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  • About
  • The Global ETD Search service is a free service for researchers to find electronic theses and dissertations. This service is provided by the Networked Digital Library of Theses and Dissertations.
    Our metadata is collected from universities around the world. If you manage a university/consortium/country archive and want to be added, details can be found on the NDLTD website.
1

Les effets indésirables des fluoroquinolones chez les sujets âgés de 65 ans et plus

Brunhes, Marie-Pierre. Paille, François January 2003 (has links) (PDF)
Reproduction de : Thèse d'exercice : Médecine : Nancy 1 : 2003. / Titre provenant de l'écran-titre.
2

Investigation of the in vitro development of fluoroquinolone-resistance in salmonellae.

January 2004 (has links)
Jin Yongjie. / Thesis (M.Phil.)--Chinese University of Hong Kong, 2004. / Includes bibliographical references (leaves 138-171). / Abstracts in English and Chinese. / Abstract (in English) --- p.i / Abstract (in Chinese) --- p.iii / Acknowledgments --- p.iv / Table of Contents --- p.v / List of Tables --- p.x / List of Figures --- p.xi / Chapter Chapter 1 --- Introduction --- p.1 / Chapter 1. --- Salmonella --- p.1 / Chapter 1.1 --- Morphology --- p.1 / Chapter 1.2 --- Antigenic structure --- p.1 / Chapter 1.3 --- Identification --- p.2 / Chapter 1.4 --- Nomenclature and classification --- p.2 / Chapter 1.5 --- Pathogenesis and virulence --- p.3 / Chapter 1.6 --- Infections --- p.4 / Chapter 1.6.1 --- Enteric fever --- p.4 / Chapter 1.6.2 --- Gastroenteritis --- p.4 / Chapter 1.7 --- Treatment --- p.5 / Chapter 1.7.1 --- Enteric fever --- p.5 / Chapter 1.7.2 --- Gastroenteritis --- p.5 / Chapter 1.8 --- Epidemiology and control --- p.6 / Chapter 1.8.1 --- Enteric fever --- p.6 / Chapter 1.8.2 --- Salmonella food poisoning --- p.6 / Chapter 2. --- Fluoroquinolones --- p.6 / Chapter 2.1 --- History of fluoroquinolones --- p.8 / Chapter 2.2 --- Mechanisms of action --- p.9 / Chapter 3. --- Antimicrobial resistance --- p.10 / Chapter 3.1 --- Microorganism-mediated resistance --- p.10 / Chapter 3.1.1 --- Intrinsic resistance --- p.11 / Chapter 3.1.2 --- Acquired resistance --- p.11 / Chapter 3.2 --- Resistance due to environmental factors --- p.12 / Chapter 3.3 --- Biological and clinical resistance --- p.12 / Chapter 3.4 --- Breakpoints --- p.13 / Chapter 4. --- Fluoroquinolone-resistance --- p.14 / Chapter 4.1 --- Increasing fluoroquinolone-resistance in bacteria --- p.14 / Chapter 4.2 --- Mechanisms of resistance to fluoroquinolones --- p.16 / Chapter 4.2.1 --- Mutations of target genes --- p.17 / Chapter 4.2.2 --- Active efflux systems and decreased membrane permeability --- p.20 / Chapter 5. --- Fluoroquinolone-resistance in Salmonella --- p.21 / Chapter 5.1 --- Prevalence of fluoroquinolone-resistant salmonellae in man and Animals --- p.21 / Chapter 5.1.1 --- Prevalence in the world --- p.21 / Chapter 5.1.2 --- Increasing resistance trend in Hong Kong --- p.24 / Chapter 5.2 --- Clinical outcome --- p.25 / Chapter 5.3 --- Mechanisms of fluoroquinolone-resistance in Salmonella --- p.25 / Chapter 5.3.1 --- Target gene mutations --- p.25 / Chapter 5.3.2 --- Other resistance mechanisms --- p.28 / Chapter 5.3.3 --- In vitro development of Salmonella resistant mutants --- p.29 / Chapter 6. --- Restricting the development of resistant mutants --- p.31 / Chapter 6.1 --- Mutant prevention concentration (MPC) --- p.31 / Chapter 6.1.1 --- Definition --- p.31 / Chapter 6.1.2 --- Development of MPC concept --- p.31 / Chapter 6.1.3 --- Significance --- p.34 / Chapter 6.2 --- Mutant selection window (MSW) --- p.35 / Chapter 6.2.1 --- The concept of mutant selection window (MSW) --- p.35 / Chapter 6.2.2 --- Significance --- p.36 / Chapter 7. --- Detection of gene mutations --- p.37 / Chapter 8. --- Objectives --- p.37 / Chapter Chapter 2 --- Materails and Methods --- p.39 / Chapter 1. --- Materials --- p.39 / Chapter 1.1 --- Bacterial strains --- p.39 / Chapter 1.1.1 --- Bacterial strains used for this study --- p.39 / Chapter 1.1.2 --- Storage of bacterial strains --- p.39 / Chapter 1.2 --- Materials --- p.40 / Chapter 2. --- Methods --- p.40 / Chapter 2.1 --- Identification --- p.40 / Chapter 2.2 --- Microbiological methods --- p.40 / Chapter 2.2.1 --- Determination of minimal inhibitory concentration (MIC) --- p.40 / Chapter 2.2.1.1 --- Preparation of antibiotic plates --- p.40 / Chapter 2.2.1.2 --- Preparation of inoculum --- p.43 / Chapter 2.2.1.3 --- Inoculation of antibiotic plates --- p.43 / Chapter 2.2.2 --- Determination of mutant prevention concentration (MPC) --- p.43 / Chapter 2.2.2.1 --- Preparation of bacterial suspension --- p.43 / Chapter 2.2.2.2 --- Inoculation of bacterial suspension --- p.43 / Chapter 2.2.2.3 --- Determination of the size of the inoculum --- p.43 / Chapter 2.2.2.4 --- Reading of results --- p.45 / Chapter 2.3 --- Molecular methods --- p.45 / Chapter 2.3.1 --- Polymerase chain reaction (PCR) --- p.45 / Chapter 2.3.2 --- Agarose gel electrophoresis --- p.47 / Chapter 2.3.3 --- Multiplex PCR amplimer conformation (MPAC) analysis --- p.47 / Chapter 2.3.3.1 --- Preparation of samples --- p.47 / Chapter 2.3.3.2 --- Electrophoresis --- p.49 / Chapter 2.3.3.3 --- Silver staining --- p.49 / Chapter 2.3.4 --- DNA Sequencing --- p.50 / Chapter 2.3.4.1 --- Purification of PCR products --- p.50 / Chapter 2.3.4.2 --- Sequencing reactions --- p.50 / Chapter 2.3.4.3 --- Electrophoresis --- p.50 / Chapter 2.3.4.4 --- Silver staining --- p.52 / Chapter 2.4 --- Stepwise selection and characterization of mutants --- p.53 / Chapter 2.4.1 --- In vitro selection of first-step strains --- p.53 / Chapter 2.4.2 --- Characterization of selected strains --- p.53 / Chapter 2.4.3 --- Subsequent selection of stepwise mutants --- p.54 / Chapter 3. --- Research plan --- p.54 / Chapter Chapter 3 --- Results --- p.55 / Chapter 1. --- Identification and fluoroquinolone MICs of Salmonella strains --- p.55 / Chapter 1.1 --- Identification of Salmonella strains --- p.55 / Chapter 1.2 --- Fluoroquinolone MICs for Salmonella strains --- p.55 / Chapter 1.2.1 --- Salmonella Typhimurium --- p.55 / Chapter 1.2.2 --- Salmonella Hadar --- p.57 / Chapter 2. --- Mutant prevention concentration (MPC) --- p.57 / Chapter 2.1 --- Salmonella Typhimurium --- p.57 / Chapter 2.1.1 --- MPC value --- p.57 / Chapter 2.1.2 --- MPC/MIC ratio --- p.57 / Chapter 2.1.3 --- Cmax/MPC ratio --- p.60 / Chapter 2.1.4 --- Frequencies of development of resistant strains --- p.60 / Chapter 2.2 --- Salmonella Hadar --- p.63 / Chapter 2.2.1 --- MPC value --- p.63 / Chapter 2.2.2 --- MPC/MIC ratio --- p.63 / Chapter 2.2.3 --- Cmax/MPC ratio --- p.63 / Chapter 2.2.4 --- Frequencies of development of resistant strains --- p.65 / Chapter 3. --- Stepwise selection of resistant mutants --- p.65 / Chapter 3.1 --- Salmonella Typhimurium --- p.70 / Chapter 3.1.1 --- Characterization of 1 St-step strains --- p.70 / Chapter 3.1.1.1 --- Antimicrobial susceptibilities --- p.70 / Chapter 3.1.1.2 --- Characterization of gene mutations --- p.75 / Chapter 3.1.1.3 --- Mutations and fluoroquinolone susceptibilities --- p.80 / Chapter 3.1.1.4 --- Mutations and nalidixic acid susceptibilities --- p.81 / Chapter 3.1.2 --- Characterization of 2nd-step mutants --- p.81 / Chapter 3.1.2.1 --- Antimicrobial susceptibilities --- p.82 / Chapter 3.1.2.2 --- Characterization of gene mutations --- p.85 / Chapter 3.1.2.3 --- Mutations and fluoroquinolone susceptibilities --- p.87 / Chapter 3.1.2.4 --- Nalidixic acid susceptibilities --- p.87 / Chapter 3.1.3 --- Characterization of 3rd-step mutants --- p.88 / Chapter 3.1.3.1 --- Antimicrobial susceptibilities --- p.88 / Chapter 3.1.3.2 --- Characterization of gene mutations --- p.92 / Chapter 3.1.3.3 --- Mutations and fluoroquinolone susceptibilities --- p.94 / Chapter 3.1.3.4 --- Nalidixic acid susceptibilities --- p.95 / Chapter 3.1.4 --- Characterization of 4th-step mutants --- p.95 / Chapter 3.1.4.1 --- Antimicrobial susceptibilities --- p.95 / Chapter 3.1.4.2 --- Characterization of gene mutations --- p.98 / Chapter 3.1.4.3 --- Nalidixic acid susceptibilities --- p.98 / Chapter 3.2 --- Salmonella Hadar --- p.98 / Chapter 3.2.1 --- Characterization of 1 St-step strains --- p.98 / Chapter 3.2.1.1 --- Antimicrobial susceptibilities --- p.99 / Chapter 3.2.1.2 --- Characterization of gene mutations --- p.102 / Chapter 3.2.1.3 --- Mutations and fluoroquinolone susceptibilities --- p.107 / Chapter 3.2.1.4 --- Mutations and nalidixic acid susceptibilities --- p.108 / Chapter 3.2.2 --- Characterization of 2nd-step mutants --- p.109 / Chapter 3.2.2.1 --- Antimicrobial susceptibilities --- p.109 / Chapter 3.2.2.2 --- Characterization of gene mutations --- p.112 / Chapter 3.2.2.3 --- Mutations and fluoroquinolone susceptibilities --- p.112 / Chapter 3.2.2.4 --- Nalidixic acid susceptibilities --- p.113 / Chapter Chapter 4 --- Discussion --- p.114 / Chapter 1. --- Susceptibility of salmonellae to fluoroquinolones --- p.114 / Chapter 2. --- The potential of fluoroquinolones to restrict the development of Salmonella resistant strains --- p.114 / Chapter 2.1 --- MPC and MPC/MIC of fluoroquinolones --- p.115 / Chapter 2.2 --- Cmax/MPC of fluoroquinolones --- p.117 / Chapter 2.3 --- Selection frequency of fluoroquinolones --- p.118 / Chapter 2.4 --- Effects of fluoroquinolones on the development of resistancein Salmonella Typhimurium and Salmonella Hadar --- p.119 / Chapter 3. --- Characterization of in vitro fluoroquinolone-resistant Salmonella mutants --- p.120 / Chapter 3.1 --- Development of resistance phenotype --- p.120 / Chapter 3.1.1 --- Microbiology --- p.120 / Chapter 3.1.2 --- Antimicrobial susceptibilities --- p.120 / Chapter 3.1.2.1 --- First-step strains --- p.120 / Chapter 3.1.2.2 --- Second-step mutants --- p.121 / Chapter 3.1.2.3 --- Third-step mutants --- p.121 / Chapter 3.1.2.4 --- Fourth-step mutants --- p.122 / Chapter 3.2 --- Contribution of target gene mutations to resistance development --- p.122 / Chapter 3.2.1 --- First-step mutants --- p.122 / Chapter 3.2.2 --- Second-step mutants --- p.125 / Chapter 3.2.3 --- Third-step mutants --- p.126 / Chapter 3.2.4 --- Fourth-step mutants --- p.128 / Chapter 3.3 --- The sequential development of gene mutations --- p.129 / Chapter 3.4 --- Other fluoroquinolone-resistance mechanisms --- p.130 / Chapter 4. --- Mutations and susceptibilities to fluoroquinolones and nalidixic acid --- p.132 / Chapter 4.1 --- Nalidixic acid - a marker for resistance to fluoroquinolones --- p.132 / Chapter 4.2 --- Breakpoint and clinical efficiency --- p.133 / Chapter 5. --- Strategies to reduce development of fluoroquinolone-resistance --- p.134 / Chapter 6. --- Conclusion --- p.136 / Chapter 7. --- Areas for future research --- p.136 / References --- p.138
3

