonal spread of resistance to mecillinam and nitrofurantoin.

  • 55.
    Praski Alzrigat, Lisa
    et al.
    Uppsala University, Disciplinary Domain of Medicine and Pharmacy, Faculty of Medicine, Department of Medical Biochemistry and Microbiology.
    Huseby, Douglas L
    Uppsala University, Disciplinary Domain of Medicine and Pharmacy, Faculty of Medicine, Department of Medical Biochemistry and Microbiology.
    Brandis, Gerrit
    Uppsala University, Disciplinary Domain of Medicine and Pharmacy, Faculty of Medicine, Department of Medical Biochemistry and Microbiology.
    Hughes, Diarmaid
    Uppsala University, Disciplinary Domain of Medicine and Pharmacy, Faculty of Medicine, Department of Medical Biochemistry and Microbiology.
    Fitness cost constrains the spectrum of marR mutations in ciprofloxacin-resistant Escherichia coli: Multiple Antibiotic-Resistance, Gram-Negative Bacteria, Multidrug Efflux Pump, Urinary-Tract-Infections, Fluoroquinolone Resistance, Quinolone Resistance, Mechanisms, Expression, Sequence, Soxs2017In: Journal of Antimicrobial Chemotherapy, ISSN 0305-7453, E-ISSN 1460-2091, Vol. 2, no 11, p. 3016-3024Article in journal (Refereed)
    Abstract [en]

    Objectives: To determine whether the spectrum of mutations in marR in ciprofloxacin-resistant clinical isolates of Escherichia coli shows evidence of selection bias, either to reduce fitness costs, or to increase drug resistance. MarR is a repressor protein that regulates, via MarA, expression of the Mar regulon, including the multidrug efflux pump AcrAB-TolC. Methods: Isogenic strains carrying 36 different marR alleles identified in resistant clinical isolates, or selected for resistance in vitro, were constructed. Drug susceptibility and relative fitness in growth competition assays were measured for all strains. The expression level of marA, and of various efflux pump components, as a function of specific mutations in marR, was measured by qPCR. Results: The spectrum of genetic alterations in marR in clinical isolates is strongly biased against inactivating mutations. In general, the alleles found in clinical isolates conferred a lower level of resistance and imposed a lower growth fitness cost than mutations selected in vitro. The level of expression of MarA correlated well with the MIC of ciprofloxacin. This supports the functional connection between mutations in marR and reduced susceptibility to ciprofloxacin. Conclusions: Mutations in marR selected in ciprofloxacin-resistant clinical isolates are strongly biased against inactivating mutations. Selection favours mutant alleles that have the lowest fitness costs, even though these cause only modest reductions in drug susceptibility. This suggests that selection for high relative fitness is more important than selection for increased resistance in determining which alleles of marR will be selected in resistant clinical isolates.

  • 56.
    Pränting, Maria
    et al.
    Uppsala University, Disciplinary Domain of Medicine and Pharmacy, Faculty of Medicine, Department of Medical Biochemistry and Microbiology.
    Andersson, Dan I
    Uppsala University, Disciplinary Domain of Medicine and Pharmacy, Faculty of Medicine, Department of Medical Biochemistry and Microbiology.
    Mechanisms and physiological effects of protamine resistance in Salmonella enterica serovar Typhimurium LT22010In: Journal of Antimicrobial Chemotherapy, ISSN 0305-7453, E-ISSN 1460-2091, Vol. 65, no 5, p. 876-887Article in journal (Refereed)
    Abstract [en]

    OBJECTIVES: Protamines are cationic peptides that exert antimicrobial activity. We have examined the evolution of bacterial resistance to protamine sulphate and the resulting effects on fitness and physiology, with the objective of increasing knowledge about mechanisms of bacterial resistance to antimicrobial peptides. METHODS: Spontaneous, protamine-resistant Salmonella enterica serovar Typhimurium (i.e. Salmonella Typhimurium) LT2 mutants were isolated on agarose plates containing protamine sulphate. Resistance mutations were identified using transposon insertions and DNA sequencing. Peptide susceptibility was determined by broth dilution tests and antibiotic susceptibility using Etests. Fitness was determined as log-phase growth rates. Growth-compensated strains were isolated by serial passage through population bottlenecks followed by visual screening for large colonies. RESULTS: Protamine-resistant mutants appeared at a rate of 2.3 x 10(-7)/cell/generation. These mutants were 2-20 times more resistant to protamine than the parental strain and less susceptible to several other antimicrobials, including colistin, gentamicin, lactoferricin and human defensin HNP-1. The resistance mutations were mapped to genes involved in haem biosynthesis and respiration, and were associated with a reduction in bacterial fitness. Some mutants could, in the absence of protamine, be evolved to higher fitness by acquiring second-site compensatory mutations. CONCLUSIONS: Spontaneous mutants resistant to protamine sulphate were readily selected in Salmonella Typhimurium LT2. These mutants were less susceptible to several other peptides and antibiotics, and had the characteristics of small colony variants, a phenotype often associated with persistent and recurrent infections that are difficult to treat and which could be a strategy for bacteria to escape the killing effects of antimicrobial peptides.

  • 57.
    Pränting, Maria
    et al.
    Uppsala University, Disciplinary Domain of Medicine and Pharmacy, Faculty of Medicine, Department of Medical Biochemistry and Microbiology.
    Lööv, Camilla
    Uppsala University, Disciplinary Domain of Medicine and Pharmacy, Faculty of Medicine, Department of Medical Biochemistry and Microbiology.
    Burman, Robert
    Uppsala University, Disciplinary Domain of Medicine and Pharmacy, Faculty of Pharmacy, Department of Medicinal Chemistry, Division of Pharmacognosy.
    Göransson, Ulf
    Uppsala University, Disciplinary Domain of Medicine and Pharmacy, Faculty of Pharmacy, Department of Medicinal Chemistry, Division of Pharmacognosy.
    Andersson, Dan I
    Uppsala University, Disciplinary Domain of Medicine and Pharmacy, Faculty of Medicine, Department of Medical Biochemistry and Microbiology.
    The cyclotide cycloviolacin O2 from Viola odorata has potent bactericidal activity against Gram-negative bacteria2010In: Journal of Antimicrobial Chemotherapy, ISSN 0305-7453, E-ISSN 1460-2091, Vol. 65, no 9, p. 1964-1971Article in journal (Refereed)
    Abstract [en]

    OBJECTIVES:

    To determine the antibacterial activity of small cyclic plant proteins, i.e. cyclotides, and the importance of the surface exposed charged residues for activity.

    METHODS:

    Prototypic cyclotides, including the Möbius kalata B1 and the bracelet cycloviolacin O2 (cyO2), were isolated using reversed-phase HPLC. Initial activity screenings were conducted using radial diffusion assays (RDAs) and MIC assays with Salmonella enterica serovar Typhimurium LT2, Escherichia coli and Staphylococcus aureus as test strains. For the most active peptide, cyO2, time-kill kinetics was determined in sodium phosphate buffer (containing 0.03% trypticase soy broth) against several Gram-negative and Gram-positive bacterial species. Charged residues in cyO2 were chemically modified and activity was determined in time-kill assays.

    RESULTS:

    CyO2 was the most active cyclotide and efficiently inhibited the growth of S. enterica serovar Typhimurium LT2 and E. coli in RDAs and MIC assays, while the other peptides were less active. In time-kill assays, cyO2 also had bactericidal activity against the Gram-negative species Klebsiella pneumoniae and Pseudomonas aeruginosa. In contrast, none of the cyclotides had high activity against S. aureus. Chemical masking of the charged Glu and Lys residues in cyO2 caused a near total loss of activity against Salmonella, while masking Arg caused a less pronounced activity reduction.

    CONCLUSIONS:

    CyO2 is a cyclotide with potent activity against Gram-negative bacteria. The charged residues in cyO2 are all required for optimum antibacterial activity. In combination with its previously demonstrated cytotoxic activity against cancer cells and the general stability of cyclotides, cyO2 provides a promising scaffold for future drug design.

  • 58. Puértolas-Balint, Fabiola
    et al.
    Warsi, Omar
    Linkevicius, Marius
    Tang, Po-Cheng
    Andersson, Dan I
    Mutations that increase expression of the EmrAB-TolC efflux pump confer increased resistance to nitroxoline in Escherichia coli.2019In: Journal of Antimicrobial Chemotherapy, ISSN 0305-7453, E-ISSN 1460-2091, article id dkz434Article in journal (Refereed)
    Abstract [en]

    OBJECTIVES: To determine the mechanism of resistance to the antibiotic nitroxoline in Escherichia coli.

