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Estudo da Similaridade Genética de Amostras de Acinetobacter baumannii Produtoras de Carbapenemases do Tipo OXA Isoladas em Diversos Hospitais Brasileiros / Genetic relationship among OXA-carbapenemase producing Acinetobacter baumannii isolated from distinct Brazilian hospitalsWerneck, Jessica Sanchez de Freitas [UNIFESP] 29 September 2010 (has links) (PDF)
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Previous issue date: 2010-09-29 / Centro de Integração Empresa Escola (CIEE) / Conselho Nacional de Desenvolvimento Científico e Tecnológico (CNPq) / Fundação de Amparo à Pesquisa do Estado de São Paulo (FAPESP) / Fundação de Apoio e Desenvolvimento ao Ensino, Pesquisa e Extensão Universitária no Acre (FUNDAPE) / Nesta dissertação são apresentados dois estudos envolvendo isolados clínicos de Acinetobacter baumannii que apresentaram mecanismos enzimáticos de resistência aos carbapenêmicos, a produção de carbapenemases do tipo OXA. O primeiro estudo avalia a relação genética de amostras de A. baumannii multirresistentes produtoras de OXA-23 provenientes de distintas cidades brasileiras. 91 amostras clínicas de A. baumannii foram isoladas em 17 centros médicos localizados nas cidades de São Paulo (SP), Rio de Janeiro (RJ), Belo Horizonte (MG), Porto Alegre (RS), Blumenau (SC), Curitiba (PR), São Luiz (MA) e Salvador (BA). Nesta coleção observamos altas taxas de resistência aos carbapenêmicos (91.2%) e presença do gene blaOXA-23-like em 83,5% dos isolados. O elemento de insersão ISAba1 à montante ao gene blaOXA-23 foi detectado em todos os 76 isolados de A. baumannii produtores de OXA-23. Nove grupos de genótipos foram observados entre os 76 isolados produtores de OXA-23. Nossos achados sugerem que o gene blaOXA-23 presente nestes isolados estava localizado DNA cromossomal. Três principais grupos (A, B e D) foram observados circulando em seis cidades das regiões sudeste e sul do Brasil, sendo que o genótipo predominante (A) apresentou relação clonal àquele primeiramente descrito na cidade de Curitiba, em 2003. Em contrapartida foi encontrado um único genótipo de A. baumannii produtor de OXA-23 na cidade de São Luís. Embora existam estudos brasileiros prévios reportando a disseminação de clones de A. baumannii pordutores de OXA-23 localmente, nenhum estudo brasileiro demonstrou antes, i) a análise comparativa dos genótipos originários de diferentes e distantes cidades brasileiras, ii) a provável localização do gene blaOXA-23 e iii) a presença do elemento ISAba1 à montante ao gene blaOXA-23,o que pode ter levado ao alto grau de resistência aos carbapenens observado. Desta maneira, o estudo demonstra a circulação de clones de A. baumannii produtores de OXA-23 no Brasil e enfatiza que medidas de controle de infecção são urgentemente necessárias para reduzir tanto a disseminação de isolados multirresistenes quanto o número de infecções causadas por este patógeno. O segundo estudo trata de um relato de caso de um paciente internado em um hospital da cidade de São Paulo, que apresentou uma infecção por A. baumannii produtor de OXA-72. O isolado em questão, A 30235, apresentou resistência a todos os antimicrobianos testados, com excessão à ampicilina/sulbactam tendo apresentado redução da sensibilidade a esta associação. Foi realizada com sucesso a transformação do plasmídeo presente no isolado A30235 na cepa de A. baumannii ATCC 19606. A confirmação da presença do gene blaOXA-72 no transformante evidenciou a localização plasmidial deste determinante de resistência. O plasmídio contendo o gene blaOXA-72 apresentou um peso molecular de aproximadamente 86 Kb. Neste estudo identificamos pela primeira vez no Brasil, um isolado clínico de A. baumannii produtor de OXA-72. A presença deste determinante de resistência codificado por um gene localizado em um elemento genético móvel demonstra a crescente diversidade de carbapenemase do tipo OXA no Brasil com potencial de disseminação. Medidas de controles adequadas deverão ser tomadas para evitar a disseminação de isolados produtores de OXA-72 entre os hospitais brasileiros, o que tem ocorrido com os isolados de A. baumannii produtor de OXA-23. / In this dissertation we present two studies involving carbapenem-resistant Acinetobacter baumannii clinical isolates due to the production of carbapenems modifying enzymes, the OXA-type carbapenemases. The first study aimed to determine the genetic relationship of multi-drug-resistant A. baumannii producing OXA-23 that was isolated in distinct Brazilian cities. A total of 91 A. baumannii clinical isolates were recovered from 17 medical centers located at eight cities, namely São Paulo (SP), Rio de Janeiro (RJ), Belo Horizonte (MG), Porto Alegre (RS), Blumenau (SC), Curitiba (PR), São Luiz (MA) and Salvador (BA). In this collection we observed high rates of carbapenems