An <i>in vitro</i> study of the susceptibilities and growth dynamics of common ocular pathogens using five fluoroquinolones

Hedlin, Peter David Skarsgard 13 September 2005
<p>Bacteria are responsible for up to 70% of all ocular infections including conjunctivitis, keratitis and endophthalmitis. If left untreated, a reduction of visual acuity, and in severe cases, sight loss, is possible. Treatment usually consists of a topically applied antibacterial preparation for patients with superficial infections. With intraocular infections, topical administration is augmented with systemic treatment or local instillation. While several types of drugs are available for ocular therapy, the fluoroquinolone class of antimicrobials is especially effective. This is due in part to their broad-spectrum of activity and low toxicity. However, as with any globally prescribed antimicrobial agent, bacterial resistance is an issue. Over the past 10 years there has been a decline in the effectiveness of older fluoroquinolones (ciprofloxacin and ofloxacin) in treating Gram-positive and, to a lesser extent, certain Gram-negative infections. In response to the declining activity of ciprofloxacin and ofloxacin, newer fluoroquinolones have been developed such as levofloxacin (L-isomer of ofloxacin), and more recently, gatifloxacin and moxifloxacin. In order to ensure the most potent drugs are being used to treat the most serious types of infection, studies need to be done to assess the activity of the current antimicrobial arsenal against pertinent infecting organisms. Three different types of experiments can be done to achieve this. In vitro potency can be tested two ways. The first is minimum inhibitory concentration (MIC). This test defines the concentration of antimicrobial drug that prevents growth of bacteria when tested against an inoculum of approximately 105 colony forming units (CFU)/ml. The second is the mutant prevention concentration (MPC), which is the amount of drug needed to inhibit a first step resistant mutant. This is a relatively new approach to measuring fluoroquinolone potency; like MIC it is not a measure of kill. A separate set of experiments are needed to assess in vitro killing. Kill curves measure the ability of an antimicrobial agent to reduce/kill a bacterial population over a period of 24 hours.</p><p>Because bacterial loads can vary greatly in in vivo infections, kill curves were conducted on a series of four inoculum sizes ranging from 106 to 109 cfu/ml. Some of the most common ocular pathogens are <i>Streptococcus pneumoniae</i>, <i>Staphylococcus aureus</i>, <i>Haemophilus influenzae</i> and <i>Pseudomonas aeruginosa</i>. <i>Mycobacterium fortuitum</i> and <i>Mycobacterium chelonae</i>, while much less commonly associated with ocular disease, are capable of causing vision-threatening infections. As a result, the above six organisms were used to test the in vitro potency of ciprofloxacin, ofloxacin, levofloxacin, moxifloxacin and gatifloxacin.</p><p>Both MIC and MPC testing found both gatifloxacin and moxifloxacin to be 4-8-fold more potent <i>in vitro</i> against the Gram-positive organisms than the older fluoroquinolones with an average potency rank order of moxifloxacin = gatifloxacin > levofloxacin > ofloxacin = ciprofloxacin. The Gram-negative results, however, revealed that the older fluoroquinolones are still the most potent of the fluoroquinolones tested with an average potency rank order of ciprofloxacin > ofloxacin = levofloxacin > gatifloxacin = moxifloxacin. Kill curve results showed a significant difference in the rate of killing between the MIC and MPC drug concentrations. At the MIC drug concentration there was generally only a noticeable reduction in viable cells following 24 hours of drug exposure and in many cases this was followed by a period of bacterial re-growth. At the MPC drug concentration, a significant bacterial count reduction was often observed as early as 4 to 6 hours for both S. pneumoniae and H. influenzae. Surprisingly, there was little difference between the five fluoroquinolones in their rates of and amount of bacterial reduction.</p><p>Because of high in vitro resistance rates in drugs like penicillin, the fluoroquinolones are an important broad-spectrum alternative. Consequently, it is imperative that measures are taken to maintain the efficacy of this class. One approach is to ensure that the most potent drug is being used to eradicate possible resistant sub-populations present in in vivo infections. The data from these experiments suggest that the new fluoroquinolones gatifloxacin and moxifloxacin are much more potent (<i>in vitro</i>) than older fluoroquinolones against Gram-positive bacteria. With Gram-negative pathogens, however, ciprofloxacin remains the most potent agent <i>in vitro</i>.</p>
4