    METHODS: Spontaneous nitroxoline-resistant mutants were selected at different concentrations of nitroxoline. WGS and strain reconstruction were used to define the genetic basis for the resistance. The mechanistic basis of resistance was determined by quantitative PCR (qPCR) and by overexpression of target genes. Fitness costs of the resistance mutations and cross-resistance to other antibiotics were also determined.

    RESULTS: Mutations in the transcriptional repressor emrR conferred low-level resistance to nitroxoline [nitroxoline MIC (MICNOX) = 16 mg/L] by increasing the expression of the emrA and emrB genes of the EmrAB-TolC efflux pump. These resistant mutants showed no fitness reduction and displayed cross-resistance to nalidixic acid. Second-step mutants with higher-level resistance (MICNOX = 32-64 mg/L) had mutations in the emrR gene, together with either a 50 kb amplification, a mutation in the gene marA, or an IS upstream of the lon gene. The latter mutations resulted in higher-level nitroxoline resistance due to increased expression of the tolC gene, which was confirmed by overexpressing tolC from an inducible plasmid in a low-level resistance mutant. Furthermore, the emrR mutations conferred a small increase in resistance to nitrofurantoin only when combined with an nfsAB double-knockout mutation. However, nitrofurantoin-resistant nfsAB mutants showed no cross-resistance to nitroxoline.

    CONCLUSIONS: Mutations in different genes causing increased expression of the EmrAB-TolC pump lead to an increased resistance to nitroxoline. The structurally similar antibiotics nitroxoline and nitrofurantoin appear to have different modes of action and resistance mechanisms.

  • 59.
    Ruth, Mike Marvin
    et al.
    Radboud Univ Nijmegen, Dept Med Microbiol, Med Ctr, Nijmegen, Netherlands.
    Sangen, Jasper J. N.
    Radboud Univ Nijmegen, Dept Med Microbiol, Med Ctr, Nijmegen, Netherlands.
    Remmers, Karlijn
    Radboud Univ Nijmegen, Dept Med Microbiol, Med Ctr, Nijmegen, Netherlands.
    Pennings, Lian J.
    Radboud Univ Nijmegen, Dept Med Microbiol, Med Ctr, Nijmegen, Netherlands.
    Svensson, Elin
    Uppsala University, Disciplinary Domain of Medicine and Pharmacy, Faculty of Pharmacy, Department of Pharmaceutical Biosciences. Radboud Univ Nijmegen, Dept Pharm, Med Ctr, Nijmegen, Netherlands.
    Aarnoutse, Rob E.
    Radboud Univ Nijmegen, Dept Pharm, Med Ctr, Nijmegen, Netherlands.
    Zweijpfenning, Sanne M. H.
    Radboud Univ Nijmegen, Dept Pulm Dis, Med Ctr, Nijmegen, Netherlands.
    Hoefsloot, Wouter
    Radboud Univ Nijmegen, Dept Pulm Dis, Med Ctr, Nijmegen, Netherlands.
    Kuipers, Saskia
    Radboud Univ Nijmegen, Dept Med Microbiol, Med Ctr, Nijmegen, Netherlands.
    Magis-Escurra, Cecile
    Radboud Univ Nijmegen, Dept Pulm Dis, Med Ctr, Nijmegen, Netherlands.
    Wertheim, Heiman F. L.
    Radboud Univ Nijmegen, Dept Med Microbiol, Med Ctr, Nijmegen, Netherlands.
    van Ingen, Jakko
    Radboud Univ Nijmegen, Dept Med Microbiol, Med Ctr, Nijmegen, Netherlands.
    A bedaquiline/clofazimine combination regimen might add activity to the treatment of clinically relevant non-tuberculous mycobacteria2019In: Journal of Antimicrobial Chemotherapy, ISSN 0305-7453, E-ISSN 1460-2091, Vol. 74, no 4, p. 935-943Article in journal (Refereed)
    Abstract [en]

    Background: Non-tuberculous mycobacteria (NTM) infections are hard to treat. New antimicrobial drugs and smarter combination regimens are needed.

    Objectives: Our aim was to determine the in vitro activity of bedaquiline against NTM and assess its synergy with established antimycobacterials.

    Methods: We determined MICs of bedaquiline for clinically relevant NTM species and Mycobacterium tuberculosis by broth microdilution for 30 isolates. Synergy testing was performed using the chequerboard method for 22 reference strains and clinical isolates of Mycobacterium abscessus (MAB) and Mycobacterium avium complex (MAC). Time-kill kinetics (TK) assays with resistance monitoring of bedaquiline alone and combined with clofazimine were performed for MAB CIP 104536 and M. avium ATCC 700898; bedaquiline/clarithromycin combinations were evaluated against M. avium ATCC 700898. Interactions were assessed for TK experiments based on Bliss independence.

    Results: Bedaquiline had modest activity against tested NTM, with MICs between <0.007and 1mg/L. Bedaquiline showed no interaction with tested drugs against MAB or MAC. Lowest mean fractional inhibitory concentration index (FICI) values were 0.79 with clofazimine for MAB and 0.97 with clofazimine and 0.82 with clarithromycin for MAC. In TK assays, bedaquiline showed a bacteriostatic effect. Clofazimine extended the bacteriostatic activity of bedaquiline against MAB and yielded a slight bactericidal effect against M. avium. The bedaquiline/clofazimine combination slowed emergence of bedaquiline resistance for M. avium but promoted it for MAB. Relative to Bliss independence, bedaquiline/clofazimine showed synergistic interaction over time for MAB and no interaction for M. avium and bedaquiline/clarithromycin showed antagonistic interaction for M. avium.

    Conclusions: Following these in vitro data, a bedaquiline/clofazimine combination might add activity to MAB and MAC treatment. The bedaquiline/clarithromycin combination might have lower activity compared with bedaquiline alone for MAC treatment.

  • 60.
    Sandegren, Linus
    et al.
    Uppsala University, Disciplinary Domain of Medicine and Pharmacy, Faculty of Medicine, Department of Medical Biochemistry and Microbiology.
    Lindqvist, Anton
    Uppsala University, Disciplinary Domain of Medicine and Pharmacy, Faculty of Medicine, Department of Medical Biochemistry and Microbiology.
    Kahlmeter, Gunnar
    Uppsala University, Disciplinary Domain of Medicine and Pharmacy, Faculty of Medicine, Department of Medical Sciences.
    Andersson, Dan I
    Uppsala University, Disciplinary Domain of Medicine and Pharmacy, Faculty of Medicine, Department of Medical Biochemistry and Microbiology.
    Nitrofurantoin resistance mechanism and fitness cost in Escherichia coli2008In: Journal of Antimicrobial Chemotherapy, ISSN 0305-7453, E-ISSN 1460-2091, Vol. 62, no 3, p. 495-503Article in journal (Refereed)
    Abstract [en]

    OBJECTIVES

    The biological fitness cost of antibiotic resistance is a key parameter in determining the rate of appearance and spread of antibiotic-resistant bacteria. We identified mutations conferring nitrofurantoin resistance and examined their effect on the fitness of clinical Escherichia coli isolates.

    METHODS

    By plating bacterial cells on agar plates containing nitrofurantoin, spontaneous nitrofurantoin-resistant E. coli mutants were isolated. The fitness of susceptible and resistant strains was measured as growth rate in the presence and absence of nitrofurantoin in rich culture medium. Time-kill kinetics of the resistant mutants was compared with the susceptible strains. Resistance mutations were identified by DNA sequencing.

    RESULTS

    Spontaneous resistant mutants of initially susceptible clinical E. coli appeared with a rate of 10(-7)/cell/generation, and these mutants showed a reduction in the growth rate compared with the susceptible parent strain. Similarly, comparison of a set of susceptible and resistant clinical isolates of E. coli showed that the average growth rate of the resistant mutants was approximately 6% lower than the susceptible strains. Furthermore, the bacterial growth rate in the presence of nitrofurantoin at therapeutic levels was greatly reduced even for nitrofurantoin-resistant mutants. The resistance-conferring mutations were identified in the nsfA and nfsB genes that encode oxygen-insensitive nitroreductases.

    CONCLUSIONS

    Nitrofurantoin resistance confers a reduction in fitness in E. coli in the absence of antibiotic. In the presence of therapeutic levels of nitrofurantoin, even resistant mutants are so disturbed in growth that they are probably unable to become enriched and establish an infection.

