Pseudomonas aeruginosa est un pathogène opportuniste induisant des infections chroniques chez les patients atteints de mucoviscidose. L'éradication de ces infections est presque impossible à l'âge adulte du fait de la formation de biofilms dans les poumons des patients. Les traitements antibiotiques actuels sont peu efficaces contre les biofilms car ce mode de vie assure d'une part la protection des bactéries contre les agents anti-microbiens par l'intermédiaire de la matrice extracellulaire, et favorise d'autre part l'émergence de mécanismes de résistance. Il est donc essentiel de trouver des alternatives thérapeutiques. La bactérie marine Pseudoalteromonas sp. 3J6 sécrète une molécule à activité anti-biofilm efficace contre la souche de laboratoire P. aeruginosa PA01 et les souches cliniques P. aeruginosa MUC-N1, MUC-N2 et MUC-P4. Ces souches ont été caractérisées aux niveaux de leur formation de biofilms in vitro et de leur virulence . Ceci a montré que ces souches sont très différentes les unes des autres et qu'une seule souche, telle que la souche de laboratoire PA01, ne peut pas être représentative des profils observés. Il est donc nécessaire de mener les études anti-biofilms sur plusieurs souches, telles que celles que nous avons sélectionnées. Le potentiel thérapeutique du surnageant de culture (SNa.Js} de Pseudoalteromonas sp. 3J6 et son extrait (Ea.Js} a été étudié en évaluant leur toxicité, la réponse inflammatoire, leur impact sur la production de facteurs de virulence et leur potentiel thérapeutique . SNa.Js et Ea.Js n'étaient pas toxiques vis-à-vis des modèles testés, n'induisaient pas de réponse inflammatoire dans les poumons de souris et n'augmentaient pas la production par P. aeruginosa des facteurs de virulence quantifiés. De plus, SNa.Js s'est avèré être aussi efficace que l'antibiotique ciprofloxacine pour traiter une infection à P. aeruginosa MUC-N2 in vivo sur modèle murin. Ces résultats sont encourageants quant à un potentiel thérapeutique de la molécule à activité anti-biofilm pour contribuer au traitement des infections à P. aeruginosa chez les patients atteints de mucoviscidose. / Pseudomonas aeruginosa is an opportunistic pathogen leading to chronic infections in patients suffering of cystic fibrosis. Eradication of these infections is almost impossible in adults because of biofilm formation in patient's lungs. Current antibiotics treatments are not efficient enough against biofilms because this lifestyle first protects bacteria from antimicrobial agents via the biofilm extracellular matrix and secondly promotes the emergence of antibiotic resistance mechanisms. lt is therefore essential to find therapeutic alternatives . The marine bacterium Pseudoalteromonas sp. 3J6 secretes an anti-biofilm molecule active against the laboratory P. aeruginosa PA01 strain and the P. aeruginosa clinical strains MUC-N1, MUC-N2 and MUC­ P4. These strains were characterized at the levels of in vitro biofilm formation, and of their virulence. This part of the research work highlighted that these strains are different from one to another and that a single strain, such as the laboratory strain PA01, cannot be representative of the various patterns. Anti-biofilm studies have thus to be performed on several strains, such as the ones we selected. The therapeutic potential of the culture supernatant (SNa.Js} of Pseudoalteromonas sp. 3J6 and its extract Ea.Js was studied by evaluating their toxicity, inflammatory response, impact on virulence factors production, and therapeutic efficiency. SNa.Js and Ea.Js were not taxie against the tested models; did not induce inflammatory response in mice lungs, and did not enhance virulence factor production by clinical P. aerugonisa strains. Moreover, SNa.Js was as efficient as the ciprofloxacin antibiotic to treat an in vivo infection by P. aeruginosa MUC-N2 on mice. These results are encouraging as for a therapeutic potential of the anti-biofilm molecule to contribute at the treatment of P. aeruginosa infections of cystic fibrosis patients.