An <i>in vitro</i> study of the susceptibilities and growth dynamics of common ocular pathogens using five fluoroquinolones

Hedlin, Peter David Skarsgard 13 September 2005 (has links)
<p>Bacteria are responsible for up to 70% of all ocular infections including conjunctivitis, keratitis and endophthalmitis. If left untreated, a reduction of visual acuity, and in severe cases, sight loss, is possible. Treatment usually consists of a topically applied antibacterial preparation for patients with superficial infections. With intraocular infections, topical administration is augmented with systemic treatment or local instillation. While several types of drugs are available for ocular therapy, the fluoroquinolone class of antimicrobials is especially effective. This is due in part to their broad-spectrum of activity and low toxicity. However, as with any globally prescribed antimicrobial agent, bacterial resistance is an issue. Over the past 10 years there has been a decline in the effectiveness of older fluoroquinolones (ciprofloxacin and ofloxacin) in treating Gram-positive and, to a lesser extent, certain Gram-negative infections. In response to the declining activity of ciprofloxacin and ofloxacin, newer fluoroquinolones have been developed such as levofloxacin (L-isomer of ofloxacin), and more recently, gatifloxacin and moxifloxacin. In order to ensure the most potent drugs are being used to treat the most serious types of infection, studies need to be done to assess the activity of the current antimicrobial arsenal against pertinent infecting organisms. Three different types of experiments can be done to achieve this. In vitro potency can be tested two ways. The first is minimum inhibitory concentration (MIC). This test defines the concentration of antimicrobial drug that prevents growth of bacteria when tested against an inoculum of approximately 105 colony forming units (CFU)/ml. The second is the mutant prevention concentration (MPC), which is the amount of drug needed to inhibit a first step resistant mutant. This is a relatively new approach to measuring fluoroquinolone potency; like MIC it is not a measure of kill. A separate set of experiments are needed to assess in vitro killing. Kill curves measure the ability of an antimicrobial agent to reduce/kill a bacterial population over a period of 24 hours.</p><p>Because bacterial loads can vary greatly in in vivo infections, kill curves were conducted on a series of four inoculum sizes ranging from 106 to 109 cfu/ml. Some of the most common ocular pathogens are <i>Streptococcus pneumoniae</i>, <i>Staphylococcus aureus</i>, <i>Haemophilus influenzae</i> and <i>Pseudomonas aeruginosa</i>. <i>Mycobacterium fortuitum</i> and <i>Mycobacterium chelonae</i>, while much less commonly associated with ocular disease, are capable of causing vision-threatening infections. As a result, the above six organisms were used to test the in vitro potency of ciprofloxacin, ofloxacin, levofloxacin, moxifloxacin and gatifloxacin.</p><p>Both MIC and MPC testing found both gatifloxacin and moxifloxacin to be 4-8-fold more potent <i>in vitro</i> against the Gram-positive organisms than the older fluoroquinolones with an average potency rank order of moxifloxacin = gatifloxacin > levofloxacin > ofloxacin = ciprofloxacin. The Gram-negative results, however, revealed that the older fluoroquinolones are still the most potent of the fluoroquinolones tested with an average potency rank order of ciprofloxacin > ofloxacin = levofloxacin > gatifloxacin = moxifloxacin. Kill curve results showed a significant difference in the rate of killing between the MIC and MPC drug concentrations. At the MIC drug concentration there was generally only a noticeable reduction in viable cells following 24 hours of drug exposure and in many cases this was followed by a period of bacterial re-growth. At the MPC drug concentration, a significant bacterial count reduction was often observed as early as 4 to 6 hours for both S. pneumoniae and H. influenzae. Surprisingly, there was little difference between the five fluoroquinolones in their rates of and amount of bacterial reduction.</p><p>Because of high in vitro resistance rates in drugs like penicillin, the fluoroquinolones are an important broad-spectrum alternative. Consequently, it is imperative that measures are taken to maintain the efficacy of this class. One approach is to ensure that the most potent drug is being used to eradicate possible resistant sub-populations present in in vivo infections. The data from these experiments suggest that the new fluoroquinolones gatifloxacin and moxifloxacin are much more potent (<i>in vitro</i>) than older fluoroquinolones against Gram-positive bacteria. With Gram-negative pathogens, however, ciprofloxacin remains the most potent agent <i>in vitro</i>.</p>
5

Caractérisation moléculaire de la résistance de F.tularensis aux fluoroquinolones / Molecular characterization of F.tularensis resistance to fluoroquinolones

Siebert, Claire 25 October 2018 (has links)
Dans des expériences récemment réalisées au laboratoire, des clones de F. tularensis ont été exposés à des concentrations croissantes d’antibiotiques. Au cours de cette expérience d’évolution expérimentale, les bactéries ont acquis un haut niveau de résistance aux FQ. Cette résistance s’est accompagnée de mutations sur la cible des FQ soient les topoisomérases de type II. Le séquençage du génome de ces souches, a montré que l’exposition aux FQ induisait également des mutations du gène fupA. Mon objectif est d’investiguer le rôle potentiel de FupA dans la résistance aux FQ à partir de deux axes : 1- Analyse phénotypique des mutants FupA : Cette analyse se fera via la complémentation des mutants des lignées d’évolution, l’évaluation des CMI et l’étude in vitro de la multiplication intracellulaire. Préalablement, les vecteurs de complémentation nécessaires seront clonés et séquencés. 2- Etude biochimique de la protéine : Cette étude consistera en la purification de la protéine recombinante suivie de la recherche des ligands potentiels. Outre des expériences de pull-down, la protéine sera utilisée pour générer des anticorps. L’étude cristallographique de cette protéine est également envisagée. / In recent experiments performed in the lab, F. tularensis strains were exposed to increasing concentrations of antibiotics. During this evolution procedure, bacteria acquired a high-level fluoroquinolone (FQ) resistance. This resistance has been associated with mutations in the FQ target genes encoding DNA gyrase and topoisomerase IV. Interestingly, the sequencing of the genomes of these strains showed that exposure to FQ also induced mutations in the gene encoding the FupA protein. My goal is to investigate the putative role of FupA in FQ resistance, an effect never previously reported. Two main approaches will be carried out. 1- phenotypic analysis of fupA mutants: This analysis will consist in complementation of mutants, evaluation of MIC and in vitro study of intracellular multiplication. Previously, the complementation vectors required will be cloned and sequenced. FQ resistance will also be evaluated on a FupA KO Francisella strain to be constructed. 2- biochemical study of the protein: We will perform cloning and purification of the full-length recombinant protein or truncated forms. Recombinant target will be useful for further identification of potential ligands by pull-down experiments. It will also be used to generate antibodies to check expression of FupA in highly resistant clones as well as for IF or EM localisation of the protein. Crystallographic study of this protein is also envisaged.
6