  • 61.
    Sandegren, Linus
    et al.
    Uppsala University, Disciplinary Domain of Medicine and Pharmacy, Faculty of Medicine, Department of Medical Biochemistry and Microbiology.
    Linkevicius, Marius
    Uppsala University, Disciplinary Domain of Medicine and Pharmacy, Faculty of Medicine, Department of Medical Biochemistry and Microbiology.
    Lytsy, Birgitta
    Uppsala University, Disciplinary Domain of Medicine and Pharmacy, Faculty of Medicine, Department of Medical Sciences, Clinical Bacteriology.
    Åsa, Melhus
    Uppsala University, Disciplinary Domain of Medicine and Pharmacy, Faculty of Medicine, Department of Medical Sciences, Clinical Bacteriology.
    Andersson, Dan I.
    Uppsala University, Disciplinary Domain of Medicine and Pharmacy, Faculty of Medicine, Department of Medical Biochemistry and Microbiology.
    Transfer of an Escherichia coli ST131 multiresistance cassette has created a Klebsiella pneumoniae-specific plasmid associated with a major nosocomial outbreak2012In: Journal of Antimicrobial Chemotherapy, ISSN 0305-7453, E-ISSN 1460-2091, Vol. 67, no 1, p. 74-83Article in journal (Refereed)
    Abstract [en]

    Objectives:

    To characterize the complete sequence, horizontal spread and stability of the CTX-M-15-encoding multiresistance plasmid of a Klebsiella pneumoniae strain involved in a large nosocomial outbreak.

    Methods:

    The 220 kbp plasmid pUUH239.2 was completely sequenced using 454 technology. The conjugational host range, conjugation frequencies, plasmid stability and fitness cost of plasmid carriage were studied in vitro. Conjugational spread during the outbreak was assessed retrospectively by multiplex PCR screening, restriction fragment length polymorphism and PFGE.

    Results:

    Plasmid pUUH239.2 encodes resistance to β-lactams (blaCTX-M-15, blaTEM-1 and blaOXA-1), aminoglycosides [aac-(6′)-1b-cr and aadA2], tetracyclines [tet(A) and tetR], trimethoprim (dhfrXII), sulphonamides (sul1) quaternary ammonium compounds (qacEΔ1), macrolides [mph(A)-mxr-mphR(A)] and heavy metal ions (silver, copper and arsenic). The plasmid consists of a backbone, highly similar to the K. pneumoniae plasmid pKPN3, and a 41 kbp resistance region, highly similar to the resistance regions of plasmids pEK499 and pC15-1a previously isolated from Escherichia coli strains belonging to the outbreak lineage ST131 (where ST stands for sequence type). The pUUH239.2 plasmid is stable in K. pneumoniae but unstable in E. coli and confers a fitness cost when introduced into a naive host cell. Transfer of pUUH239.2 from the outbreak K. pneumoniae clone to the E. coli of the patients’ intestinal floras has occurred on multiple occasions during the outbreak.

    Conclusions:

    The plasmid pUUH239.2 is a composite of the pKPN3 K. pneumoniae plasmid backbone and the blaCTX-M-15-encoding multiresistance cassette associated with the internationally recognized outbreak strain E. coli ST131. The resulting plasmid differs in stability between K. pneumoniae and E. coli, and this has probably limited the spread of this plasmid during the outbreak.

  • 62.
    Schalkwijk, Stein
    et al.
    Radboud Univ Nijmegen, RIHS, Dept Pharm, Med Ctr, Nijmegen, Netherlands;Radboud Univ Nijmegen, Dept Pharmacol & Toxicol, RIMLS, Med Ctr, Nijmegen, Netherlands.
    ter Heine, Rob
    Radboud Univ Nijmegen, RIHS, Dept Pharm, Med Ctr, Nijmegen, Netherlands.
    Colbers, Angela
    Radboud Univ Nijmegen, RIHS, Dept Pharm, Med Ctr, Nijmegen, Netherlands.
    Capparelli, Edmund
    Univ Calif San Diego, Skaggs Sch Pharm & Pharmaceut Sci, San Diego, CA 92103 USA;Univ Calif San Diego, Sch Med, San Diego, CA 92103 USA.
    Best, Brookie M.
    Univ Calif San Diego, Skaggs Sch Pharm & Pharmaceut Sci, San Diego, CA 92103 USA;Univ Calif San Diego, Sch Med, San Diego, CA 92103 USA.
    Cressey, Tim R.
    Chiang Mai Univ, Fac Associated Med Sci, Chiang Mai, Thailand;Harvard TH Chan Sch Publ Hlth, Boston, MA USA;Univ Liverpool, Dept Mol & Clin Pharmacol, Liverpool, Merseyside, England.
    Greupink, Rick
    Radboud Univ Nijmegen, Dept Pharmacol & Toxicol, RIMLS, Med Ctr, Nijmegen, Netherlands.
    Russel, Frans G. M.
    Radboud Univ Nijmegen, Dept Pharmacol & Toxicol, RIMLS, Med Ctr, Nijmegen, Netherlands.
    Molto, Jose
    Hosp Badalona Germans Trias & Pujol, Fundacio Lluita Sida, Badalona, Spain;Hosp Badalona Germans Trias & Pujol, Infect Dis Dept, Badalona, Spain;Univ Autonoma Barcelona, Barcelona, Spain.
    Mirochnick, Mark
    Boston Univ, Sch Med, Dept Pediat, Boston, MA 02118 USA.
    Karlsson, Mats
    Uppsala University, Disciplinary Domain of Medicine and Pharmacy, Faculty of Pharmacy, Department of Pharmaceutical Biosciences.
    Burger, David M.
    Radboud Univ Nijmegen, RIHS, Dept Pharm, Med Ctr, Nijmegen, Netherlands.
    Evaluating darunavir/ritonavir dosing regimens for HIV-positive pregnant women using semi-mechanistic pharmacokinetic modelling2019In: Journal of Antimicrobial Chemotherapy, ISSN 0305-7453, E-ISSN 1460-2091, Vol. 74, no 5, p. 1348-1356Article in journal (Refereed)
    Abstract [en]

    Background: Darunavir 800mg once (q24h) or 600 mg twice (q12h) daily combined with low-dose ritonavir is used to treat HIV-positive pregnant women. Decreased total darunavir exposure (17%-50%) has been reported during pregnancy, but limited data on unbound exposure are available. Objectives: To evaluate total and unbound darunavir exposures following standard darunavir/ritonavir dosing and to explore the value of potential optimized darunavir/ritonavir dosing regimens for HIV-positive pregnant women. Patients and methods: A population pharmacokinetic analysis was conducted based on data from 85 women. The final model was used to simulate total and unbound darunavir AUC(0-tau) and C-trough during the third trimester of pregnancy, as well as to assess the probability of therapeutic exposure. Results: Simulations predicted that total darunavir exposure (AUC(0-tau)) was 24% and 23% lower in pregnancy for standard q24h and q12h dosing, respectively. Unbound darunavir AUC(0-tau) was 5% and 8% lower compared with post-partum for standard q24h and q12h dosing, respectively. The probability of therapeutic exposure (unbound) during pregnancy was higher for standard q12h dosing (99%) than for q24h dosing (94%). Conclusions: The standard q12h regimen resulted in maximal and higher rates of therapeutic exposure compared with standard q24h dosing. Darunavir/ritonavir 600/100 mg q12h should therefore be the preferred regimen during pregnancy unless (adherence) issues dictate q24h dosing. The value of alternative dosing regimens seems limited.

  • 63. Schon, T.
    et al.
    Jureen, P.
    Chryssanthou, E.
    Giske, C. G.
    Kahlmeter, Gunnar
    Uppsala University, Disciplinary Domain of Medicine and Pharmacy, Faculty of Medicine, Department of Medical Sciences, Clinical Bacteriology.
    Hoffner, S.
    Angeby, K.
    Rifampicin-resistant and rifabutin-susceptible Mycobacterium tuberculosis strains: a breakpoint artefact?2013In: Journal of Antimicrobial Chemotherapy, ISSN 0305-7453, E-ISSN 1460-2091, Vol. 68, no 9, p. 2074-2077Article in journal (Refereed)
    Abstract [en]

    It has long been assumed that some rifampicin-resistant Mycobacterium tuberculosis strains are susceptible to, and thus treatable with, rifabutin. However, clinical breakpoints for susceptibility testing of rifabutin as well as the evidence for a clinical effect of rifabutin in rifampicin-resistant strains remains poorly defined. The objective of this study was to re-evaluate the breakpoint for rifabutin in relation to its MIC wild-type distribution and the presence of mutations in rpoB. The MIC in 7H10 Middlebrook medium was determined for clinical isolates of M. tuberculosis (n95), where a majority were multidrug resistant. Additionally, all strains were screened for rpoB mutations by sequencing and the GenoType MTBDRplus assay. Rifampicin resistance was confirmed by genotypical and/or phenotypical tests in 73 isolates (76.8). Nineteen isolates, defined as rifampicin resistant and rifabutin susceptible according to the present breakpoint, exhibited significantly higher MICs of rifabutin (0.0640.5 mg/L) than rifabutin-susceptible isolates without any detectable mutations in rpoB (P0.001). These 19 isolates were clearly resistant to rifampicin (MIC 2256 mg/L) and all but one had mutations in rpoB, with 9 (47.4) specifically in Asp516Val. Our results indicate that rifampicin-resistant but rifabutin-susceptible isolates according to the present breakpoints harbour rpoB mutations and have a rifabutin MIC significantly higher than strains without any detectable mutations in rpoB. So far there are no clinical, pharmacological or microbiological data to confirm that such isolates can be treated with rifabutin and we suggest a revision of the current breakpoints.