Made available in DSpace on 2017-06-01T18:20:37Z (GMT). No. of bitstreams: 1 dissertacao_jose_carlos_vilar.pdf: 1152016 bytes, checksum: 278c9de3058893d719e16b46c0048cf0 (MD5) Previous issue date: 2012-10-21 / Conselho Nacional de Desenvolvimento Científico e Tecnológico / Pseudomonas aeruginosa is an opportunistic pathogen microorganism was considered one of the most important agents of nosocomial infections due to its multiple resistance to many existing antibiotics. It is found in soil, water, plants, animals, food and various hospital settings, due to its high potential for resistance can perform processes of bioremediation. One of the biggest environmental problems generated in the activities of a textile industry is the large number of highly polluting effluents generated among the pollutants out to the black dye Remazol, which has caused serious environmental problems such as transparency and affect the aesthetics of water bodies thus preventing the penetration of sunlight, and thus the photosynthetic activity. In this regard, initial studies were performed with P. aeruginosa (UCP1567) isolated from hospital environment through the elimination of plasmids related to antibiotic resistance, using cadmium. The sample was acclimatized twice in Luria Bertani medium containing glucose, through the Minimum Inhibitory Concentration determined for cadmium (32μg/mL and 52 mg / mL, respectively) using the dilution method. The results suggest that heavy metal was effective in eliminating the resistance plasmids, corresponding to 84.61% for acclimation to cadmium 32μg/mL, and 92.30% for 52 mg / mL. Then, after eliminating the culture of resistance by cadmium, was submitted to the decolorization of Remazol black through a 23 factorial design, with independent variables agitation, the dye concentration and inoculum size, and as the response variable discoloration of the dye. The results showed that the potential for bioremediation of P. aeruginosa was not affected, and that culture after removal of microbial resistance, caused a discoloration of 85 to 94.4% of the dye, under conditions of rest, and 52% and agitation of 100 rpm and 45% under agitation at 200 rpm. / A Pseudomonas aeruginosa é um microorganismo patógeno oportunista, sendo considerado um dos mais importantes agentes de infecções hospitalares, devido a sua múltipla resistência aos diversos antibióticos existentes. É encontrada no solo, água, vegetais, animais, alimentos e em diversos ambientes hospitalares, devido ao seu elevado potencial de resistência pode realizar processos de biorremediação. Um dos maiores problemas ambientais gerados nas atividades de uma indústria têxtil é a grande quantidade de efluentes altamente poluidores gerados, dentre os poluentes destaca se o corante preto de remazol, que tem causado sérios problemas ambientais, como afetar a transparência e estética dos corpos aquáticos, impedindo assim a penetração da luz solar, e por conseguinte a atividade fotossintética. Neste sentido, foram realizados estudos iniciais com a P. aeruginosa (UCP1567) isolada de ambiente hospitalar, através da eliminação de plasmídeos relacionados à resistência aos antibióticos, utilizando o cádmio. A amostra foi aclimatada duas vezes em meio Luria Bertani contendo glicose, através da Concentração Mínima Inibitória determinada para o cádmio (32µg/mL e 52 µg/mL, respectivamente) utilizando o método de diluição. Os resultados obtidos sugeriram que o metal pesado foi eficiente na eliminação dos plasmídeos de resistência, correspondendo a 84,61% para a aclimatação com cádmio a 32µg/mL, e 92,30% para 52 µg/mL. Em seguida, a cultura após eliminação da resistência pelo cádmio, foi submetida ao processo de descoloração do preto de remazol através de um planejamento fatorial 23, tendo como variáveis independentes a agitação, a concentração do corante e o tamanho do inóculo, e como variável resposta, a descoloração do corante. Os resultados obtidos demonstraram que o potencial de biorremediação da P. aeruginosa não foi afetado, e que a cultura microbiana após eliminação de resistência, promoveu uma descoloração de 85-94,4% do corante, sob condições de repouso, e de 52% sob agitação de 100 rpm e 45% sob agitação de 200 rpm.