resistance (91.2%). Also, the blaOXA- 23-like gene was present in 83.5% of isolates. The insertion sequence ISAba1 was positioned upstream the blaOXA-23 gene in all OXA-23-producing A. baumannii identified. Nine clusters were observed among OXA-23 producers. Our fidings suggest that the blaOXA-23 gene was probably chromosomally-located in all isolates studied. Three clusters (A, B and D) were found in six cities from southeast and southern reagions of Brazil. In addition, the predominant cluster (A) was clonally related to that first described in Curitiba, in2003. In contrast, a single cluster of A. baumannii producing OXA-23 was found in São Luís city. Although there were previous reports regarding the spread of OXA-23-producing A. baumannii in Brazil the following features had not yet been assessed: i) the comparative analysis of OXA-23 producers genotypes originating in distinct and distant Brazilian cities, ii) the genetic location of the blaOXA-23 gene and iii) the presence of ISAba1 upstream blaOXA-23 probably resulting in high degree of carbapenem resistance. Thus, our study demonstrates that the clonal dissemination of OXA-23- producing A. baumannii had occurred in Brazil. These findings emphasize that infection control measures are urgently needed to reduce both the spread of multidrug resistantstrains and the number of infections caused by this pathogen. The second study refers to a case report involving a hospitalized patient that presented an wound infection due to OXA-72-producing A. baumannii. The referred clinical isolate, A 30235, was resistant to most antibiotics tested and showed reduced susctptibility to ampicillin/sulbactam. Successful transformation assays using A. baumannii ATCC 19606 as the recipient strain revealed the plasmid location of the blaOXA-72 gene. This plasmid showed molecular weight of about 86 Kb. We identified for the first time in Brazil, an A. baumannii clinical isolate producing OXA-72. The presence of this resistance determinant encoded by a gene located in a mobile genetic elemet, points out to an increasing diversity of OXA-type carbapenemase in Brazil with potential spread. Appropriate control measures should be taken to prevent the spread of OXA-72 producers among Brazilian hospitals, which it we have experienced with OXA-23- producing-A. baumannii in this country. / TEDE / BV UNIFESP: Teses e dissertações
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Résistance aux carbapénèmes médiée par les carbapénèmases de type OXA-48 chez les entérobactéries / carbapenems resistance mediated by OXA-48-like carbapenemasesPotron, Anaïs 03 December 2013 (has links)
Les carbapénèmes constituent le traitement de dernier recours des infections associées à des germes multirésistants producteurs de -lactamases à spectre étendu. Les entérobactéries ont cependant développé des mécanismes de résistance à l’encontre de cette classe d’antibiotiques, notamment par la production de carbapénèmases. La carbapénèmase OXA-48 a rapidement disséminé en Europe et dans le pays du pourtour méditerranéen depuis 2010. Les objectifs de ce travail ont englobé, dans une première partie, la caractérisation de trois variants de la carbapénèmase OXA-48, possédant chacun des particularités phénotypiques ou génétiques. Nous nous sommes ensuite intéressés à l’épidémiologie de la carbapénèmase OXA-48 afin de comprendre ses mécanismes de dissémination puis à la variabilité de son environnement génétique. Le dernier objectif était de déterminer les facteurs génétiques à l’origine de la dissémination de la carbapénèmase OXA-48. Nous avons ainsi montré que les carbapénèmases de type OXA-48 bénéficient de tous les éléments moléculaires pour assurer leur succès : mobilisation par un transposon actif pour certains variants, transfert efficace de plasmides et dissémination clonale de souches. / Carbapenems are often the last therapeutic option for treating infections involving multiresistant ESBL-producing bacteria. Nevertheless, enterobacteria have developped resistance mechanisms toward this class of antibiotics, including carbapenemases production. Carbapenemase OXA-48 has rapidly spread throughout Europe and various countries of Mediterranean area since 2010. The aim of this work was first to characterize three variants of the carbapenemase OXA-48, each possessing phenotypic or genetic characteristics. We focused on the epidemiology of carbapenemase OXA-48 in order to understand the mechanisms of its dissemination and on the variability of its genetic environment. The last objective was to determine the genetic factors responsible for the spread of carbapenemase OXA-48. We have shown that the OXA-48-type carbapenemases possess all the molecular elements to ensure its success: mobilization by an active transposon for some variants, efficient transfer of plasmids and clonal spread of strains.