  • 64. Schon, Thomas
    et al.
    Jureen, Pontus
    Giske, Christian G.
    Chryssanthou, Erja
    Sturegard, Erik
    Werngren, Jim
    Kahlmeter, Gunnar
    Uppsala University, Disciplinary Domain of Medicine and Pharmacy, Faculty of Medicine, Department of Medical Sciences.
    Hoffner, Sven E.
    Angeby, Kristian A.
    Evaluation of wild-type MIC distributions as a tool for determination of clinical breakpoints for Mycobacterium tuberculosis2009In: Journal of Antimicrobial Chemotherapy, ISSN 0305-7453, E-ISSN 1460-2091, Vol. 64, no 4, p. 786-793Article in journal (Refereed)
    Abstract [en]

    Objectives: The aim of this study was to establish wild-type MIC distributions of first-line drugs for Mycobacterium tuberculosis, as well as to explore the usefulness of such distributions when setting clinical breakpoints. Methods: We determined the MICs of rifampicin, isonlazid and ethambutol for M. tuberculosis using a Middlebrook 7H10 dilution method for 90 consecutive clinical isolates, 8 resistant strains and 16 isolates from the WHO proficiency test panel. M. tuberculosis H37Rv was used for quality control and susceptibility results using 7H10 were compared with the results obtained with BACTEC460. Results: The agreement with BACTEC460 was very high for isonlazid (99.1%) and rifampicin (99.1%) but lower for ethambutol (94.7%). Intra- and inter-assay variation was below one MIC dilution. The MIC distributions for isoniazid and rifampicin provided a clear separation between susceptible and resistant strains. Regarding ethambutol, the current breakpoint for 7H10 (5 mg/L) is close to the wild-type and all strains (n=6) showing a disagreement between BACTEC460 and 7H10 were distributed very close to the breakpoint (MIC 4-8 mg/L). This problematic relation was confirmed by investigating isolates from the WHO panel with an agreement <95% (64%-88% among 26 laboratories, n=4) for which the MICs were 4-8 mg/L . Conclusions: Utilizing the wild-type MIC distribution was found to be as useful in M. tuberculosis as in other bacteria when setting clinical breakpoints. We suggest that the present clinical breakpoints should be re-evaluated, taking into account wild-type MIC distributions and available pharmacokinetic data.

  • 65. Silva, M
    et al.
    Ferreira, P E
    Otienoburu, S D
    Calçada, C
    Ngasala, Billy
    Uppsala University, Disciplinary Domain of Medicine and Pharmacy, Faculty of Medicine, Department of Women's and Children's Health, International Maternal and Child Health (IMCH), International Child Health and Nutrition. Muhimbili University of Health and Allied Sciences, Dar es Salaam, Tanzania.
    Björkman, A
    Mårtensson, Andreas
    Uppsala University, Disciplinary Domain of Medicine and Pharmacy, Faculty of Medicine, Department of Women's and Children's Health, International Maternal and Child Health (IMCH), International Child Health and Nutrition.
    Gil, José Pedro
    Uppsala University, Disciplinary Domain of Medicine and Pharmacy, Faculty of Medicine, Department of Women's and Children's Health, International Maternal and Child Health (IMCH), International Child Health and Nutrition. Division of Pharmacogenetics, Department of Physiology and Pharmacology, Karolinska Institutet, Stockholm, Sweden;Center for Biodiversity, Functional & Integrative Genomics, Faculdade de Ciências, Universidade de Lisboa, Lisbon, Portugal.
    Veiga, M I
    Plasmodium falciparum K13 expression associated with parasite clearance during artemisinin-based combination therapy2019In: Journal of Antimicrobial Chemotherapy, ISSN 0305-7453, E-ISSN 1460-2091, Vol. 74, no 7, p. 1890-1893Article in journal (Refereed)
    Abstract [en]

    BACKGROUND: Delayed parasite clearance and, consequently, reduced efficacy of artemisinin-based combination therapies have been linked with Plasmodium falciparum K13 gene SNPs in Southeast Asia. In Africa, significantly prolonged clearance has not yet been observed and the presently restricted variation in parasite clearance cannot be explained by K13 polymorphisms.

    OBJECTIVES: Our aim was to study the in vivo pfK13 transcriptional response in patients treated with artemether-lumefantrine and explore whether the pfk13 transcripts can explain the patients' parasite clearance outcomes.

    PATIENTS AND METHODS: A total of 47 Tanzanian children with microscopically confirmed uncomplicated P. falciparum malaria were hospitalized and received artemether-lumefantrine treatment (clinical trial ID: NCT00336375). RNA was extracted from venous blood samples collected before treatment initiation and at five more timepoints after treatment. cDNA was synthesized and pfk13 transcripts measured by real-time PCR.

    RESULTS: A wide range of pfk13 transcript variation was observed throughout all timepoints after artemether-lumefantrine treatment. Taking parasite clearance data together with the pfk13 transcripts profile, we observed a negative correlation inferring that pfk13 down-regulation is associated with longer parasite clearance time.

    CONCLUSIONS: The findings suggest that a reduced PfK13 transcriptional response may represent a first step towards artemisinin tolerance/resistance.

  • 66. Skov, R
    et al.
    Smyth, R
    Yusof, A
    Karlsson, A
    Mills, K
    Frimodt-Moller, N
    Kahlmeter, G
    Uppsala University, Disciplinary Domain of Medicine and Pharmacy, Faculty of Medicine, Department of Medical Sciences.
    Effects of temperature on the detection of methicillin resistance in Staphylococcus aureus using cefoxitin disc diffusion testing with Iso-Sensitest agar2009In: Journal of Antimicrobial Chemotherapy, ISSN 0305-7453, E-ISSN 1460-2091, Vol. 63, no 4, p. 699-703Article in journal (Refereed)
    Abstract [en]

    Cefoxitin is today the substance of choice for the phenotypic detection of methicillin-resistant Staphylococcus aureus (MRSA). We investigated the influence of incubation temperature in the standard range, i.e. 35-37 degrees C, and time, i.e. 18-20 h, versus a full 24 h. Cefoxitin disc testing was examined at incubation temperatures of 35 and 36 degrees C and times of 18-20 and 24 h, respectively, for 94 mecA-negative and 49 mecA-positive S. aureus on Iso-Sensitest agar using a semi-confluent inoculum. Cefoxitin inhibition zones on Iso-Sensitest agar were larger at temperatures above 35 degrees C; two isolates (4%, 95% confidence interval=0.5-14%) incubated at 36 degrees C were falsely categorized as susceptible to methicillin. Incubation time across 18-24 h did not impact results. Detection of methicillin resistance in S. aureus using the cefoxitin disc method with a semi-confluent inoculum on Iso-Sensitest agar is influenced by incubation temperature, and the temperature should not exceed 35 degrees C for the reliable detection of MRSA.

  • 67.
    Sundqvist, Martin
    et al.
    Uppsala University, Disciplinary Domain of Medicine and Pharmacy, Faculty of Medicine, Department of Medical Sciences, Infectious Diseases.
    Geli, Patricia
    Andersson, Dan I.
    Uppsala University, Disciplinary Domain of Medicine and Pharmacy, Faculty of Medicine, Department of Medical Biochemistry and Microbiology.
    Sjölund-Karlsson, M.
    Runehagen, A.
    Cars, H.
    Abelson-Storby, K.
    Cars, Otto
    Uppsala University, Disciplinary Domain of Medicine and Pharmacy, Faculty of Medicine, Department of Medical Sciences, Infectious Diseases.
    Kahlmeter, Gunnar
    Uppsala University, Disciplinary Domain of Medicine and Pharmacy, Faculty of Medicine, Department of Medical Sciences, Clinical Bacteriology.
    Little evidence for reversibility of trimethoprim resistance after a drastic reduction in trimethoprim use2010In: Journal of Antimicrobial Chemotherapy, ISSN 0305-7453, E-ISSN 1460-2091, Vol. 65, no 2, p. 350-360Article in journal (Refereed)
    Abstract [en]

    Objectives The worldwide rapid increase in antibiotic-resistant bacteria has made efforts to prolong the lifespan of existing antibiotics very important. Antibiotic resistance often confers a fitness cost in the bacterium. Resistance may thus be reversible if antibiotic use is discontinued or reduced. To examine this concept, we performed a 24 month voluntary restriction on the use of trimethoprim-containing drugs in Kronoberg County, Sweden. Methods The intervention was performed on a 14 year baseline of monthly data on trimethoprim resistance and consumption. A three-parameter mathematical model was used to analyse the intervention effect. The prerequisites for reversion of resistance (i.e. fitness cost, associated resistance and clonal composition) were studied on subsets of consecutively collected Escherichia coli from urinary tract infections. Results The use of trimethoprim-containing drugs decreased by 85% during the intervention. A marginal but statistically significant effect on the increase in trimethoprim resistance was registered. There was no change in the clonal composition of E. coli and there was no measurable fitness cost associated with trimethoprim resistance in clinical isolates. The frequency of associated antibiotic resistances in trimethoprim-resistant isolates was high. Conclusions A lack of detectable fitness cost of trimethoprim resistance in vitro together with a strong co-selection of other antibiotics could explain the rather disappointing effect of the intervention. The result emphasizes the low possibility of reverting antibiotic resistance once established and the urgent need for the development of new antibacterial agents.