Risk factors for the use of macrolide and fluoroquinolone antimicrobials by human populations in Canada 2000-2006

Glass, Shiona K 27 August 2009 (has links)
Multivariable linear and negative binomial models were produced to assess relationships among socioeconomic and influenza rate data with the use of macrolide and fluoroquinolone antimicrobials by Canadians. Varying results were found both among and between the macrolide and fluoroquinolone groups; however, a pattern of accessibility to care was apparent. Cheaper antimicrobials were used most often in the most disadvantaged populations, and more expensive antimicrobials were used most frequently in advantaged populations. Significant interactions were found between influenza and socioeconomic variables relating to unemployment, education, and degree of poverty in a population. Results suggest that antimicrobials are being prescribed and consumed at inappropriate rates in both disadvantaged and affluent populations in Canada. In order to reduce antimicrobial use and the further development of antimicrobial resistance in Canada, we suggest that responsible antimicrobial stewardship be practiced and promoted by all physicians in community and hospital settings, particularly during the influenza season. / Public Health Agency of Canada
7

Evaluation of the Quality of Amiodarone with Macrolides and Fluoroquinolones Drug-Drug Interactions Reported in the Literature

Do, Brian, Patel, Pritesh, Yee, Kevin, Malone, Daniel January 2015 (has links)
Class of 2015 Abstract / Objectives: To determine the quality of evidence in the literature reporting the potential effect of QT prolongation and cardiovascular interactions of amiodarone with fluoroquinolones and macrolides. Methods: A thorough database search was conducted utilizing PubMed, Embase, Micromedex, and Facts and Comparison. Studies were eligible if they involved human subjects, original submission in English, and focusing on any drugs within the macrolide class along with amiodarone or any drugs within the fluoroquinolone class along with amiodarone was included. Drug-drug interactions (DDI) within the literature were evaluated using one of two tools: (1) van Roon to assess the quality of randomized controlled studies, and (2) the Drug Interaction Probability Scale (DIPS) to assess case reports. Results: Five case reports were included for evaluation. None of the patients within the case reports were less than 65 years old. Four of the five case reports included ciprofloxacin as part of the proposed drug interaction with amiodarone. The range of DIPS scores were 4-7 with a median score of 6. Conclusions: The evidence purporting this drug-drug interaction is of poor quality and low quantity. Additional studies of high quality must be conducted on the subject of this DDI to provide clinicians the ability to make more informed clinical decisions.
8

Approches moléculaires de l'épidémiologie de la légionellose et de la résistance aux antibiotiques chez Legionella pneumophila / Molecular approaches of the epidemiology of legionellosis and the antibiotic resistance of Legionella pneumophila