Submitted by Biblioteca Central (biblioteca@unicap.br) on 2017-11-01T18:01:40Z No. of bitstreams: 1 selma_neide_rodrigues_lopes_silva.pdf: 57370089 bytes, checksum: 3257bf7b1893f51caf087a9f9d294c8a (MD5) / Made available in DSpace on 2017-11-01T18:01:40Z (GMT). No. of bitstreams: 1 selma_neide_rodrigues_lopes_silva.pdf: 57370089 bytes, checksum: 3257bf7b1893f51caf087a9f9d294c8a (MD5) Previous issue date: 2010-03-04 / Surfactants are amphipathic agents with application in different industries, such as the petroleum, food and pharmaceutical. Many kinds of chemical surfactants are being used nowadays, although the development of alternative products, with biodegradable nature and lower toxicity, as the so called biosurfactants, metabolites from microorganisms, is a sound strategy for the development of compounds with ecological acceptability and for the knowledge of specific properties and application of these compounds. Different biosurfactants have been produced, but few are being commercialised due the high production costs regarded the utilisation of substrates and purification techniques. Thus, the purpose of this work is to combine the glycerol generated from biodiesel at low cost with the ability of the bacterium Pseudomonas aeruginosa UCP0992 to produce a biosurfactant with ability to remove a hydrophobic pollutant from the petroleum industry. First, the influence of glycerol concentration (2-7%), of type (NaNO3, NH4NO3, urea, (NH4)2SO4, peptone, yeast extract and corn steep liquor), and concentration (0,05-0,6%) of the nitrogen source and of the cultivation conditions (temperature – 28 and 37ºC, aeration – 60, 80 and 90% and agitation – 150 and 200 rpm) was studied for biosurfactant production by Pseudomonas aeruginosa UCP0992. The kinetic of growth and production of the biosurfactant has been described for the medium supplemented with 3% glycerol and 0.6% NaNO3, at 28 ºC during 120 hours under 200 rpm. A parallel relation between biomass, consume of glycerol, biosurfactant production, surface tension reduction and hexadecane emulsification showed a growth-associated production. The isolated biosurfactant corresponded to a yield of 8.0 g/L after 96 hours with a biomass of 4.0 g/L. The medium surface tension was reduced from 56 mN/m for 27,4 mN/ and the hexadecane emulsification remained unchanged after 72 hours, with values around 75%. The biosurfactant showed a CMC of 700 mg/L an interfacial tension against hexadecane of 2 mN/m, thermal (4-120ºC) and pH (4-12) stability regarding the surface tension reduction and the emulsification capacity of vegetable oils and diesel, and tolerance under salt concentrations (2-10%). Little changes in the surface tension and in the emulsification activity were observed when the cell-free broth containing the biosurfactant was submitted under 90°C during two hours. The biosurfactant has been characterized as a group of rhamnolipids with anionic nature. The crude biosurfactant did not show toxicity against the micro crustacean Artemia salina and the cabbage (Brassica oleracea) in the conditions tested, while the isolated biosurfactant showed toxicity against the micro crustacean depending on the concentration used. The potential application of the biosurfactant in petroleum and diesel recovery from sand was demonstrated by the percentiles of oils removal (85%). The promising results obtained in this work are noteworthy for possible biosurfactant production from glycerol. / Os surfactantes são poderosos agentes anfipáticos com aplicação nas indústrias petrolífera, alimentícia e farmacêutica, entre outras. Vários surfactantes quimicamente sintetizados são hoje utilizados, embora o desenvolvimento de produtos biodegradáveis e menos tóxicos, os chamados biossurfactantes, agentes obtidos por via microbiológica, torna-se uma estratégia importante na obtenção de componentes compatíveis com o meio ambiente. Muitos biossurfactantes têm sido produzidos, embora poucos sejam comercializados em virtude do alto custo de produção envolvido na obtenção desses compostos, principalmente no que se refere à utilização de substratos caros a e aos processos de purificação. Neste sentido, a utilização de glicerol, substrato gerado em grandes quantidades a custos cada vez mais reduzidos em função da crescente demanda mundial de biodiesel, aliada a habilidade da bactéria Pseudomonas aeruginosa UCP0992 em produzir biossurfactantes, foi avaliada para a produção de um biossurfactante com vistas à aplicação na área ambiental. Inicialmente, a influência da concentração do glicerol (2-7%), do tipo (NaNO3, NH4NO3, uréia, (NH4)2SO4, peptona, extrato de levedura e milhocina) e da concentração (0,05-0,6%) da fonte de nitrogênio e das condições de cultivo (temperatura – 28 e 37ºC, aeração – 60, 80 e 90% e agitação – 150 e 200 rpm) foram avaliadas na produção do biossurfactante por Pseudomonas aeruginosa UCP0992. A cinética de crescimento do microrganismo e de produção do biossurfactante foi descrita para o meio mineral suplementado com 3% de glicerol e 0,6% de NaNO3, a 28 ºC durante 120 horas sob agitação de 200 rpm. Uma relação paralela entre o crescimento do microrganismo, o consumo de glicerol, a obtenção do biossurfactante e a emulsificação do hexadecano foi observada, indicando uma produção associada ao crescimento. O rendimento em biossurfactante isolado após 96 horas foi de 8,0 g/L, para uma biomassa de 4,0 g/L. A tensão tensão superficial do meio foi reduzida de 56 mN/m para 27,4 mN/ e a emulsificação do hexadecando permaneceu inalterada a partir das 72 horas, com percentual de 75%. O biossurfactante apresentou CMC de 700 mg/L, tensão interfacial contra o hexadecano de 2 mN/m, estabilidade térmica (4-120ºC) e estabilidade a diferentes pH (4-12) relacionadas à capacidade de redução da tensão superficial e à atividade de emulsificação de óleos vegetais e diesel, além de tolerância a concentrações salinas (2-10%). O biossurfactante demonstrou pequenas variações na tensão superficial e na capacidade de emulsificação quando submetido a 90°C por durante dias horas. O biossurfactante foi caracterizado como um grupo de raminolipídeos de natureza aniônica. O biossurfactante bruto não apresentou toxicidade frente ao microcrustáceo Artemia salina e ao repolho (Brassica oleracea) nas condições testadas, enquanto que o biossurfactante isolado apresentou toxicidade frente ao microcrustáceo em função da concentração testada. O potencial de aplicação do biossurfactante na remoção de diesel foi demonstrado pelos elevados percentuais de remoção (85%). Os resultados promissores obtidos nesse trabalho indicam a viabilidade de produção de biossurfactantes potentes a partir de glicerol como fonte de carbono.