  • 68.
    Sundqvist, Martin
    et al.
    Uppsala University, Disciplinary Domain of Medicine and Pharmacy, Faculty of Medicine, Department of Medical Sciences, Infectious Diseases.
    Kahlmeter, Gunnar
    Uppsala University, Disciplinary Domain of Medicine and Pharmacy, Faculty of Medicine, Department of Medical Sciences, Clinical Bacteriology.
    Effect of excluding duplicate isolates of Escherichia coli and Staphylococcus aureus in a 14 year consecutive database2007In: Journal of Antimicrobial Chemotherapy, ISSN 0305-7453, E-ISSN 1460-2091, Vol. 59, no 5, p. 913-918Article in journal (Refereed)
    Abstract [en]

    OBJECTIVES: It is recommended that duplicate isolates are excluded when reporting resistance rates. The rationale for this is that failing to do so will yield falsely high resistance rates. We analysed a 14 year consecutive database of Escherichia coli (n=62,380) and Staphylococcus aureus (n=28,178) using various cut-off algorithms to determine the importance of excluding duplicates and principal differences between the bacteria. METHODS: Susceptibility testing was performed according to the Swedish Reference Group for Antibiotics guidelines. Duplicates were excluded on the basis of species, individual and time (exclusion cut-offs of 7, 14, 30, 45, 90, 180, 270 and 365 days) from the first isolate. RESULTS: Although 30% of the isolates were excluded using a 365 day exclusion algorithm, the effects on resistance rates of excluding duplicates were small. Irrespective of cut-off, resistance in S. aureus decreased when duplicates were excluded. Using 7-30 days cut-offs, resistance in E. coli decreased or was not affected, whereas higher resistance rates were obtained when exclusion was based on a 365 day cut-off. Fluoroquinolone resistance was a clear exception to this rule. CONCLUSIONS: Although the effect of exclusion of duplicates was minor, we suggest that exclusion cut-offs should match the study timeline. The data presented on E. coli, from urinary tract infections, and S. aureus, from skin and soft tissue infections, suggest that E. coli infection, >90 days after the first culture, is mainly caused by new less-resistant strains. Patients with S. aureus continue to be colonized with the same strain.

  • 69.
    Sundqvist, Martin
    et al.
    Uppsala University, Disciplinary Domain of Medicine and Pharmacy, Faculty of Medicine, Department of Medical Sciences.
    Kahlmeter, Gunnar
    Uppsala University, Disciplinary Domain of Medicine and Pharmacy, Faculty of Medicine, Department of Medical Sciences.
    'Pre-emptive culturing' will improve the chance of 'getting it right' when empirical therapy of urinary tract infections fails2009In: Journal of Antimicrobial Chemotherapy, ISSN 0305-7453, E-ISSN 1460-2091, Vol. 64, no 2, p. 227-228Article in journal (Refereed)
    Abstract [en]

    Antibiotic resistance is increasing and beginning to affect the outcome of empirical antimicrobial therapy of urinary tract infections. Associated resistance, i.e. the fact that a bacterium resistant to one antibiotic is often much more likely to be resistant to other antibiotics, drastically decreases our chances of getting a second empirical attempt right. To increase the use of narrow-spectrum antibiotics, which are considered to be less selective for resistance, we need to develop new strategies from the laboratory to support our clinical colleagues. We suggest that 'pre-emptive culturing' of urine (i.e. a culture obtained before empirical treatment is instituted) will prevent clinicians from making a second improper empirical choice or having to resort to expensive broad-spectrum antimicrobials, which may drive resistance further. This strategy will be especially important in settings with high levels of resistance.

  • 70.
    Svensson, Elin M.
    et al.
    Uppsala University, Disciplinary Domain of Medicine and Pharmacy, Faculty of Pharmacy, Department of Pharmaceutical Biosciences.
    Karlsson, Mats O
    Uppsala University, Disciplinary Domain of Medicine and Pharmacy, Faculty of Pharmacy, Department of Pharmaceutical Biosciences.
    Modelling of mycobacterial load reveals bedaquiline's exposure-response relationship in patients with drug-resistant TB2017In: Journal of Antimicrobial Chemotherapy, ISSN 0305-7453, E-ISSN 1460-2091, Vol. 72, no 12, p. 3398-3405Article in journal (Refereed)
    Abstract [en]

    Background: Bedaquiline has been shown to reduce time to sputum culture conversion (SCC) and increase cure rates in patients with drug-resistant TB, but the influence of drug exposure remains uncharacterized.

    Objectives: To investigate whether an exposure-response relationship could be characterized by making better use of the existing information on pharmacokinetics and longitudinal measurements of mycobacterial load.

    Methods: Quantitative culture data in the form of time to positivity (TTP) in mycobacterial growth indicator tubes obtained from a randomized placebo-controlled Phase IIb registration trial were examined using non-linear mixed-effects methodology. The link to individual bedaquiline exposures and other patient characteristics was evaluated.

    Results: The developed model included three simultaneously fitted components: a longitudinal representation of mycobacterial load in patients, a probabilistic component for bacterial presence in sputum samples, and a time-to-event model for TTP. Data were described adequately, and time to SCC was well predicted. Individual bedaquiline exposure was found to significantly affect the decline in mycobacterial load. Consequently, the proportion of patients without SCC at week 20 is expected to decrease from 25% (95% CI 20%-31%) without bedaquiline to 17% (95% CI 13%-21%), 12% (95% CI 8%-16%) and 7% (95% CI 4%-11%), respectively, with half the median, median and double the median bedaquiline exposure observed in patients with standard dosing. Baseline bacterial load and level of drug resistance were other important predictors.

    Conclusions: To our knowledge, this is the first successful description of bedaquiline's exposure-response relationship and may be used when considering dose optimization. Characterization of this relationship was possible by integrating quantitative information in existing clinical data using novel models.

  • 71.
    Svensson, Elin M.
    et al.
    Uppsala University, Disciplinary Domain of Medicine and Pharmacy, Faculty of Pharmacy, Department of Pharmaceutical Biosciences.
    Murray, Stephen
    Karlsson, Mats O.
    Uppsala University, Disciplinary Domain of Medicine and Pharmacy, Faculty of Pharmacy, Department of Pharmaceutical Biosciences.
    Dooley, Kelly E.
    Rifampicin and rifapentine significantly reduce concentrations of bedaquiline, a new anti-TB drug2015In: Journal of Antimicrobial Chemotherapy, ISSN 0305-7453, E-ISSN 1460-2091, Vol. 70, no 4, p. 1106-1114Article in journal (Refereed)
    Abstract [en]

    Objectives: Bedaquiline is the first drug of a new class approved for the treatment of TB in decades. Bedaquiline is metabolized by cytochrome P450 (CYP) 3A4 to a less-active M2 metabolite. Its terminal half-life is extremely long (5-6 months), complicating evaluations of drug-drug interactions. Rifampicin and rifapentine, two anti-TB drugs now being optimized to shorten TB treatment duration, are potent inducers of CYP3A4. This analysis aimed to predict the effect of repeated doses of rifampicin or rifapentine on the steady-state pharmacokinetics of bedaquiline and its M2 metabolite from single-dose data using a model-based approach. Methods: Pharmacokinetic data for bedaquiline and M2 were obtained from a Phase I study involving 32 individuals each receiving two doses of bedaquiline, alone or together with multiple-dose rifampicin or rifapentine. Sampling was performed over 14 days following each bedaquiline dose. Pharmacokinetic analyses were performed using non-linear mixed-effects modelling. Models were used to simulate potential dose adjustments. Results: Rifamycin co-administration increased bedaquiline clearance substantially: 4.78-fold [ relative standard error (RSE) 9.10%] with rifampicin and 3.96-fold (RSE 5.00%) with rifapentine. Induction of M2 clearance was equally strong. Average steady-state concentrations of bedaquiline and M2 are predicted to decrease by 79% and 75% when given with rifampicin or rifapentine, respectively. Simulations indicated that increasing the bedaquiline dosage to mitigate the interaction would yield elevated M2 concentrations during the first treatment weeks. Conclusions: Rifamycin antibiotics reduce bedaquiline concentrations substantially. In line with current treatment guidelines for drug-susceptible TB, concomitant use is not recommended, even with dose adjustment.