Shadoud, Lubana 17 June 2014 (has links)
Legionella pneumophila est une bactérie à Gram négatif, intracellulaire facultative, responsable de la légionellose (ou maladie des Légionnaires) chez l'Homme. Les fluoroquinolones et les macrolides sont utilisés en première intention dans le traitement antibiotique de cette maladie. Cependant, les échecs thérapeutiques sont fréquents, et le taux de mortalité demeure élevé (10-15% des cas, plus de 30% chez le patient immunodéprimé). Bien qu'aucune souche de L. pneumophila résistante à ces antibiotiques n'ait été isolée à ce jour, ces échecs peuvent faire évoquer la possibilité d'une sélection in vivo de mutants résistants. Le mécanisme génétique principal d'acquisition de la résistance aux fluoroquinolones correspond à l'accumulation de mutations au niveau des gènes codant pour l'ADN gyrase et la topoisomérase IV ; en particulier celles affectant les codons en positions 83 et 87 du QRDR (quinolone resistance determining region) du gène gyrA entrainent une résistance de haut niveau à ces antibiotiques. Le première aspect de notre projet était d'élaborer un test de PCR en temps réel permettant de détecter chez L. pneumophila des mutants gyrA résistants aux fluoroquinolones et de les différencier des souches sauvages par analyse des températures de fusion des amplifias obtenus. Après optimisation, ce test nommé qPCRgyrALp amplifie spécifiquement une portion du QRDR du gène gyrA de l'espèce L. pneumophila et permet de détecter et de différencier les mutations gyrA83 et gyrA87. Nous avons ensuite utilisé ce test pour la recherche de mutants gyrA directement dans divers prélèvements respiratoires provenant de 82 patients atteints de légionellose, certains en échec thérapeutique après traitement par une fluoroquinolone. Les résultats ont montré pour quatre patients un profil de courbe de fusion semblable à celui du mutant gyrA83. Le séquençage du QRDR de gyrA à partir de ces prélèvements respiratoires a confirmé cette mutation chez deux patients. L'utilisation de la technique de séquençage à haut débit a permis de quantifier ces mutants gyrA83 chez ces deux patients, permettant de montrer un remplacement progressif in vivo de la population de L. pneumophila sensible aux fluoroquinolones par une population résistante à ces antibiotiques. Le deuxième aspect de notre travail a été de développer des tests de PCR quantitative en temps réel (qPCR) permettant de quantifier la charge bactérienne à L. pneumophila dans les prélèvements cliniques des patients infectés, avant et au cours du traitement antibiotique, dans la but de prédire l'évolution clinique et le pronostic final de ces patients. Nous avons utilisé deux tests de qPCR, ciblant soit le gène codant pour l'ARNr16s (qPCR16S) soit le gène mip (qPCRmip) dans des prélèvements respiratoires de 116 patients atteints de légionellose. Chez certains patients, nous avons pu déterminer la cinétique de la charge bactérienne au cours du temps, alors que les patients recevaient une antibiothérapie adaptée. Les premières cinétiques recueillies montrent la possibilité de différencier les patients qui répondent rapidement au traitement antibiotique et évoluent favorablement au cours de la 1ère semaine d'hospitalisation, de ceux qui présentent une réponse modeste au traitement et nécessitent une hospitalisation prolongée, voire décèdent. La PCR en temps réel semble donc représenter un outil pronostique d'intérêt au cours de la légionellose. Le type de cinétique observé chez un patient donné semble pouvoir prédire l'évolution des patients et la nécessité d'ajuster le traitement antibiotique. / Legionella pneumophila is a Gram- negative, facultative intracellular bacterium responsible for legionellosis (or Legionnaires' disease ) in humans. The fluoroquinolones and the macrolides are used as first-line antibiotic treatment of this disease. However, treatment failures are common, and the mortality rates remain high (10-15 % of cases, more than 30% in immunocompromised patients). Although L. pneumophila strain resistant to these antibiotics have never been isolated, treatment failures may suggest the possibility of in vivo selection of resistant mutants. The main genetic mechanisms associated with acquired resistance to fluoroquinolones correspond to the accumulation of mutations in the genes encoding DNA gyrase and topoisomerase IV, especially those affecting codons 83 and 87 of the QRDR (quinolone resistance determining region) of the gyrA gene, which are associated with high level resistance to these antibiotics. The first aspect of our project was to develop a real-time PCR test to detect gyrA QRDR mutants and differentiate them from wild-type strains of L. pneumophila by analysis of melting temperatures of the amplified DNA. After optimization, the qPCRgyrALp test specifically amplified a portion of the gyrA QRDR of L. pneumophila and could detect and differentiate gyrA83 and gyrA87 mutations. Then, we checked the presence of gyrA mutants directly in respiratory samples collected in 82 legionellosis patients, including some after treatment failure with a fluoroquinolone. For four patients, results corresponded to a melting curve profile similar to that of the gyrA83 mutant. Amplification and sequencing of the gyrA QRDR directly from these respiratory samples confirmed this mutation in two patients. The use of high-throughput sequencing technology allowed us to quantify the gyrA83 mutants in these two patients, allowing demonstration of in vivo gradual replacement of the fluoroquinolones susceptible population of L. pneumophila by a resistant one. The second aspect of our work was to develop quantitative real-time PCR tests offering the possibility to quantify the L. pneumophila bacterial load in respiratory specimens before and during antibiotic treatment, in order to predict the clinical course and the final prognosis of these patients. We used two qPCR tests, either targeting the gene encoding 16S rRNA (qPCR16S ) or the mip gene (qPCRmip ) in respiratory samples from 116 patients with Legionnaires' disease. In some patients, we determined the kinetics of bacterial loads over time, while patients received appropriate antibiotic therapy. The kinetics we observed allowed differentiation of patients who respond quickly to antibiotic treatment and were released from hospital within the first week following admission, from those with a modest response to treatment and requiring prolonged hospitalization or finally died. Thus, our real-time PCR tests seem to be good prognostic tools for evaluation of legionellosis prognosis. The type of kinetics observed in a given patient may allow the clinician to predict the evolution of patients and the need to adjust the antibiotic treatment.
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Desenvolvimento e validação de métodos SPE-LC-MS e MEPS-LC-MS para quantificação de fluoroquinolonas em matrizes aquosas / Development and validation of methods SPE-LC-MS and MEPS-LC-MS for determination of fluoroquinolones in water samples