La mucoviscidose reste une maladie génétique grave sans traitement curatif. L’amélioration de la prise en charge de la maladie s’est accompagnée d’une augmentation importante de l’espérance de vie qui était de moins de un an dans les années 1950 et dépasse en moyenne 40 ans pour les enfants nés aujourd’hui. <p>La mucoviscidose se caractérise au niveau pulmonaire par un mucus anormalement épais qui est difficilement évacué. Le mucus s’accumule alors dans les voies respiratoires et les bactéries y persistent et s’y multiplient. 80 à 95% des patients décèdent des suites de l’insuffisance respiratoire causée par les infections chroniques bactériennes concomitantes aux inflammations des voies aériennes. Les patients souffrent particulièrement d’infections à P. aeruginosa qui est la bactérie la plus fréquemment rencontrée dans la mucoviscidose. Lors des premières infections à P. aeruginosa les bactéries sont de phénotype non mucoïde et sont relativement sensibles aux antibiotiques. Les bactéries finissent par s’adapter dans les voies aériennes des patients atteints de mucoviscidose et forment des biofilms multiresistants aux antibiotiques. Les patients sont alors colonisés par P. aeruginosa de façon permanente. Les infections chroniques à P. aeruginosa sont responsables du déclin de la fonction respiratoire, d’exacerbations pulmonaires nécessitant une hospitalisation et de la mortalité des patients atteints de mucoviscidose. Il est donc urgent d’améliorer les traitements symptomatiques existants dans le but d’augmenter la qualité et l’espérance de vie des patients en attendant de nouveaux traitements curatifs visant à corriger l’anomalie génétique.<p>Ce travail s’est donc orienté vers l’étude de l’efficacité in vitro de différentes combinaisons d’antibiotiques vis-à-vis des biofilms bactériens. Parmi l’ensemble des associations d’antibiotiques testées une synergie d’activité a été observée entre la tobramycine et la clarithromycine sur des biofilms âgés de 24 heures mais aussi de 12 jours. Notamment parmi les 26 souches testées une synergie d’action est observée chez 11 souches après une coadministration pendant 28 jours de tobramycine et clarithromycine en doses répétées sur un biofilm « mûr » âgé de 12 jours. Parmi ces 11 souches, 7 biofilms sont entièrement détruits à la fin du traitement. <p>Sur l’ensemble des souches étudiées le traitement avec l’association est soit supérieur ou équivalent à celui de la tobramycine seule et a permis d’obtenir une action dans 53.8% des cas pour la combinaison versus 30.8% pour la tobramycine.<p>Cependant aucune corrélation n’est trouvée entre le profil génétique, le phénotype mucoïde ou non des souches et la présence de synergie d’action ou la destruction entière du biofilm.<p>Cette dernière étude s’inspire du traitement actuel de référence TOBI® qui s’administre par cycle de 28 jours, 2 fois par jour. <p>Les différents travaux de cette thèse ont permis tout d’abord de développer un modèle in vitro proche des conditions rencontrées in vivo chez les patients atteints de mucoviscidose. Ce modèle pourra également être utilisé pour une série d’autres études et mises au point de nouveaux médicaments sous forme de molécule isolée ou en combinaison. <p>Ce travail a également permis d’envisager le développement d’un nouveau médicament dans le traitement d’infections chroniques à P. aeruginosa chez les patients atteints de mucoviscidose.<p>Cependant des études futures in vivo sur souris mais aussi toxicologiques et cliniques seront indispensables pour confirmer le profil d’efficacité et de sécurité de cette association.<p><p> / Doctorat en sciences pharmaceutiques / info:eu-repo/semantics/nonPublished

Pseudomonas aeruginosa PA14 est une bactérie pathogène très résistante aux antibiotiques et impliquée dans de nombreuses infections nosocomiales. Toutefois, la disponibilité d'agents antibactériens efficaces contre cette bactérie manque cruellement à ce jour. Explorer la physiologie de P. aeruginosa au niveau des modifications post-traductionnelles (PTMs) pourrait fortement contribuer au développement de nouveaux agents thérapeutiques. En effet, il a été montré certaines corrélations entre les PTMs et la virulence, l’adaptation et la résistance bactérienne. De plus, les progrès récents en protéomique ont permis d’accéder à un nombre croissant des protéines modifiées. Pourtant, leur description reste un véritable challenge.Dans une première partie, nous avons étudié l'impact des kinases et des phosphatases sur la physiologie de P. aeruginosa