  • 72.
    Svensson, Robin J.
    et al.
    Uppsala University, Disciplinary Domain of Medicine and Pharmacy, Faculty of Pharmacy, Department of Pharmaceutical Biosciences.
    Gillespie, Stephen H.
    Univ St Andrews, Sch Med, St Andrews KY16 9TF, Fife, Scotland..
    Simonsson, Ulrika S. H.
    Uppsala University, Disciplinary Domain of Medicine and Pharmacy, Faculty of Pharmacy, Department of Pharmaceutical Biosciences.
    Improved power for TB Phase IIa trials using a model-based pharmacokinetic-pharmacodynamic approach compared with commonly used analysis methods2017In: Journal of Antimicrobial Chemotherapy, ISSN 0305-7453, E-ISSN 1460-2091, Vol. 72, no 8, p. 2311-2319Article in journal (Refereed)
    Abstract [en]

    Background: The demand for new anti-TB drugs is high, but development programmes are long and costly. Consequently there is a need for new strategies capable of accelerating this process.

    Objectives: To explore the power to find statistically significant drug effects using a model-based pharmacokinetic-pharmacodynamic approach in comparison with the methods commonly used for analysing TB Phase IIa trials.

    Methods: Phase IIa studies of four hypothetical anti-TB drugs (labelled A, B, C and D), each with a different mechanism of action, were simulated using the multistate TB pharmacometric (MTP) model. cfu data were simulated over 14 days for patients taking once-dailymonotherapy at four different doses per drug and a reference (10mg/kg rifampicin). The simulated data were analysed using t-test, ANOVA, mono-and bi-exponential models and a pharmacokinetic-pharmacodynamic model approach (MTP model) to establish their respective power to find a drug effect at the 5% significance level.

    Results: For the pharmacokinetic-pharmacodynamic model approach, t-test, ANOVA, mono-exponential model and bi-exponential model, the sample sizes needed to achieve 90% power were: 10, 30, 75, 20 and 30 (drug A); 30, 75, 245, 75 and 105 (drug B); 70, > 1250, 315, > 1250 and >1250 (drug C); and 30, 110, 710, 170 and 185 (drug D), respectively.

    Conclusions: A model-based design and analysis using a pharmacokinetic-pharmacodynamic approach can reduce the number of patients required to determine a drug effect at least 2-fold compared with current methodologies. This could significantly accelerate early-phase TB drug development.

  • 73. Theuretzbacher, Ursula
    et al.
    Van Bambeke, Francoise
    Canton, Rafael
    Giske, Christian G.
    Mouton, Johan W.
    Nation, Roger L.
    Paul, Mical
    Turnidge, John D.
    Kahlmeter, Gunnar
    Uppsala University, Disciplinary Domain of Medicine and Pharmacy, Faculty of Medicine, Department of Medical Sciences, Clinical Microbiology and Infectious Medicine, Clinical Bacteriology.
    Reviving old antibiotics2015In: Journal of Antimicrobial Chemotherapy, ISSN 0305-7453, E-ISSN 1460-2091, Vol. 70, no 8, p. 2177-2181Article in journal (Refereed)
    Abstract [en]

    In the face of increasing antimicrobial resistance and the paucity of new antimicrobial agents it has become clear that new antimicrobial strategies are urgently needed. One of these is to revisit old antibiotics to ensure that they are used correctly and to their full potential, as well as to determine whether one or several of them can help alleviate the pressure on more recent agents. Strategies are urgently needed to 're-develop' these drugs using modern standards, integrating new knowledge into regulatory frameworks and communicating the knowledge from the research bench to the bedside. Without a systematic approach to re-developing these old drugs and rigorously testing them according to today's standards, there is a significant risk of doing harm to patients and further increasing multidrug resistance. This paper describes factors to be considered and outlines steps and actions needed to re-develop old antibiotics so that they can be used effectively for the treatment of infections.

  • 74.
    Thorsted, Anders
    et al.
    Uppsala University, Disciplinary Domain of Medicine and Pharmacy, Faculty of Pharmacy, Department of Pharmaceutical Biosciences.
    Kristoffersson, Anders N.
    Uppsala University, Disciplinary Domain of Medicine and Pharmacy, Faculty of Pharmacy, Department of Pharmaceutical Biosciences. Pharmetheus AB, Uppsala, Sweden.
    Maarbjerg, Sabine F.
    Aarhus Univ Hosp, Dept Pediat & Adolescent Med, Aarhus, Denmark.
    Schrøder, Henrik
    Aarhus Univ Hosp, Dept Pediat & Adolescent Med, Aarhus, Denmark.
    Wang, Mikala
    Aarhus Univ Hosp, Dept Clin Microbiol, Aarhus, Denmark.
    Brock, Birgitte
    Aarhus Univ Hosp, Dept Clin Biochem, Aarhus, Denmark;Steno Diabet Ctr Copenhagen, Gentofte, Denmark.
    Nielsen, Elisabet I.
    Uppsala University, Disciplinary Domain of Medicine and Pharmacy, Faculty of Pharmacy, Department of Pharmaceutical Biosciences.
    Friberg, Lena E.
    Uppsala University, Disciplinary Domain of Medicine and Pharmacy, Faculty of Pharmacy, Department of Pharmaceutical Biosciences.
    Population pharmacokinetics of piperacillin in febrile children receiving cancer chemotherapy: the impact of body weight and target on an optimal dosing regimen2019In: Journal of Antimicrobial Chemotherapy, ISSN 0305-7453, E-ISSN 1460-2091, Vol. 74, no 10, p. 2984-2993Article in journal (Refereed)
    Abstract [en]

    Background The β-lactam antibiotic piperacillin (in combination with tazobactam) is commonly chosen for empirical treatment of suspected bacterial infections. However, pharmacokinetic variability among patient populations and across ages leads to uncertainty when selecting a dosing regimen to achieve an appropriate pharmacodynamic target.

    Objectives To guide dosing by establishing a population pharmacokinetic model for unbound piperacillin in febrile children receiving cancer chemotherapy, and to assess pharmacokinetic/pharmacodynamic target attainment (100% fT>1xMIC and 50% fT>4xMIC) and resultant exposure, across body weights.

    Methods Forty-three children admitted for 89 febrile episodes contributed 482 samples to the pharmacokinetic analysis. The typical doses required for target attainment were compared for various dosing regimens, in particular prolonged infusions, across MICs and body weights.

    Results A two-compartment model with inter-fever-episode variability in CL, and body weight included through allometry, described the data. A high CL of 15.4L/h (70kg) combined with high glomerular filtration rate (GFR) values indicated rapid elimination and hyperfiltration. The target of 50% fT>4xMIC was achieved for an MIC of 4.0mg/L in a typical patient with extended infusions of 2-3 (q6h) or 3-4 (q8h)h, at or below the standard adult dose (75 and 100mg/kg/dose for q6h and q8h, respectively). Higher doses or continuous infusion were needed to achieve 100% fT>1xMIC due to the rapid piperacillin elimination.

    Conclusions The licensed dose for children with febrile neutropenia (80mg/kg q6h as a 30min infusion) performs poorly for attainment of fT>MIC pharmacokinetic/pharmacodynamic targets. Given the population pharmacokinetic profile, feasible dosing regimens with reasonable exposure are continuous infusion (100% fT>1xMIC) or prolonged infusions (50% fT>4xMIC).

  • 75.
    Tängdén, Thomas
    et al.
    Uppsala University, Disciplinary Domain of Medicine and Pharmacy, Faculty of Medicine, Department of Medical Sciences, Infectious Diseases.
    Adler, Marlen
    Uppsala University, Disciplinary Domain of Medicine and Pharmacy, Faculty of Medicine, Department of Medical Biochemistry and Microbiology.
    Cars, Otto
    Uppsala University, Disciplinary Domain of Medicine and Pharmacy, Faculty of Medicine, Department of Medical Sciences, Infectious Diseases.
    Sandegren, Linus
    Uppsala University, Disciplinary Domain of Medicine and Pharmacy, Faculty of Medicine, Department of Medical Biochemistry and Microbiology.
    Löwdin, Elisabeth
    Uppsala University, Disciplinary Domain of Medicine and Pharmacy, Faculty of Medicine, Department of Medical Sciences, Infectious Diseases.
    Frequent emergence of porin-deficient subpopulations with reduced carbapenem susceptibility in ESBL-producing Escherichia coli during exposure to ertapenem in an in vitro pharmacokinetic model2013In: Journal of Antimicrobial Chemotherapy, ISSN 0305-7453, E-ISSN 1460-2091, Vol. 68, no 6, p. 1319-1326Article in journal (Refereed)
    Abstract [en]

    OBJECTIVES:

    Ertapenem resistance is increasing in Enterobacteriaceae. The production of extended-spectrum β-lactamases (ESBLs) and reduced expression of outer membrane porins are major mechanisms of resistance in ertapenem-resistant Klebsiella pneumoniae. Less is known of ertapenem resistance in Escherichia coli. The aim of this study was to explore the impact of ESBL production in E. coli on the antibacterial activity of ertapenem.