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Estudo teórico de propriedades químicas de sistemas hetero-macrocíclos que complexam metais de transição divalentes da primeira e segunda filas / Theoretical study of chemistry proprieties of the hetero-macrocycle systems that complex bivalentes transition metals of the first and second-rowLima, Francisco das Chagas Alves 06 May 2008 (has links)
Um estudo teórico detalhado das estruturas e energias do ligante 1, 7, 11, 17-tetraoxa-2, 6, 12, 16-trazaocicloocsano ([20]aneN4O4) coordenado com íons metálicos de transição Fe2+, Co2+, Ni2+, Ru2+, Rh2+ e Pd2+ foi realizado em nível de teoria B3LYP/Lanl2DZ. As geometrias dos complexos foram totalmente otimizados em simetria Cs com os íons metálicos coordenados com quatro átomos de nitrogênio (complexos 1a e 1aq) ou quatro átomos de oxigênios (complexos 1b e 1bq) e duas moléculas de água. Os arranjos octaédricos (1a e 1b) e quadrado-planares (1aq e 1bq) foram consideremos neste trabalho. A estrutura teórica está em excelente acordo com a estrutura de difração de raio-x experimental determinada para o complexo octaédrico de Ni2+ de [20]AneN4O4. Os cátions M2+ ligam-se preferencialmente aos átomos de nitrogênios com energia de ligação que aumenta na ordem Fe2+ < Ru2+ < Co2+ < Ni2+ < Rh2+ < Pd2+. Para os metais de transição da primeira fila, os complexos de spin alto são mais estáveis que os complexos de spin baixo. Em contraste, para os metais de transição da segunda fila, os estados de spin baixo mostraram-se mais estáveis que os estados de spin alto. As ligações metal-ligante nos complexos foram analisadas em termo das interações covalentes e iônicas e ajudaram a entender porque os complexos (1a e 1aq) são mais estáveis que os complexos (1b e 1bq). Os complexos poliaminas [20]aneN4 e poliéteres [20]aneO4 foram obtidos substituindo os átomos de nitrogênio e oxigênio da posição alfa dos macrociclos [20]aneN4O4 e [20]aneO4N4, respectivamente. O macrociclo [20]aneO4 tem preferência em complexar íons metálicos da primeira fila, enquanto o macrociclo [20]aneN4 prefere complexar os íons metálicos da segunda fila. / A detailed theoretical study of structures and energies of the 1,7,1l,17-tetraoxa-2,6,12,16-tetraaza-cycloeicosane ligand ([20]AneN4O4) coordinated to Fe2+, Co2+, Ni2+, Ru2+, Rh2+ and Pd2+ transition metals ions was carried out with the B3LYP/Lanl2DZ method. The geometries of the complexes were fully optimized in Cs symmetry with the metal ions coordinated either to four atoms nitrogen (complexes 1a e 1b) or to the four atoms oxygen (complexes 1aq e 1bq). The octahedral and square planar arrangements were considered in this work. The theoretical structure is in excellent agreement with the experimental X-ray diffraction structure determination for the [20]AneN4O4 octahedral Ni2+ complex. The M2+ cations bind preferentially to the nitrogen atoms with binding energies that increase in the order Fe2+ < Ru2+ < Co2+ < Ni2+ < Rh2+ < Pd2+. For the first-row transition metals, the highspin complexes are more stable than the low-spin complexes. In contrast, for the second-row of transition metals, the low-spin states were found more stable than the high spin states. The metal-ligand bonds in the complexes were analyzed in terms of the covalent and ionic interactions and helped to understand why complexes (1a e 1aq) are more stable than complexes (1b e 1bq). The polyamines [20]aneN4 and polyethers [20]aneO4 complexes were obtained substituting the atoms N or O of the alfa position of the macrocycles [20]aneN4O4 and [20]aneO4N4, respectively. The macrocycle [20]aneO4 prefers to complex first-row transition metals; however, the macrocycle [20]aneN4 prefers to complex second-row transition metals.
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Estudo teórico de propriedades químicas de sistemas hetero-macrocíclos que complexam metais de transição divalentes da primeira e segunda filas / Theoretical study of chemistry proprieties of the hetero-macrocycle systems that complex bivalentes transition metals of the first and second-rowFrancisco das Chagas Alves Lima 06 May 2008 (has links)
Um estudo teórico detalhado das estruturas e energias do ligante 1, 7, 11, 17-tetraoxa-2, 6, 12, 16-trazaocicloocsano ([20]aneN4O4) coordenado com íons metálicos de transição Fe2+, Co2+, Ni2+, Ru2+, Rh2+ e Pd2+ foi realizado em nível de teoria B3LYP/Lanl2DZ. As geometrias dos complexos foram totalmente otimizados em simetria Cs com os íons metálicos coordenados com quatro átomos de nitrogênio (complexos 1a e 1aq) ou quatro átomos de oxigênios (complexos 1b e 1bq) e duas moléculas de água. Os arranjos octaédricos (1a e 1b) e quadrado-planares (1aq e 1bq) foram consideremos neste trabalho. A estrutura teórica está em excelente acordo com a estrutura de difração de raio-x experimental determinada para o complexo octaédrico de Ni2+ de [20]AneN4O4. Os cátions M2+ ligam-se preferencialmente aos átomos de nitrogênios com energia de ligação que aumenta na ordem Fe2+ < Ru2+ < Co2+ < Ni2+ < Rh2+ < Pd2+. Para os metais de transição da primeira fila, os complexos de spin alto são mais estáveis que os complexos de spin baixo. Em contraste, para os metais de transição da segunda fila, os estados de spin baixo mostraram-se mais estáveis que os estados de spin alto. As ligações metal-ligante nos