PA14. Cependant, aucune différence de phénotype n'a été observée entre les 8 mutants de ces enzymes et la souche sauvage.Dans une deuxième partie, nous avons caractérisé le succinylome et l'acétylome de la lysine chez P. aeruginosa PA14 dans quatre sources de carbone (glucose, citrate, succinate et glutamate) par enrichissement par anticorps couplé à la spectrométrie de masse. Ainsi, 1 530 sites succinylés (617 protéines) et 1 109 sites acétylés (526 protéines) ont été identifiés. De façon intéressante, 622 sites (312 protéines) ont été observés acétylés ou succinylés sur la même lysine, révélant ainsi l'existence de protéoformes pour une même protéine. Les protéines modifiées sont impliquées dans tous les processus biologiques. Toutefois, certaines d'entre elles ont des fonctions dans la résistance aux antibiotiques, le chimiotactisme et la virulence.Nous avons également quantifié les peptides succinylés et/ou acétylés dans les 4 sources de carbone. Les peptides succinylés étaient principalement sur-exprimés en citrate, mais aucune différence significative n’a été observée pour les peptides acétylées.Dans une troisième partie, nous avons étudié par immunoprécipitation le succinylome et l’acétylome de la lysine des protéines extracellulaire de P. aeruginosa. Nous avons montré que certaines lysines des protéines LasB et CbpD, deux facteurs de virulence, sont modifiées par 9 PTMs différentes. Une approche d’électrophorèse bi-dimensionnelle (2D) a permis de révéler et de quantifier les protéoformes des protéines extracellulaires et plus spécifiquement de ces facteurs de virulence.Dans une quatrième partie, une approche quantitative « label-free » a permis de mettre en avant 581 protéines qui varient différemment selon la source de carbone. Parmi ces protéines, 67 biomarqueurs ont été identifiés par approche statistique.Ces travaux constituent un point de départ prometteur pour de futures études sur le rôle de la succinylation de la lysine et d'autres PTMs chez P. aeruginosa. / Pseudomonas aeruginosa PA14 is a multi-drug resistant human pathogen largely involved in nosocomial infections. Unfortunately, today, effective antibacterial agents lacked. Explore its physiology at the post-translational modification (PTMs) level may contribute to the renewal of combat tactics. Indeed, some correlations between PTMs and the bacterial virulence, adaptation and resistance have been shown. The recent improvements in proteomics have increased the number of modified proteins. However, their characterization believes a real challenge.In the first part, we focused on the impact of kinases and phosphatases on bacterial physiology of P. aeruginosa PA14. For this purpose, we compared different phenotypes of 8 mutants of kinase and phosphatase with the WT strain. Unfortunately, no difference was observed.In the second part, we characterized the lysine succinylome and acetylome in P. aeruginosa PA14 in 4 carbon sources (glucose, citrate, succinate and glutamate) by mass spectrometry. Overall, a total of 1 530 succinylated sites (617 proteins) and 1 109 acetylated sites (526 proteins) were identified. Interestingly, we noticed that 622 sites (312 proteins) can be either acetylated or succinylated on the same lysine. This reveals the existence of proteoforms for a same protein. As expected, many modified proteins are involved in a wide range of biological processes but some of these proteins have interesting functions like antibiotic resistance, chemotaxis and virulence.We also tried to quantify succinylated and/or acetylated peptides in the 4 carbon sources. Succinylated peptides were mainly over-represented in the citrate condition whereas no significant difference was observed for the acetylated forms.In the third part, we investigated the lysine succinylome and acetylome of P. aeruginosa in extracellular compartment by immunoprecipitation. We showed that some lysines of two virulence factors, LasB and CbpD, were modified by 9 different PTMs. We also used a 2-dimensional gel approach to reveal and quantify proteoforms of extracellular proteins and more specifically virulence factors. In the fourth part, we did a label-free quantitative approach to obtain protein abundance in each carbon source. In total, 581 proteins vary differently depending on the carbon source. Among these proteins, 67 biomarkers were identified by statistical approach.This work is a promising starting point for further investigations on the biological role of lysine succinylation, and others PTMs, in P. aeruginosa.