    METHODS:

    Two E. coli strains, with and without ESBL production, were exposed to ertapenem in vitro for 48 h at concentrations simulating human pharmacokinetics with conventional and higher dosages.

    RESULTS:

    Isolates with non-susceptibility to ertapenem (MICs 0.75-1.5 mg/L) were detected after five of nine time-kill experiments with the ESBL-producing strain. All of these isolates had ompR mutations, which reduce the expression of outer membrane porins OmpF and OmpC. Higher dosage did not prevent selection of porin-deficient subpopulations. No mutants were detected after experiments with the non-ESBL-producing strain. Compared with other experiments, experiments with ompR mutants detected in endpoint samples showed significantly less bacterial killing after the second dose of ertapenem. Impaired antibacterial activity against E. coli with ESBL production and ompR mutation was also demonstrated in time-kill experiments with static antibiotic concentrations.

    CONCLUSIONS:

    The combination of ESBL production and porin loss in E. coli can result in reduced susceptibility to ertapenem. Porin-deficient subpopulations frequently emerged in ESBL-producing E. coli during exposure to ertapenem at concentrations simulating human pharmacokinetics. Inappropriate use of ertapenem should be avoided to minimize the risk of selection of ESBL-producing bacteria with reduced susceptibility to carbapenems.

  • 76.
    Tängdén, Thomas
    et al.
    Uppsala University, Disciplinary Domain of Medicine and Pharmacy, Faculty of Medicine, Department of Medical Sciences, Infectious Diseases.
    Eriksson, Britt-Marie
    Uppsala University, Disciplinary Domain of Medicine and Pharmacy, Faculty of Medicine, Department of Medical Sciences, Infectious Diseases.
    Melhus, Åsa
    Uppsala University, Disciplinary Domain of Medicine and Pharmacy, Faculty of Medicine, Department of Medical Sciences, Clinical Bacteriology.
    Svennblad, Bodil
    Uppsala University, Disciplinary Domain of Medicine and Pharmacy, Medicinska och farmaceutiska vetenskapsområdet, centrumbildningar mm , UCR-Uppsala Clinical Research Center.
    Cars, Otto
    Uppsala University, Disciplinary Domain of Medicine and Pharmacy, Faculty of Medicine, Department of Medical Sciences, Infectious Diseases.
    Radical reduction of cephalosporin use at a tertiary hospital after educational antibiotic intervention during an outbreak of extended-spectrum beta-lactamase-producing Klebsiella pneumoniae2011In: Journal of Antimicrobial Chemotherapy, ISSN 0305-7453, E-ISSN 1460-2091, Vol. 66, no 5, p. 1161-1167Article in journal (Refereed)
    Abstract [en]

    Objectives: During an outbreak of extended-spectrum beta-lactamase (ESBL)-producing Klebsiella pneumoniae at our hospital, we performed an educational antibiotic intervention aimed at reducing prescriptions of second- and third-generation cephalosporins and preventing increased use of fluoroquinolones and carbapenems. In this report, we describe the implementation strategy used and evaluate the intervention effect according to Cochrane recommendations. Methods: New recommendations for empirical intravenous antibiotic treatment were communicated to prescribers throughout the hospital by infectious diseases physicians working with Strama (the Swedish strategic programme against antibiotic resistance). No restrictive measures were used. The intervention effect was analysed with interrupted time series (ITS) regression analysis of local and national monthly antibiotic sales data. Results: A radical immediate and sustained reduction was demonstrated for the cephalosporins targeted in the intervention, whereas consumption of piperacillin/tazobactam and penicillin G increased substantially. Fluoroquinolone and carbapenem use was essentially unchanged. The ESBL outbreak subsided and no increased resistance to piperacillin/tazobactam was detected in K. pneumoniae, Escherichia coli or Pseudomonas aeruginosa blood isolates during the 2.5 year follow-up. Conclusions: Our study clearly demonstrates that an educational intervention can have an immediate and profound effect on antibiotic prescription patterns at a large tertiary hospital. ITS regression analysis of local and national antibiotic sales data was valuable to readily assess the immediate and sustained effects of the intervention.

  • 77.
    Ungphakorn, Wanchana
    et al.
    Uppsala University, Disciplinary Domain of Medicine and Pharmacy, Faculty of Pharmacy, Department of Pharmaceutical Biosciences.
    Tängdén, Thomas
    Uppsala University, Disciplinary Domain of Medicine and Pharmacy, Faculty of Medicine, Department of Medical Sciences, Infectious Diseases.
    Sandegren, Linus
    Uppsala University, Disciplinary Domain of Medicine and Pharmacy, Faculty of Medicine, Department of Medical Biochemistry and Microbiology.
    Nielsen, Elisabet I.
    Uppsala University, Disciplinary Domain of Medicine and Pharmacy, Faculty of Pharmacy, Department of Pharmaceutical Biosciences.
    A pharmacokinetic-pharmacodynamic model characterizing the emergence of resistant Escherichia coli subpopulations during ertapenem exposure2016In: Journal of Antimicrobial Chemotherapy, ISSN 0305-7453, E-ISSN 1460-2091, Vol. 71, no 9, p. 2521-2533Article in journal (Refereed)
    Abstract [en]

    Resistant subpopulations with reduced expression of outer membrane porins have been observed in ESBL-producing Escherichia coli during exposure to ertapenem. The aim of this work was to develop a pharmacokinetic-pharmacodynamic (PKPD) model to characterize the emergence of resistant E. coli during exposure to ertapenem and to predict bacterial killing following different dosing regimens of ertapenem. Data from in vitro time-kill experiments were used to develop a mechanism-based PKPD model for three E. coli strains: a native strain, an ESBL-producing strain, and an ESBL-producing strain with reduced expression of porins OmpF and OmpC. Each strain was exposed to static ertapenem concentrations (1-512aEuroSxaEuroSMIC) for 24 h using starting inocula of similar to 10(6) and 10(8) cfu/mL. The developed PKPD model consisted of three bacterial states: susceptible growing, less susceptible non-growing, and non-susceptible non-growing bacteria. A pre-existing bacterial subpopulation was used to describe the emergence of resistance. The PKPD model adequately characterized the data of the three E. coli strains investigated. Results from predictions suggest that the conventional dosage (1 g intravenously once daily) might result in regrowth of resistant subpopulations when used to treat infection caused by ESBL-producing strains. Resistant subpopulations frequently emerged in E. coli when exposed to ertapenem, supporting that the time course of emergence of resistance should be taken into consideration when selecting dosing regimens.

  • 78. Westman, Eva
    et al.
    Melhus, Åsa
    Uppsala University, Disciplinary Domain of Medicine and Pharmacy, Faculty of Medicine, Department of Medical Sciences.
    The treatment of Haemophilus influenzae acute otitis media with amoxicillin protects against reinfection but not against structural changes2002In: Journal of Antimicrobial Chemotherapy, ISSN 0305-7453, E-ISSN 1460-2091, Vol. 49, no 1, p. 141-7Article in journal (Refereed)
    Abstract [en]

    Acute otitis media (AOM) is the most common reason for outpatient antimicrobial therapy today. With increasing problems with antibiotic resistance, a more restrictive use of antibiotics has been advocated for both single and recurrent episodes. The arguments advanced have been immunological as well as ecological. To study the effects of a 5 day course of amoxicillin on recurrent AOM caused by non-typeable Haemophilus influenzae, Sprague-Dawley rats were used. Amoxicillin was introduced at the clinical peak of the first infection. One month later the animals were rechallenged. Local and systemic changes were monitored by otomicroscopy, bacterial cultures, and analyses of systemic IgG responses and histological changes. Antibiotic treatment accelerated the resolution of the primary infection. After resolution, only minor morphological changes were observed. The protective rate at rechallenge was 100% in the treatment group, compared with 80% in the untreated control group. Although the production of serum IgG antibodies was initially slightly impeded by the treatment, it was significantly higher in treated animals after rechallenge. Major structural changes could not be avoided at the second infection, however, and a significantly higher frequency of myringosclerosis was observed in treated animals. These experimental findings constitute support for further studies of antimicrobial drugs and recurrent AOM.