Amparo, Maura Roquete 19 April 2013 (has links)
Os antimicrobianos, especialmente a classe das fluoroquinolonas (FQs), são utilizados em grandes quantidades na medicina humana e veterinária. Uma atenção especial deve ser dada à ocorrência desses fármacos em diferentes matrizes ambientais, devido a potencialidade de propagação da resistência bacteriana. As principais fontes dessa contaminação são os esgoto industrial, urbanos, esgoto sanitário de hospital e de fazendas que utilizam antibióticos com finalidades veterinárias. Após a ingestão, os antimicrobianos são excretados na sua forma inalterada e, devido a baixa eficiência dos sistemas convencionais de tratamento de esgoto, são eventualmente liberados para o meio aquático. Diferentes métodos têm sido desenvolvidos para a determinação de FQs em amostras aquosas diversas, tais como esgoto sanitário , água de abastecimento, águas superficiais e esgoto sanitário de hospital. A maior parte dessas amostras ambientais é complexa e exige uma série de etapas de preparo, limpeza e pré-concentração; de maneira que, nos últimos anos, extensos esforços têm sido feitos para o desenvolvimento de novas técnicas de preparo de amostra que reduzam o tempo, trabalho, consumo de solvente e que permitam melhor desempenho do processo analítico. Nesse estudo foram desenvolvidos dois métodos de extração - a extração em fase sólida (SPE ) e a microextração por sorvente empacotado (MEPS) - sendo a separação, identificação e quantificação feitos por HPLC-MS/MS. Os métodos foram avaliados e validados segundo os parâmetros: precisão, exatidão, recuperação, linearidade, limite de detecção (LD), limite de quantificação (LQ), seletividade, efeito matriz, eficiência total do processo e robustez. Posteriormente, foi feita aplicação dos métodos desenvolvidos para investigação de FQs em águas superficiais e amostra de esgoto coletadas em diferentes pontos da cidade de São Carlos-SP. Os métodos apresentaram valores de recuperação maiores que 80% para as FQs estudadas, e valores de exatidão e precisão menores que 30% . A comparação entre as técnicas de extração desenvolvidas permitiu listar vantagens e desvantagens particulares de cada técnica. Além do menor consumo de solventes e volume de amostras, valores insignificantes de efeito matriz foram alcançados para a técnica MEPS; no entanto a SPE, devido ao seu maior fator de concentração, permitiu a quantificação de duas fluoroquinolonas em amostra de esgoto doméstico e detecção das mesmas em amostra de rio. / Antimicrobials, particularly the fluoroquinolones (FQs) class, are widely used in human and veterinary medicine. Particular attention must be given to the occurrence of these drugs in different environmental matrices, due to the potential spread of bacterial resistance. Effluents from industries, residential districts, hospitals and animal farms are the main sources of contamination by antibiotics. After ingestion, the antimicrobials are excreted in its unchanged form. Due to the low efficiency of conventional wastewater treatments, these antimicrobials are eventually released into the aquatic environment. Several methods have been developed for the determination of FQs in different water samples, such as municipal wastewater, tap water, river water, and hospital sewage. Most of these environmental samples is complex and requires a number of preparation steps, cleaning and preconcentration. For this reason, recently, extensive efforts have been made to develop new techniques for sample preparation in order to reduce: time, number of steps, solvent consumption and achieve better performance on the analytical process. This work describes the development of two methods of extraction - by solid phase extraction (SPE) and microextraction by packed sorbent (MEPS) - and separation, identification and quantification by HPLC-MS/MS. These methods were evaluated and validated by studying the following parameters: accuracy, precision, recovery, linearity, limit of detection (MDL), limit of quantification (MQL), selectivity, matrix effect, process efficiency and robustness. These methods were subsequently applied for FQs investigation in surface water and sewage sample collected at different points in the city of Sao Carlos/SP, Brazil. The methods recoveries achieved values greater than 80% for the studied FQS and the accuracy and precision values were satisfactory when compared to the values acceptable by regulatory agencies such as EPA and AOAC. A comparison between the extraction techniques developed allowed listing advantages and disadvantages of each particular technique. Besides the lowest solvent consumption and volume of samples, negligible values of matrix effects were achieved for MEPS technique. However, SPE, due to its higher pre-concentration, allowed the quantification of two fluoroquinolones in a sample of sewage and the detection in river sample.
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Modelagem farmacocinética populacional na avaliação do papel da glicoproteína-P na penetração tecidual de fluoroquinolonas / Population pharmacokinetic modeling on evaluation of role P-glycoprotein on fluoroquinolones tissue penetration