This study aims to investigate the potential effects of environmental variables and the toxicity of nanosilver colloids synthesized by chemical reduction method on growth and development of phytoplankton community (the Microcystis genus dominance) in the eutrophication Tien lake water, Hanoi city, Vietnam. The variables analyzed including: physical (pH and Turbidity), chemical (content of NH4 +, PO4 3- and silver metal), biological (content of Chlorophyll-a, cell density). The characteristic of nanomaterial was confirmed by using UV-visible spectrophotometer, TEM and HR-TEM methods. The obtained silver nanoparticles (AgNPs) showed that their spherical form and uniform size varied from 10 to 15 nm. The experimental results showed that the samples treated with AgNPs inhibition on growth against M. aeruginosa at concentration 1 mg/l after 8 days. The content of silver in aquarium water decreased from 1 mg/l (D0) to 0.8 mg/l (D8). The contents of chlorophyll-a of phytoplankton community, including Microcystis genus in samples exposed with AgNPs were declined from 11.27 ± 0.56μg/L (D0) to 1.98 ± 0.37 μg/L (D8) . The environmental variables such as: pH, temperature, dissolved oxygen, turbidity, ammonium, phosphate...in the experiment were below the limit of the Vietnam Standard 08:2015/MONRE for surface water quality. / Mục đích của nghiên cứu này là khảo sát ảnh hưởng của vật liệu nano bạc tổng hợp bằng phương pháp khử hóa học đến sinh trưởng và phát triển của quần xã thực vật nổi (chủ yếu là chi Microcystis) trong nước hồ Tiền phú dưỡng, tại Hà Nội, Việt Nam. Các thông số phân tích bao gồm: thủy lý (pH và độ đục), hóa học (hàm lượng amoni, photphat và hàm lượng bạc kim loại), sinh học (hàm lượng chất diệp lục, mật độ tế bào). Đặc trưng của vật liệu được xác định bằng các phương pháp quang phổ UV-VIS, TEM và HR-TEM. Vật liệu nano bạc có dạng hình cầu, kích thước đồng nhất trong khoảng 10-15nm. Kết quả thử nghiệm sau 8 ngày cho thấy các mẫu có bổ sung vật liệu nano bạc ức chế sinh trưởng đối với vi khuẩn lam M. aeruginosa ở nồng độ 1mg/l. Hàm lượng bạc kim loại giảm từ 1 mg/l (ngày đầu tiên) xuống còn 0.8 mg/l (vào ngày cuối cùng). Sinh khối thực vật nổi trong đó có chi Microcystis trong mẫu xử lý với AgNPs đã giảm tương ứng từ 11.27 ± 0.56 μg/L (ngày đầu tiên, D0) xuống 1.98 ± 0.37 μg/L (ngày cuối cùng, D8). Các thông số môi trường của nước hồ đều nằm dưới giới hạn cho phép của QCVN 08:2015/BTNMT đối với chất lượng nước mặt.

This study aims to investigate the toxicity of copper material synthesized by chemical reduction method and effects of environmental variables on growth of phytoplankton community (dominated by Microcystis genus) in the Tien eutrophic lake, Hanoi, Vietnam. The variables analyzed include: physical (pH and Turbidity), chemical (content of NH4+, PO43- and copper metal), biological (content of Chlorophyll-a, cell density). The characteristic of nanomaterial was confirmed by using UVvisible spectrophotometer, XRD, SEM and TEM methods. The CuNPs showed they spherical form and uniform size about 20-40 nm. The experimental results showed that the treated with CuNPs inhibition on growth against phytoplankton after 8 days. The cell density of phytoplankton community and Microcystis genus in samples exposure with CuNPs declined after 8 days from 647.037 and 467.037 down to 381.111 and 202.592, respectively. / Mục đích của nghiên cứu này là khảo sát độc tính của vật liệu nano đồng được tổng hợp bằng phương pháp khử hóa học và ảnh hưởng của các yếu tố môi trường đến sinh trưởng và phát triển của quần xã thực vật nổi (chủ yếu là chi Microcystis) trong nước hồ Tiền phú dưỡng, tại Hà Nội, Việt Nam. Các thông số phân tích bao gồm: thủy lý (pH và độ đục), hóa học (hàm lượng amoni, photphat và hàm lượng đồng kim loại), sinh học (hàm lượng chất diệp lục, mật độ tế bào). Đặc trưng của vật liệu được xác định bằng các phương pháp quang phổ UV-VIS, XRD, SEM và TEM. Vật liệu nano đồng có dạng hình cầu, kích thước đồng nhất từ 20 đến 40 nm. Kết quả thử nghiệm sau 8 ngày cho thấy các mẫu có bổ sung vật liệu nano đồng ức chế sinh trưởng quần xã thực vật nổi ở nồng độ 1mg/l. Mật độ quần xã thực vật nổi và chi Microcystis trong mẫu xử lý với CuNPs đã giảm tương ứng sau 8 ngày từ 647.037 và 467.037 xuống còn 381.111 và 202.592.