complexos foram analisadas em termo das interações covalentes e iônicas e ajudaram a entender porque os complexos (1a e 1aq) são mais estáveis que os complexos (1b e 1bq). Os complexos poliaminas [20]aneN4 e poliéteres [20]aneO4 foram obtidos substituindo os átomos de nitrogênio e oxigênio da posição alfa dos macrociclos [20]aneN4O4 e [20]aneO4N4, respectivamente. O macrociclo [20]aneO4 tem preferência em complexar íons metálicos da primeira fila, enquanto o macrociclo [20]aneN4 prefere complexar os íons metálicos da segunda fila. / A detailed theoretical study of structures and energies of the 1,7,1l,17-tetraoxa-2,6,12,16-tetraaza-cycloeicosane ligand ([20]AneN4O4) coordinated to Fe2+, Co2+, Ni2+, Ru2+, Rh2+ and Pd2+ transition metals ions was carried out with the B3LYP/Lanl2DZ method. The geometries of the complexes were fully optimized in Cs symmetry with the metal ions coordinated either to four atoms nitrogen (complexes 1a e 1b) or to the four atoms oxygen (complexes 1aq e 1bq). The octahedral and square planar arrangements were considered in this work. The theoretical structure is in excellent agreement with the experimental X-ray diffraction structure determination for the [20]AneN4O4 octahedral Ni2+ complex. The M2+ cations bind preferentially to the nitrogen atoms with binding energies that increase in the order Fe2+ < Ru2+ < Co2+ < Ni2+ < Rh2+ < Pd2+. For the first-row transition metals, the highspin complexes are more stable than the low-spin complexes. In contrast, for the second-row of transition metals, the low-spin states were found more stable than the high spin states. The metal-ligand bonds in the complexes were analyzed in terms of the covalent and ionic interactions and helped to understand why complexes (1a e 1aq) are more stable than complexes (1b e 1bq). The polyamines [20]aneN4 and polyethers [20]aneO4 complexes were obtained substituting the atoms N or O of the alfa position of the macrocycles [20]aneN4O4 and [20]aneO4N4, respectively. The macrocycle [20]aneO4 prefers to complex first-row transition metals; however, the macrocycle [20]aneN4 prefers to complex second-row transition metals.
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Le glycérol comme base structurale de coeurs de dendrimères obtenus par addition d'oxa-Michael sur des dérivés acryliques / Glycerol as structural base of dendrimer cores obtained by oxa-Michael addition to acrylic compoundsNadeau, Frédéric 13 October 2017 (has links)
Le glycérol est une molécule bio-sourcée, abondamment disponible, issue de la saponification des corps gras et de la transestérification des huiles végétales. Les travaux portent sur l'utilisation du glycérol comme base structurale de cœurs de dendrimère, en particulier par addition d'oxa-Michael sur des dérivés acryliques. La fonctionnalisation en surface de dendrimères par des motifs imidazolium est explorée afin d’obtenir un dendrimère liquide ionique (DLI) aux propriétés thermosensibles. Le chapitre bibliographique est consacré dans une première partie, aux méthodes de synthèse de dendrimère mettant en jeu des dérivés acryliques et à leurs applications et dans une seconde partie, aux travaux consacrés à l'addition oxa-Michael d'alcools sur des dérivés acryliques. Le deuxième chapitre porte sur les synthèses, à partir du glycérol, de la base structurale du cœur de dendrimère. La réaction d’acylation du glycérol par le chlorure d’acryloyle est présentée ainsi que les différentes constructions de dendrimères poly(estersulfure) à partir du triacrylate de glycérol. L’addition nucléophile du glycérol sur l’acroléine, l’acrylamide, des acrylates et l’acrylonitrile a été étudiée. Avec les acrylates, la réaction d'addition nucléophile est en compétition avec la réaction de transestérification, à l’exception de l'acrylate de t-butyle résistant en milieu basique. Avec l’acrylonitrile, la synthèse du 1,2,3-tricyanoéthylglycéryléther a pu être menée sans solvant, en 5 heures à température ambiante, en présence d’un catalyseur peu coûteux (la soude 4 mol %) avec un rendement de 88% et une pureté de 99% sans méthode de purification. Les intermédiaires de réaction mono- et di-cyanoéthylglycéryléther ont été caractérisés et ont permis un suivi cinétique de la réaction. La synthèse de dendrimères poly(amidoamine) à partir du 1,2,3-tricyanoéthylglycéryl éther fait l’objet du troisième chapitre : synthèse des générations G0 à G2,5, caractérisation des dendrimères. Des défauts de structure dus à une cyclisation intramoléculaire ont été mis en évidence par HRMS pour les générations entières et les dendrimères de demi-génération Gn+5 (n entier) sont purifiés par colonne chromatographique. Différentes voies de synthèse pour l’obtention d’un DLI à termini imidazolium sont présentées / Glycerol is a bio-based molecule, abundantly available, from the saponification of triglyceride and the transesterification of vegetable oils. The work described in this PhD thesis concerns the use of glycerol as structural base of dendrimer's core, in particular by oxa-Michael addition on acrylic derivatives. The surface functionalization of dendrimers by imidazolium units is explored in order to obtain an ionic liquid dendrimer (DLI) with thermosensitive properties. In the first part, the bibliographic chapter presents the methods of dendrimers synthesis involving acrylic