Antibiotics are a foundation of modern medicine, helping to save millions of lives since their discovery in 1928. But the improper and excessive use of these drugs over the last few decades has led to an alarming increase in antimicrobial resistance; coupled with the recent decrease in antibiotic discovery, it is widely thought that we are approaching a post-antibiotic era. A less well-understood problem is that of drug tolerance. Even at high doses, antibiotics often cannot kill all the bacteria in an infection because of cells that are able to tolerate antibiotic treatment. Evidence points to drug-tolerant cells, also called persisters, to be a major cause of treatment failure and chronic and recurring infections It is imperative that we develop insight and methods to prevent the spread of antimicrobial resistance and combat antimicrobial tolerance. One key effort is characterizing bacterial responses to antibiotic drug treatment to generate a more comprehensive understanding of the factors that contribute to cell death and to elucidate potential targets for new therapies. Quinolones are an important class of antibiotics that target DNA replication. They bind to topoisomerase II and IV, leading to eventual DNA fragmentation and death. However, the precise mechanism by which they work is not well understood. Because they inhibit DNA replication, quinolones lead to up-regulation of the SOS response, which allows for increased mutagenesis and the potential for increased antimicrobial resistance, thus making quinolones an interesting class of antibiotics to study. Although quinolones are one of the most effective classes of antibiotics, there are many conditions in which they do not kill, such as in stationary-phase cultures. Understanding the mechanism behind quinolone killing, quinolone-induced mutagenesis and tolerance to quinolones is important to improve quinolone efficacy. Here I have presented my work on understanding quinolones: sensitivity, mutagenesis and tolerance. In understanding quinolone sensitivity, I focus on DNA repair and its involvement in quinolone-mediated death. I then probe the field of stress-induced mutagenesis by quinolones, uncovering phenotypes of dose-dependent mutagenesis that have previously been uncharacterized. Finally, I focus on drug tolerance and how density-dependent tolerance to quinolones can be reversed by up-regulating cellular respiration through the addition of a carbon source and electron acceptor. / 2018-11-02T00:00:00Z

La magnétite, Fe3O4, est un oxyde de fer naturel à valence mixte Fe(II-III), qui sous sa forme nanométrique, a un fort potentiel d’applications technologiques dans des domaines allant de la biomédecine au traitement des eaux. Les nano-magnétites sont très efficaces pour l’adsorption ainsi que la réduction et l’oxydation de divers polluants environnementaux. Elles peuvent catalyser l’oxydation de type Fenton hétérogène induisant une dégradation efficace des polluants organiques et ceci dans un large domaine de pH. Cependant, les mécanismes impliqués restent mal connus. L’objectif principal de cette étude est d’explorer la capacité de la nano-magnétite à catalyser des réactions radicalaires de type Fenton hétérogène sans ajout d’oxydants forts, mais en utilisant uniquement l’oxygène de l’air. Ces réactions pourraient par la suite constituer la base de nouveaux procédés de remédiation efficace et éco-compatible pour l’élimination des polluants organiques dans différents compartiments de l’environnement. L’acide nalidixique, un antibiotique appartenant à la famille des quinolones, a été utilisé comme contaminant modèle, car ce composé polaire et ionisable se révèle persistant dans l’environnement et récalcitrant aux traitements classiques.Après synthèse de nano-magnétite offrant une surface spécifique élevée, la sorption de l’acide nalidixique sur ce support a été étudiée en conditions anoxiques et une adsorption supérieure à 98 % a été obtenue. En présence d’oxygène, cette sorption est suivie d’une transformation du contaminant modèle. Après désorption selon un protocole qui a été développé, un taux de dégradation d’environ 60 % a été évalué après seulement 30 minutes d’oxygénation, et 80% après 90 minutes. Cinq sous-produits de NAL ont été identifiés par chromatographie liquide couplée à la spectrométrie de masse (UHPLC-MS/MS) et un schéma de dégradation a été proposé. L’analyse de la phase solide par la diffraction des rayons X et par spectroscopie d’absorption au seuil K du fer (XANES et EXAFS) démontre une oxydation significative de la magnétite en maghémite (jusqu’à 40 %). Complétés d’une part par le suivi de la teneur en Fe(II) dissous, et des expériences réalisées en présence d’un piège à radicaux hydroxyles, et d’autres part par l’interprétation des effets du pH et de son évolution lors de la réaction, ces résultats ont permis de proposer un mécanisme réactionnel qui implique la formation des espèces réactives d’oxygène suite à l’oxydation de la magnétite.Les conclusions tirées des résultats expérimentaux prouvent les potentialités prometteuses des oxydes de fer mixte dans la remédiation des sols et eaux contaminés par des composés organiques. / Magnetite, Fe3O4, is a natural mixed iron oxide Fe(II-III), that has a wide range of applications in biomedicine as well as in water treatment. Nanosized magnetite presents high capacities to adsorb and transform a wide range of contaminants via oxidative or reductive reactions. It was shown as an active catalyst for heterogeneous Fenton reactions in the removal of organic compounds under a broad range of pH. However, the mechanisms of these reactions are not well defined.The main objective of this study was to explore the nanomagnetite capacity to catalyze heterogeneous Fenton reactions in presence of dissolved oxygen, thus avoiding the use of strong chemical oxidants. These reactions could form the basis of a new efficient and eco-friendly process for the removal of organic pollutants. Nalidixic acid (NAL), an ionizable quinolone antibiotic known for being persistent and recalcitrant to classical treatments, was used as a model contaminant.We synthesized large surface area single-cristalline nanomagnetite with high NAL sorption ability (98%) under anoxic conditions. Furthermore, a desorption protocol was developed to recover the sorbed amount of NAL in order to measure the degradation percentage.Moreover, under oxic conditions, the model contaminant was transformed, up to nearly 60% and 80 % after a 30 and 90 minutes exposure to air bubbling, respectively. Five by-products issuing from the nalidixic acid oxidative degradation were identified by liquid chromatography-mass spectrometry and a degradation pathway was suggested. X-ray powder diffraction and Iron K-edge X-ray absorption spectroscopy were used to investigate mineralogical and iron redox changes in the solid phase over the course of the reaction. Magnetite was oxidized (up to about 40%) into maghemite, -Fe2O3, as the sole product of the oxidation, and without significant change in the size of the particles. These results, in addition to the monitoring of dissolved Fe(II), and experiences conducted in the presence of ethanol as hydroxyl radicals scavenger and at static pH, lead to a better understanding of the reaction mechanism and on the role of pH in the reaction efficiency. In conclusion, this study points out the promising potentialities of mixed valence iron oxides for the treatment of contaminated soils and wastewater by organic pollutants.