Made available in DSpace on 2014-07-29T15:30:34Z (GMT). No. of bitstreams: 1 Dissertacao Diana Christina P S Goncalves.pdf: 438422 bytes, checksum: 43edf5906fb5547aa85bb981cb94aaf2 (MD5) Previous issue date: 2009-02-18 / P. aeruginosa is frequently isolated in hospitals and the clinical importance has been increased due to gravity of infections. The metallo-beta-lactamase (MBL) production is an emergent mechanism of resistance in P. aeruginosa. The study aimed to determine the antimicrobial susceptibility profile of P. aeruginosa isolated of patients admitted in a hospital in Goiânia, to verify the MBL production by diffusion test and detect MBL genes by PCR technique. A total of 75 samples were evaluated, isolated of various clinical samples, in the period of January/2005 to January/2007. The biochemical identification was performed by automation technique system (API 20E ®) and antimicrobial susceptibility profile by Kirby- Bauer method. The 75 P. aeruginosa presented multi-drug resistance and, the resistance profile was: 90.7% to ceftazidime: 30.7% to aztreonam, 97.3% to ciprofloxacin; 48.0% of resistance to piperacilin/tazobactam, 88.0% to cefepime; amicacin, gentamicin and tobramicina whit resistance profile of 78.7%, 84.0% and 77.4%, respectively. The MBL production by difusion disc method was 46.7% (35/75). The gene blaSPM-1 was detected in 39 (52.0%) and gene blaIMP-1 in three (4.0%) isolates. The high frequency of P. aeruginosa resistant and MBL production alert to necessity of control the dissemination of bacteria multi-drug resistant in hospital, as well as the adoption of preventive actions and explanation of the health workers about rational use of antibiotics. / P. aeruginosa é frequentemente isolada em ambientes hospitalares e sua importância clínica têm aumentado devido à gravidade das infecções. A produção de metalo-beta-lactamase (MBL) é um mecanismo de resistência emergente entre P. aeruginosa. O estudo teve como objetivo determinar o perfil de suscetibilidade antimicrobiana de P. aeruginosa isoladas de pacientes internados em um hospital de Goiânia, realizar a triagem fenotípica para verificar a produção de MBL e detectar genes que codificam MBL pela técnica de PCR. Foram avaliadas 75 amostras, isoladas de diversos sítios, no período de janeiro de 2005 a janeiro de 2007. A identificação bioquímica foi realizada pelo sistema API 20E e o antibiograma pelo método de Kirby-Bauer. Todos os 75 isolados de P. aeruginosa apresentaram multirresistência, 82,7% foram resistentes ao imipenem; ceftazidima 90,7%; aztreonam 30,7%; ciprofloxacina 97,3%; 48,0% de resistência a piperacilina/tazobactam, 88,0% ao cefepime; amicacina, gentamicina e tobramicina, com resistência de 78,7%, 84,0% e 77,4%, respectivamente. A produção de MBL pelo método de disco aproximação foi detectada em 46,7% (35/75). O gene blaSPM-1 foi detectado em 39 (52,0%) e o blaIMP-1 em três (4,0%) amostras através da técnica de PCR. A frequência elevada de P. aeruginosa multirresistentes e produtoras de MBL alerta para necessidade de controle da disseminação de resistência no ambiente hospitalar, bem como a adoção de medidas preventivas e esclarecimento das equipes de saúde sobre uso racional dos antimicrobianos.