derivatives and their applications and, in a second part, the work introduces the oxa-Michael addition of alcohols to acrylic derivatives. The second chapter deals with the synthesis of the structural base of the dendrimer core from glycerol. The reaction of acylation of glycerol with acryloyl chloride is presented as well as the various constructions of poly(estersulfide) dendrimers from glycerol triacrylate. The nucleophilic addition of glycerol to acrolein, acrylamide, acrylates and acrylonitrile has been studied. With the acrylates, the nucleophilic addition reaction is in competition with the transesterification reaction, with the exception of t-butyl acrylate, resistant in basic medium. With acrylonitrile, the synthesis of 1,2,3-tricyanoethylglycerylether was carried out without solvent in 5 hours at room temperature in the presence of an inexpensive catalyst (4 mol% sodium hydroxide) in a yield of 88% and purity of 99% without purification method. The mono- and di-cyanoethylglycerylether reaction intermediates were characterized and allowed kinetic monitoring of the reaction. The synthesis of poly(amidoamine) dendrimers from 1,2,3-tricyanoethylglycerylether is the subject of the third chapter: synthesis of generations G0 to G2,5 and characterization of dendrimers. The defects in the structure due to intramolecular cyclization have been demonstrated by HRMS for generations Gn and the half-generation dendrimers Gn+5 (n=0, 1, 2) were purified by chromatographic column. Several routes of synthesis for the synthesis of DLI with imidazolium termini are presented
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Achromobacter xylosoxidans : épidémiologie au centre de ressources et de compétences de la mucoviscidose de Dijon et réservoir environnemental / Achromobacter xylosoxidans : epidemiology among cystic fibrosis patients in Dijon, Burgundy and environmental reservoirAmoureux-Boyer, Lucie 12 December 2013 (has links)
Achromobacter xylosoxidans est un bacille à Gram négatif non fermentaire émergent dans la mucoviscidose. A Dijon, la prévalence de patients colonisés est élevée au Centre de Ressources et de Compétences de la Mucoviscidose par rapport à la moyenne nationale, et augmente également chez des patients hospitalisés atteints d’autres pathologies. Les réservoirs et le mode de contamination des patients sont inconnus.Les objectifs de ce travail étaient de décrire la situation épidémiologique dans notre CRCM et de mettre en évidence d’éventuelles sources de contamination pour les malades en Bourgogne.Dans une première étude rétrospective, toutes les souches d’A. xylosoxidans isolées chez les patients du CRCM depuis le début de leur suivi jusqu’en 2010 ont été analysées. Sur 120 patients, 21 ont présenté au moins une culture positive. Le génotypage par électrophorèse en champ pulsé n’a montré que de rares cas de transmissions croisées. Les résistances acquises aux antibiotiques sont fréquentes : ciprofloxacine (dès primocolonisation), ceftazidime et carbapénèmes (colonisations chroniques). Notre protocole de détection d’A. xylosoxidans dans l’environnement a permis d’isoler 50 souches (lavabos, douches, rivières, lacs) : 33 à l’hôpital, 9 à domicile et 8 dans la nature. Ces isolats partagent des caractéristiques avec les souches cliniques : 6 génotypes communs et résistance constante à la ciprofloxacine. Le risque de contamination des patients à domicile ou à l’hôpital est donc réel.D’autres études sont en cours afin de mieux comprendre l’émergence de cette bactérie, ses mécanismes de résistance acquise aux antibiotiques et les facteurs favorisant sa sélection dans l’environnement. / Achromobacter xylosoxidans is an aerobic nonfermentative Gram-Negative rod considered as an important emerging pathogen among cystic fibrosis (CF) patients worldwide. In Dijon (Burgundy), the prevalence of colonised patients is high among CF patients and increasing among non-CF hospitalised patients. The natural habitat of this organism as well as the possible sources of patient contamination remain unknown.The aims of this study were to report the first epidemiological data about A. xylosoxidans in a French CF centre and to identify potential reservoirs of this organism in Burgundy. In a retrospective study, all the isolates recovered from the patients affiliated with our centre in 2010 since their first visit were analysed. Out of 120 patients, 21 (17.5%) had at least one positive culture with A. xylosoxidans. Genotyping analysis by Pulsed Field Gel Electrophoresis revealed that cross-Contamination was very rare. Acquired resistance was frequent to ciprofloxacin (since primocolonisation), to ceftazidime and to carbapenems (in persistent colonisations). Thanks to our protocol designed to detect A. xylosoxidans in environment, a total of 50 strains were isolated in hospital (33 isolates), domestic (9 isolates) and outdoor (8 isolates) samples, mainly in handwashing sinks, showers, and water. These environmental isolates shared characteristics with clinical ones: 6 genotypes in common and a constant resistance to ciprofloxacin. These results reveal potential sources of contamination for the patients at home or in hospital. Further studies are needed to help understanding the emergence of this bacterium and the mechanisms involved in acquired antibiotic resistance.