  • 79.
    Wiuff, Camilla
    et al.
    Uppsala University, Disciplinary Domain of Medicine and Pharmacy, Faculty of Medicine, Department of Medical Biochemistry and Microbiology.
    Andersson, Dan I.
    Uppsala University, Disciplinary Domain of Medicine and Pharmacy, Faculty of Medicine, Department of Medical Biochemistry and Microbiology.
    Antibiotic treatment in vitro of phenotypically tolerant bacterial populations2007In: Journal of Antimicrobial Chemotherapy, ISSN 0305-7453, E-ISSN 1460-2091, Vol. 59, no 2, p. 254-263Article in journal (Refereed)
    Abstract [en]

    Objectives: Most pharmacodynamic models used for design of treatment regimens are based on time-kill data obtained with normal cells in the susceptible state without taking into account the killing kinetics of the antibiotic-tolerant cells in the population. We compared the microbiological efficacy of six antibiotics against tolerant cells and by mathematical modelling explored the potential clinical implications of tolerance. Methods: Tolerant cells were obtained by filtration of bacterial cultures of Escherichia coli MG1655 after antibiotic exposure. Killing kinetics of the tolerant cells was compared with that of exponentially growing naive cells. To examine the nutrient dependency of the reversion from the tolerant state to the susceptible state, tolerant cells were re-suspended in Luria-Bertani and PBS and re-exposed to antibiotics. A mathematical model was used to explore the clinical implications of antibiotic tolerance. Results: Streptomycin was the most efficient drug against tolerant cells. Ciprofloxacin and ampicillin had intermediate activity against tolerant cells while rifampicin, tetracycline and erythromycin had poor activity against tolerant cells. No correlation could be established between the microbiological efficacies against susceptible and tolerant cells. Reversion from tolerance to susceptibility was dependent on the presence of nutrients and growth. Computer simulations demonstrated that the efficacy of antibiotics against tolerant cell populations has a large influence on treatment outcome. Conclusions: The in vitro killing kinetics of tolerant cells is antibiotic-dependent and different from that of cells in the susceptible state. This difference in efficacy could have an influence on treatment outcome and tolerance should therefore be studied further in vivo.

  • 80.
    Zorzet, Anna
    et al.
    Uppsala University, Disciplinary Domain of Medicine and Pharmacy, Faculty of Medicine, Department of Medical Biochemistry and Microbiology.
    Andersen, Jytte Mark
    Nilsson, Annika I
    Møller, Niels Frimodt
    Andersson, Dan I
    Uppsala University, Disciplinary Domain of Medicine and Pharmacy, Faculty of Medicine, Department of Medical Biochemistry and Microbiology.
    Compensatory mutations in agrC partly restore fitness in vitro to peptide deformylase inhibitor-resistant Staphylococcus aureus2012In: Journal of Antimicrobial Chemotherapy, ISSN 0305-7453, E-ISSN 1460-2091, Vol. 67, no 8, p. 1835-1842Article in journal (Refereed)
    Abstract [en]

    OBJECTIVES

    To determine how the fitness cost of deformylase inhibitor resistance conferred by fmt mutations can be genetically compensated.

    METHODS

    Resistant mutants were isolated and characterized with regard to their growth rates in vitro and in neutropenic mice, MIC and DNA sequence. Faster-growing compensated mutants were isolated by serial passage in culture medium, and for a subset of the resistant and compensated mutants whole-genome sequencing was performed.

    RESULTS

    Staphylococcus aureus mutants resistant to the peptide deformylase inhibitor actinonin had mutations in the fmt gene that conferred high-level actinonin resistance and reduced bacterial growth rate. Compensated mutants that remained fully resistant to actinonin and showed increased growth rates appeared within 30-60 generations of growth. Whole-genome sequencing and localized DNA sequencing of mutated candidate genes showed that alterations in the gene agrC were present in the majority of compensated strains. Resistant and compensated mutants grew at similar rates as the wild-type in a mouse thigh infection model.

    CONCLUSIONS

    Resistance to deformylase inhibitors due to fmt mutations reduces bacterial growth rates, but these costs can be reduced by mutations in the agrC gene. Mutants defective in fmt (with or without compensatory agrC mutations) grew well in an animal model, implying that they can also cause infection in a host.

  • 81. Zvada, Simbarashe P.
    et al.
    Denti, Paolo
    Donald, Peter R.
    Schaaf, H. Simon
    Thee, Stephanie
    Seddon, James A.
    Seifart, Heiner I.
    Smith, Peter J.
    McIlleron, Helen M.
    Simonsson, Ulrika S. H.
    Uppsala University, Disciplinary Domain of Medicine and Pharmacy, Faculty of Pharmacy, Department of Pharmaceutical Biosciences.
    Population pharmacokinetics of rifampicin, pyrazinamide and isoniazid in children with tuberculosis: in silico evaluation of currently recommended doses2014In: Journal of Antimicrobial Chemotherapy, ISSN 0305-7453, E-ISSN 1460-2091, Vol. 69, no 5, p. 1339-1349Article in journal (Refereed)
    Abstract [en]

    To describe the population pharmacokinetics of rifampicin, pyrazinamide and isoniazid in children and evaluate the adequacy of steady-state exposures. We used previously published data for 76 South African children with tuberculosis to describe the population pharmacokinetics of rifampicin, pyrazinamide and isoniazid. Monte Carlo simulations were used to predict steady-state exposures in children following doses in fixed-dose combination tablets in accordance with the revised guidelines. Reference exposures were derived from an ethnically similar adult population with tuberculosis taking currently recommended doses. The final models included allometric scaling of clearance and volume of distribution using body weight. Maturation was included for clearance of isoniazid and clearance and absorption transit time of rifampicin. For a 2-year-old child weighing 12.5 kg, the estimated typical oral clearances of rifampicin and pyrazinamide were 8.15 and 1.08 L/h, respectively. Isoniazid typical oral clearance (adjusted for bioavailability) was predicted to be 4.44, 11.6 and 14.6 L/h for slow, intermediate and fast acetylators, respectively. Higher oral clearance values in intermediate and fast acetylators also resulted from 23 lower bioavailability compared with slow acetylators. Simulations based on our models suggest that with the new WHO dosing guidelines and utilizing available paediatric fixed-dose combinations, children will receive adequate rifampicin exposures when compared with adults, but with a larger degree of variability. However, pyrazinamide and isoniazid exposures in many children will be lower than in adults. Further studies are needed to confirm these findings in children administered the revised dosages and to optimize pragmatic approaches to dosing.

  • 82. Ängeby, Kristian A.
    et al.
    Jureen, P.
    Giske, C. G.
    Chryssanthou, E.
    Sturegård, E.
    Nordvall, M.
    Johansson, A. G.
    Werngren, J.
    Kahlmeter, Gunnar
    Uppsala University, Disciplinary Domain of Medicine and Pharmacy, Faculty of Medicine, Department of Medical Sciences.
    Hoffner, S. E.
    Schön, T.
    Wild-type MIC distributions of four fluoroquinolones active against Mycobacterium tuberculosis in relation to current critical concentrations and available pharmacokinetic and pharmacodynamic data2010In: Journal of Antimicrobial Chemotherapy, ISSN 0305-7453, E-ISSN 1460-2091, Vol. 65, no 5, p. 946-952Article in journal (Refereed)
    Abstract [en]

    OBJECTIVES: To describe wild-type distributions of the MIC of fluoroquinolones for Mycobacterium tuberculosis in relation to current critical concentrations used for drug susceptibility testing and pharmacokinetic/pharmacodynamic (PK/PD) data. METHODS: A 96-stick replicator on Middlebrook 7H10 medium was used to define the MICs of ciprofloxacin, ofloxacin, moxifloxacin and levofloxacin for 90 consecutive clinical strains and 24 drug-resistant strains. The MICs were compared with routine BACTEC 460 susceptibility results and with MIC determinations in the BACTEC MGIT 960 system in a subset of strains using ofloxacin as a class representative. PK/PD data for each drug were reviewed in relation to the wild-type MIC distribution. RESULTS: The wild-type MICs of ciprofloxacin, ofloxacin, moxifloxacin and levofloxacin were distributed from 0.125 to 1, 0.25 to 1, 0.032 to 0.5 and 0.125 to 0.5 mg/L, respectively. The MIC data correlated well with the BACTEC 960 MGIT and BACTEC 460 results. PD indices were the most favourable for levofloxacin, followed by moxifloxacin, ofloxacin and ciprofloxacin. CONCLUSIONS: We propose S (susceptible)

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