Indiana University-Purdue University Indianapolis (IUPUI) / Pseudomonas aeruginosa is a gram negative opportunistic pathogen with the capacity to cause serious disease by forming biofilms, most notably in the lungs of cystic fibrosis (CF) patients. Biofilms are communities of microorganisms that adhere to a solid surface, undergo global regulatory changes, secrete exopolysaccharides, and are innately antibiotic resistant. Virulence modulation is an important tool utilized by P. aeruginosa to propagate infection and biofilm formation in the CF airway. Many different virulence modulatory pathways and proteins have been identified including the protein, MgtE. MgtE has recently been discovered and has been implicated in virulence modulation, as an isogeneic mutation of mgtE leads to increased cytotoxicity. To further elucidate the role of MgtE in P. aerugionsa infections, transcriptional and translational regulation of this protein following antibiotic treatment has been explored. I have demonstrated that mgtE is transcriptionally upregulated following antibiotic treatment of most of the twelve antibiotics tested utilizing RT-PCR and QRT-PCR. A novel model system was employed, which utilizes cystic fibrosis bronchial epithelial (CFBE) cells homozygous for the ΔF508 mutation for these studies. This model system allows P. aeruginosa biofilms to form on CFBE cells modeling the P. aeruginosa in the CF airway. Translational effects of antibiotic treatment on MgtE have been attempted via Western blotting and cytotoxicity assays. Furthermore, to explore the possibility that mgtE is interacting with a known regulatory pathway, a transposon-mutant library was utilized and the regulatory proteins, AlgR and NarX, among others have been identified as possibly interacting with MgtE. Lastly, an MgtE homologue from Staphylococcus aureus was utilized to further demonstrate the virulence modulatory effects of MgtE by demonstrating the expression of the homologue results in decreased cytotoxicity, exactly like expression of the native P. aeruginosa MgtE. This research explores a newly discovered protein that impacts cytotoxicity and biofilm formation and provides valuable information about P. aeruginosa virulence.

La pyocyanine (PYO) est une phénazine de couleur bleu-vert, produite spécifiquement par la bactérie pathogène opportuniste Pseudomonas aeruginosa (Pa). La toxicité aérobie de la PYO envers les cellules de mammifères, les levures et les bactéries a été décrite de longue date, mais la compréhension des mécanismes d’action est encore lacunaire, en particulier en conditions de limitation en O2 (conditions rencontrées dans le contexte infectieux). De plus, il a récemment été montré que la PYO peut apporter des effets bénéfiques pour la souche productrice en hypoxie. Au cours de ce travail, nous avons réexaminé les effets de la PYO sur un large panel de bactéries dont son propre producteur (Pa) ainsi que sur un modèle cellulaire eucaryote Saccharomyces cerevisiae exposées à différentes tensions en O2. Nos données suggèrent que la toxicité aérobie de la PYO envers S. cerevisiae est multifactorielle, impliquant à la fois une interaction avec le complexe III de la chaîne respiratoire et l’induction d’un stress oxydatif. Pour la première fois, nous avons mis en évidence une toxicité de la PYO exacerbée en anaérobiose chez un eucaryote (S. cerevisiae). Le mécanisme d’action impliquerait le PYO radical. Nous avons également montré que la PYO peut inhiber la croissance aérobie et anaérobie des microorganismes concurrents, plus particulièrement S. aureus en bloquant le complexe III de la chaîne respiratoire. A l’inverse, la PYO peut stimuler la respiration de Pa surtout dans les conditions mimant le contexte infectieux (hypoxie, vie ralentie). Le complexe III et/ou les oxydases terminales cbb3 serait impliqué favorablement. En conclusion, la PYO jouerait à la fois un rôle de poison hypoxique mais aussi un rôle de navette redox bénéfique pour la survie et la virulence de Pa en hypoxie. / Pyocyanin (PYO) is a blue-green phenazin, specifically produced by the opportunistic bacterium Pseudomonas aeruginosa (Pa). Aerobic toxicity of PYO toward mammalian cells, yeast and bacteria has been known for a long time, but the understanding of its mechanisms of action remains unclear, especially in conditions of limited O2 (conditions encountered during infection). In addition, it has recently been shown that PYO can bring benefits to the producer strain under hypoxia. In this study, we reexamined the effects of PYO toward a large panel of bacteria including its own producer Pa as well as a model of eukaryotic cells Saccharomyces cerevisiae exposed to different oxygen tensions. Our results suggest that the aerobic toxicity of PYO toward S. cerevisiae is multifactorial: involving both interaction with the respiratory chain at the level of complex III and induction of oxidative stress. For the first time, we have shown that PYO exerts an increased toxicity toward the eukaryotic cell, S. cerevisiae under anaerobiosis. The mechanism could involve the production of PYO radical. We have also shown that PYO can inhibit the aerobic and anaerobic growth of competing microorganisms, especially S. aureus by blocking the complex III of the respiratory chain. Conversely, PYO can stimulate the respiration of Pa, in mainly in conditions similar to those encountered during infection (hypoxia, slowed growth). The complex III and/or the cbb3 oxidases could be favorably involved. To conclude, PYO could act as a hypoxic poison as well as a redox shuttle beneficial for the survival and the virulence of Pa under hypoxia.