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Role of insertion sequences in the control of antibiotic resistance in Acinetobacter baumanniiLopes, Bruno Silvester January 2012 (has links)
Acinetobacter baumannii is an emerging multiresistant pathogen increasingly known to cause infections in the immuno-compromised patients. Carbapenems and colistin are considered to be the last resorts in treatment of infections involving multidrug resistant strains of A. baumannii. Resistance to carbapenems is well known due to the presence of intrinsic carbapenemase gene blaOXA-51-like, which may be governed by insertion elements, or by acquired carbapenemases like blaOXA-23-like, blaOXA-58-like or blaOXA-40-like genes, most of which are frequently associated with insertion elements. The acquired carbapenemases can be integrated with the host chromosome making the bacterium strongly resistant to a range of antibiotics. Recent reports also suggest that the ubiquitous and intrinsic enzymes encoded by the blaOXA-51-like gene can be mobilized on a plasmid. In this thesis, the prevalence of antibiotic resistance was examined for 96 strains isolated from various parts of the world. The resistances to aminoglycosides, fluoroquinolones and cephalosporins were studied with a major focus on resistance to carbapenems. Section 1 shows the transposition of ISAba1 and its varied influence in controlling the blaOXA-51-like gene and the blaADC gene. It explains how ISAba1 being a strong factor in influencing antibiotic resistance genes contributes to the plasticity of the organism Section 2 is related with a novel insertion element ISAba125 controlling the blaADC gene and as an element providing high resistance to ceftazidime in comparison to ISAba1. Section 3 analyses the multi-drug resistant profile of strains isolated from Cochabamba, Bolivia. Besides the classification of carbapenem resistance for the clinical strains, the aminoglycoside resistance and ciprofloxacin mechanisms are examined in this project Section 4 relates with the pattern of resistance in strains isolated from the Aberdeen Royal Infirmary. It describes two novel variants of the blaOXA-51-like gene, namely blaOXA-216 and blaOXA-217 and also the acquisition of the blaOXA-23-like gene in two isolates from different years and deemed identical by their PFGE pattern. Section 5 describes the influence of ISAba825 in controlling the blaOXA-51-like gene and the blaOXA-58-like from clinical isolates Section 6 is related with the insertional inactivation of the blaOXA-132 gene and the carbapenem resistance caused by the activation of the blaOXA-58 gene in isolate Ab244 Section 7 describes the influence of insertion elements in strains having high ciprofloxacin resistance. This project is concerned with the role of efflux pump control system adeRS and how they influence the adeABC operon causing increased and decreased expression of the genes. Section 8 describes the multi drug resistant pattern of 36 strains each isolated from Europe and the United States In conclusion, there are various factors that influence the resistance profile of multidrug resistant A. baumannii isolates with insertion sequences such as ISAba1, ISAba2, ISAba3, ISAba825, IS1008, ISAba125, ISAba16 governing the expression or providing alternate mechanisms of resistance for the better fitness of the bacterium. Mutations in the genes identified in this study also have a crucial role in imparting resistance to this bacterium.
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Significance of the OXA-51-like β-lactamases of Acinetobacter baumanniiEvans, Benjamin January 2010 (has links)
The genus Acinetobacter currently contains 34 species, the vast majority of which are not regularly implicated in causing infection. However, incidences of hospitalacquired infection with Acinetobacter species are increasing, mainly due to the rise in the number of infections caused by the species Acinetobacter baumannii in immunocompromised patients particularly in intensive care units (ICUs). Due to high levels of resistance in A. baumannii to many classes of antibiotic, the carbapenems have been portrayed as the ‘drugs of choice’ for treating infections with this species. However, the activity of the carbapenems against A. baumannii has come under threat with the identification of four groups of class D β-lactamases carried by members of the species. Of these, the OXA-51-like enzymes have been suggested to be ubiquitous and intrinsic enzymes within A. baumannii. This presents the worrying scenario of the potential for all A. baumannii to become carbapenem resistant, leaving few treatment options available for this species. This project aimed to investigate the epidemiological spread of the OXA-51-like β-lactamases, examine the diversity within these enzymes, and whether this diversity may have implications for their ability to confer resistance to the carbapenems. A functional map showing the amino-acid similarities between the OXA-51-like enzymes demonstrated that the enzymes fall into distinct closely-related groups, with notable clusters surrounding OXA-66, OXA-69 and OXA-98. PCR and sequencing analysis of a geographically diverse group of 64 A. baumannii isolates demonstrated that isolates forming specific sequence groups (SGs) as defined by Turton et al (2007) also contained the same or closely related blaOXA-51-like gene. Higher minimum inhibitory concentrations (MICs) of carbapenems were found in association with acquired carbapenemases, or with the presence of ISAba1 upstream of the blaOXA-51- like gene. Pulsed-field gel electrophoresis (PFGE) analysis of the isolates did not demonstrate relatedness between isolates which formed the same sequence group. Multilocus sequence typing (MLST) of a subset of 44 isolates grouped isolates more consistently with the SGs and blaOXA-51-like alleles, indicating that PFGE is unreliable for use with A. baumannii unless studying a short time period, and that blaOXA-51-like alleles are a good epidemiological marker. Mutation studies using meropenem with five A. baumannii isolates encoding different OXA-51-like enzymes, while resulting in an increase in meropenem MICs of between 8- and 128-fold, did not result in a nucleotide substitution in the blaOXA- 51-like genes or a change in the upstream region of the genes in any isolate suggesting that the carbapenems may not be producing a strong selective pressure on the blaOXA- 51-like genes. Analysis of πN/πS ratios for the blaOXA-51-like genes, MLST genes and the TEM, SHV and CTX-M β-lactamase families showed the blaOXA-51-like genes to be under less positive selection than these other β-lactamases, though under less purifying selection than the MLST genes. Phylogenetic analysis of the MLST genes and the blaOXA-51-like genes indicates that the blaOXA-51-like genes have been evolving within A. baumannii since its speciation, and that different groups of blaOXA-51-like genes have been evolving at different rates corresponding to different rates of evolution within their parent lineages. Structural modelling studies based upon the published crystal structure for OXA-40 indicated that amino-acid variation at particular sites in the OXA-51-like enzymes are likely to have an effect of enzyme function. Alterations at amino-acid position 167 change the shape of the entrance to the active site which may affect hydrolysis by accommodating the antibiotic differently, or may affect the substrate profile of the enzyme. The substitution of glutamine for proline at position 194 may significantly alter the shape of the enzyme thereby affecting substrate hydrolysis. This project found that specific groups of blaOXA-51-like genes are associated with specific A. baumannii lineages and that these genes could serve as convenient epidemiological markers. Most of the diversity within the OXA-51-like enzymes is due to their continued evolution within A. baumannii since the species’ emergence. However, certain amino-acid changes may play a role in altering the rate of hydrolysis or substrate profile of these enzymes.