Zimmermann, Estevan Sonego January 2015 (has links)
Objetivos: O objetivo deste trabalho foi desenvolver modelo farmacocinético (popPK) populacional para descrever simultaneamente as concentrações das fluoroquinolonas (levofloxacino – LEV e ciprofloxacino – CIP) no plasma, pulmão e próstata na presença e ausência do inibidor da P-gp tariquidar (TAR) visando determinar a contribuição desse transportador de efluxo na distribuição tecidual desses antimicrobianos. Método: Para alcançar este objetivo as seguintes etapas foram realizadas: i) foi validado o método analítico de HPLC-fluorescência para quantificação de CIP em amostras de plasma e microdialisado; ii) foram estabelecidas as condições para microdiálise para o CIP e as taxas de recuperação in vitro, por diálise e retrodiálise, e em tecido pulmonar e prostático in vivo por retrodiálise; iii) foi avaliada a farmacocinética do LEV após administração a ratos Wistar via i.v. bolus e por nebulização intratraqueal na dose de 7mg/kg na ausência e após administração prévia de TAR (15 mg/Kg i.v.); iv) foi desenvolvido um modelo popPK para prever as concentrações do LEV simultaneamente no plasma, pulmão e próstata após administração intravenosa e intratraqueal na presença e ausência do TAR; v) foi desenvolvido o modelo popPK para descrever as concentrações de CIP simultaneamente no plasma, pulmão e próstata após administração a ratos Wistar da dose de 7 mg/kg i.v. bolus na presença e ausência de TAR (15 mg/kg i.v.); vi) Para ambos os fármacos os dados foram avaliados por análise não-compartimental e modelados por modelo de quatro compartimentos modificado, com ajuda do software NONMEN®. Resultados e Conclusões. i) Método analítico foi desenvolvido e validado com sucesso para quantificação de CIP em HPLC/fluorescência mostrando-se linear na faixa de 10–2000 ng/mL em plasma e 5–1000 ng/mL em microdialisado com coeficientes de determinação (r2) superiores a 0,99. Os valores obtidos de erro padrão relativo para ensaios de precisão intra e inter-dia foram entre 8,8 e 6,0 %, para microdialisado entre 11,1 e 7,4 % para plasma, respectivamente. Os valores de exatidão foram 86,1% entre 114.3% para microdialisado e 85,6% entre 108,2% para plasma; ii) A avaliação do CIP por microdiálise mostrou recuperação concentração independente (0,25 - 1,5 μg/mL). Além disso, não houve diferença entre as recuperações obtidas por diálise e retrodiálise para o mesmo fluxo. No fluxo selecionado para os experimentos (1,5 μL/min) as recuperações médias por diálise e retrodiálise foram 23,0 ± 2,8% e 22,8 ± 1,6 %, respectivamente. A recuperação relativa das sondas in vivo foi de 11,3 ± 1,9 e 13,1 ± 2,7 % para pulmão e próstata, respectivamente; iii) A análise dos perfis plasmáticos e teciduais LEV após dose intravenosa do grupo controle (sem TAR) mostrou boa penetração tecidual na próstata (ƒT = 0,68) e no pulmão (ƒT = 0,69). Para a mesma via de administração, o grupo TAR mostrou uma penetração praticamente inalterada para o pulmão (ƒT = 0,81) e um aumento de mais de 2 vezes na penetração prostática (ƒT= 1,64). Na dose intratraqueal houve um aumento significativo na biodisponibilidade para o grupo TAR (F = 0,86) em relação ao controle (F = 0,4). Nessa via de administração foi detectado um aumento significativo na exposição (ASC) do pulmão ao LEV no grupo TAR demonstrando que o transporte por efluxo no pulmão é mais relevante quando o fármaco é administrado pela via intratraqueal; iv) Para o LEV, o modelo popPK de quatro compartimentos foi capaz de descrever simultaneamente os dados no plasma, pulmão e próstata na presença e ausência do TAR. Além disso, o modelo para administração intravenosa foi estendido e adaptado para administração intratraqueal. Foi possível analisar o impacto do transporte por efluxo sobre a penetração tecidual do LEV por diferentes vias de administração utilizando o modelo popPK; v) A avaliação do perfil farmacocinético plasmático do CIP após administração intravenosa, na presença e ausência de TAR, demonstrou diferença significativa entre todos os parâmetros calculados por análise não-compartimental, exceto para a constante de velocidade de eliminação (= 0,05). Em relação à penetração tecidual do CIP na próstata e pulmão, não houve alteração significativa nos parâmetros de eliminação e exposição tecidual do fármaco na presença do inibidor de efluxo TAR ( = 0,05), demonstrando que o transporte por efluxo possui papel minoritário no processo de distribuição do fármaco para os tecidos estudados. O modelo popPK de quatro compartimentos foi capaz de descrever as concentrações plasmáticas totais, livres no pulmão e próstata em presença e ausência de TAR, simultaneamente; vi) O modelo popPK desenvolvido permitiu o estudo mais profundo do processo de distribuição do LEV e do CIP no pulmão e próstata. / Objectives: The aim of this study was to develop a population pharmacokinetic model (popPK) able to simultaneously describe fluoroquinolones (levofloxacin – LEV and ciprofloxacin – CIP) concentrations in plasma, lung and prostate in the presence and absence of the inhibitor of P-gp tariquidar (TAR) to determine the contribution of this efflux transporter on the tissue distribution of these antimicrobials. Methods: To achieve this goal the following steps were taken: i) An analytical method by HPLC-fluorescence was developed and validated for CIP analysis in plasma and microdialysate samples; ii) microdialysis conditions were established for CIP including determination of in vitro relative recovery by dialysis and retrodialysis. The relative recovery was also determined in vivo, in lung and prostate, by retrodialysis; iii) LEV pharmacokinetics was evaluated after intravenous (i.v.) bolus and intratracheal (i.t.) administration of 7 mg/kg dose alone and following TAR administration (15 mg/kg i.v.) to Wistar rats; iv) a popPK model was developed to describe and predict LEV concentrations in plasma, lung and prostate following i.v. and i.t. dosing with and without TAR co-administration; v) the popPK model developed was used to describe CIP concentrations in plasma, lung and prostate after i.v. bolus administration of 7 mg/kg in presence and absence of TAR; vi) For both drugs non-compartmental analysis was performed besides data modeling by four compartment model using NONMEN®. Results and Conclusions i) The analytical method was developed and successfully validated for quantification of CIP by HPLC/fluorescence. The method was linear in the range of 10-2000 ng/mL in plasma and 5-1000 ng/mL in tissues microdialysate samples with coefficients of determination (r2) higher than 0.99. The relative standard error (RSD) obtained for intra and inter-day precision were lower than 8.8% and 6.0% for microdialysate and lower than 11.1 and 7.4% for plasma, respectively. The accuracy was 86.1% to 114.3% for microdialysate and 85.6 to 108.2 % for plasma samples; ii) the evaluation of CIP microdialysis probes relative recovery in vitro showed that the recovery was concentration independent (0.25 to 1.5 μg/mL). In addition, there was no statistical difference between the recoveries determined by dialysis and retrodialysis at the same flow rate. Using the selected flow rate (1.5 μL/min) the recoveries by dialysis and retrodialysis were 23.0 ± 2.8% and 22.8 ± 1.6%, respectively. CIP relative recoveries in vivo by retrodialysis were 11.3 ± 1.9 and 13.1 ± 2.7% for lung and prostate, respectively; iii) the analysis of LEV plasma and tissues concentration-time profiles after i.v. dosing showed a good tissue penetration of LEV in the prostate (ƒT = 0.68) and lung (ƒT = 0.69). For the same route of administration, TAR group showed virtually the same penetration into lung (ƒT = 0.81) and an increase of over 2 fold in drug levels in prostate (ƒT = 1.64). For the i.t. dose, there was a significant increase on LEV bioavailability for TAR group (F = 0.86) compared to control (F = 0.4). Furthermore, a significant increase was detected on lung exposure to LEV for TAR group indicating that efflux transport in the lung is more relevant when the drug is administered by the i.t. route; iv) For LEV, a four compartment model was able to describe the data simultaneously in plasma, lung and prostate in the presence and absence of TAR. Moreover, the intravenous model was extended to adapt the intratracheal dosing route. The popPK model allowed to analyze the impact of efflux transport on tissue LEV penetration of different routes of administration; v) the evaluation of plasma CIP profiles after i.v. dosing with and without TAR showed a significant difference in all parameters determined by non-compartmental analysis in the TAR group, except the elimination rate constant (α = 0.05). The CIP tissue penetration in prostate and lung, no significant difference was observed in tissues exposure and elimination rate when TAR was present demonstrating that efflux transporter play a minor role on CIP distribution to tissues investigated (α = 0.05). The popPK model with four compartments was able to describe CIP concentrations in plasma, lung and prostate in the presence and absence of TAR, simultaneously; vi) the popPK model developed allowed a more detailed investigation of LEV and CIP distribution process in lung and prostate.

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