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Method Development for the Stereoselective Synthesis of Medium-Sized Cyclic Ethers and Application to Natural Product Synthesis: Part I. Organocatalytic Oxa-Conjugate Addition for α,α´-trans-Oxepanes Part II. Gold(I)-Catalyzed Alkoxylation for α,α´-cis-Oxocenes Part III. Studies toward the SynthesisLanier, Megan January 2015 (has links)
<p>Medium-sized cyclic ethers are challenging synthetic targets due to enthalpic and entropic barriers. Methods for the stereoselective synthesis of α,α΄-disubstituted medium-sized cyclic ethers began to appear with the discovery of naturally-occurring, ladder-shaped polycyclic ethers, such as brevetoxin B, and monocyclic ethers, such as (+)-laurencin. Despite the progress made in this field, limitations remain including competing formation of smaller ring sizes and scarcity of catalytic methods. Our aim has been to develop stereoselective syntheses for 7- and 8-membered cyclic ethers which have potential for application in natural product synthesis. The C-O bond disconnection was selected for the methods described within because cyclization and stereoinduction could be achieved simultaneously. In the case of 7-membered cyclic ethers, an organocatalytic oxa-conjugate addition reaction promoted by the gem-disubstituent (Thorpe−Ingold) effect has been developed to stereoselectively provide α,α′-trans-oxepanes. A gold(I)-catalyzed alkoxylation reaction has also allowed access to α,α′-cis-oxocenes. This method has been probed for feasibility in the stereoselective synthesis of (+)-intricenyne, an 8-membered cyclic ether belonging to the C15 nonterpenoid acetogenin natural product class. These methods have the potential to become general and efficient routes to highly functionalized oxepanes and oxocenes.</p> / Dissertation
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Análise conformacional e estudo das interações eletrônicas de algumas (α-oxa-α-tio-) e (α-oxa-α-seleno-) acetofenonas-para-substituídas / Conformational analysis and electronic interactions study of some (α-oxa-α-thio-) and (α-oxa-α-seleno-) acetophenones-para-substitutedTraesel, Henrique Joel 26 October 2018 (has links)
A presente tese trata da análise conformacional e estudo das interações eletrônicas de algumas 2-(fenilselanil)-2-(metoxi)-4\'-substituídas-acetofenonas (série I) e das 2-(4-substituídasfenilsulfanil)-2-(metoxi)-4\'-substituídas-acetofenonas (série II) contendo grupos doadores, hidrogênio e atraentes de elétrons na posição para dos anéis fenacílico [4\'-Y-PhC(O)] e fenilsulfanílico (S-Ph-4-X). Através da resolução das bandas de νCO, foi constatada a ocorrência de isomerismo conformacional nos solventes de constante dielétrica crescente (n-C6H14, CCl4, CHCl3, CH2Cl2, CH3CN) em ambas as séries. Cinco compostos da série II apresentaram ressonância de Fermi (RF) no infravermelho e precisaram ser deuterados. Os dados espectroscópicos são concordantes tanto com os resultados dos cálculos de otimização e frequência em fase gasosa quanto com a simulação do efeito do solvente (PCM), no nível de teoria B3LYP/6-31+G(d,p). Em ambas as séries, foram encontradas três conformações estáveis c1, c2 e c3, sendo que c2 é significativamente mais estável que c1 e c3. O confôrmero c1 possui o oxigênio metoxílico e o carbonílico em sinperiplanar enquanto os grupos -SePh/- SPh estão em anticlinal. Os confôrmeros c2 e c3 possuem o hidrogênio metínico em sinperiplanar com a carbonila enquanto os grupos -OMe e -SePh/-SPh estão na anticlinal. O confôrmero c1 pode ser atribuído ao componente de banda (IR) de maior frequência νCO. Da mesma forma, c2 e c3 são o componente de frequência intermediária e de menor frequência, respectivamente, na série I enquanto que na série II eles coalescem em apenas um componente de menor frequência. Os cálculos de PCM mimetizam os dados espectroscópicos, i.e. é possível constatar a inversão populacional entre c1 e c2, conforme a polaridade do solvente aumenta. Através do cálculo de frequências anarmônicas e construção da PED, foram encontrados dois modos vibracionais que combinados interagem com νCO originando a RF (série II). Um modo é de alta frequência e envolve uma porcentagem considerável de δC-H(metínico), o outro modo é esqueletal. A análise das interações orbitalares (NBO) mostrou que os confôrmeros c1 e c2 são estabilizados pela transferência de cargas entre LpO5→σ*CS/Se (ca. 18 kcal mol-1) enquanto c3 é estabilizado pela interação menos intensa LpO5→σ*C-C (ca. 9,5 kcal mol-1). As interações eletrônicas foram investigadas a partir dos contatos interatômicoscurtos com apoio das cargas atômicas parciais. Apesar de c1 e c2 apresentarem praticamente a mesma estabilidade em termos de interações orbitalares, o primeiro é fortemente desestabilizado pela repulsão eletrostática entre os átomos Oδ-CO...Oδ-OMe negativamente carregados separados por uma distância menor que a soma dos raios de van der Waals. Este contato curto entre os dipolos Cδ+=Oδ- e Cδ+−Oδ- é responsável pela maior frequência de νCO de c1 frente c2 e c3. A maior estabilidade de c1 nos solventes mais polares, porém, deve-se à melhor solvatação local destes dipolos. As análises dos monocristais (raios-X) indicou que os compostos assumem, no estado sólido, a conformação menos estável c1, obtida em fase gasosa. Não foram encontradas diferenças significativas no que se refere aos substituintes Y e X. Conclui-se que a estabilidade global das conformações é determinada pelo balanço energético entre as interações orbitalares e eletrostáticas, nas duas séries estudadas. / This thesis reports the conformational analysis and the electronic interactions study of some 2-(phenylselanyl)-2-(methoxy)-4\'-substituted-acetophenones (series I) and 2-(4-substitutedphenylsulphanyl)-2-(methoxy)-4\'-substituted-acetophenones (series II) bearing electron donating, hydrogen and electron attracting substituents in the para position of the phenacyl [4\'-Y-PhC(O)] and phenylsulphanyl (S-Ph-4-X). The occurrence of conformational isomerism in crescent dielectric constant solvents (n-C6H14, CCl4, CHCl3, CH2Cl2, CH3CN) in the two series was studied through the νCO infrared band decomposition. From the series II compounds, five presented Fermi resonance (FR) and had to be deuterated. The spectroscopic data are in line with theoretical calculations results for optimization and frequency in gas phase as well as for solvent effect simulation (PCM), both in B3LYP/6-31+G(d,p) level. Three stable conformers (c1, c2 and c3) were found for both series being the c2 significantly more stable than c1 and c3. Particularly in the c1 conformer, the methoxyl and the carbonyl oxygens are in sinperiplanar orientation while the -SePh/-SPh groups are in anticlinal. In the c2 and c3 conformers the methyne hydrogen assume a sinperiplanar geometry with respect to the carbonyl whereas both -OMe and -SePh/-SPh are in anticlinal. The c1 conformer can be ascribed to the highest frequency νCO band component (IR). So, c2 and c3 ones can be ascribed to the medium and the lowest frequency νCO component, respectively, in the series I whereas in the series II they come together (coalescence) in one single band component. The PCM calculations are in good agreement with spectroscopic data i.e. it is possible to observe a populational inversion between c1 and c2 conformers as the solvent polarity increases. The anharmonic frequencies and PED theoretical treatment revealed that the combination of two vibrational modes interact with the νCO giving rise to the FR (series II): the high frequency one involves a considerable δC-H(methyne) percentage; whilst the other is skeletal. The orbital interactions analysis (NBO) showed that the c1 and c2 conformers are mainly stabilized by charge transfer LpO5→σ*CS/Se (ca. 18 kcal mol-1) while c3 is stabilized by the less intense interaction LpO5→σ*C-C (ca. 9,5 kcal mol-1). The electronic interactions were investigated by means of interatomic short contacts supported by partial atomic charges calculations. In contrast with the almost similar orbital interactions stability of c1 and c2, the former is strongly destabilized by the electrostatic repulsion between Oδ-CO...Oδ-OMe atoms which are negatively charged separated by a distance shorter than the van der Waals radius sum. This short contact between Cδ+=Oδ- and Cδ+−Oδ- dipoles is responsible for the highest νCO frequency of c1 compared to c2 and c3. In the other hand, the greater stabilization of c1 in the more polar solvents is due to the better solvation of those local dipoles. Last, but not least, the X-ray analysis showed that the compounds in the solid state assume the less stable conformation c1, obtained in gas phase. No significative differences were found in terms of the Y and X substituents. It is suggestive that an energetic balance between orbital and electrostatic interactions determines the global stability of conformations in both studied series.
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