Made available in DSpace on 2015-05-14T12:56:01Z (GMT). No. of bitstreams: 1 arquivototal.pdf: 944601 bytes, checksum: c211a613a7ca088bee34c901c685111c (MD5) Previous issue date: 2011-12-16 / Coordenação de Aperfeiçoamento de Pessoal de Nível Superior - CAPES / The objective of this research was to investigate the existence of antimicrobial plant compounds and anti-quorum sensing in oral biofilms. The plants used in this study were: Schinus terebinthifolius Raddi (aroeira da praia); Lippia sidoides Scham (alecrim pimenta); Abarema cochiliacarpos (Gomes) B. & J.W. Grimes (barbatimão); Anacardium occidentale Linn (cajueiro); Psidium guajava Linn (goiabeira); Tabebuia avellanedae Lorentz ex Griseb (ipê-roxo); Hymenaea courbaril L. (jatobá) e a Bowdichia virgiloides Kunth (sucupira), Anadenanthera macrocarpa Benth (angico), Abarema cochiliacarpos (Gomes) B. & J.W. Grimes (barbatimão), Ziziphus joazeiro Mart (joazeiro), Sida cordifolia L. (malva branca), Chenopodium ambrosioides L. (mastruz), Punica granatum L. (romã) and Zingiber officinale (gengibre). The extracts were submitted in good yield to the partition with solvents in order of increasing polarity, hexane, dichloromethane and ethyl acetate. The antimicrobial activity was determined by the techniques of minimum inhibitory concentration (MIC) and minimum bactericidal concentration (MBC) and use the model QSIs (Quorum Sensing Inhibitor System) proposed by Rasmussen and colleagues (2005). The results proved promising for the development of phytomedicines. The anti-quorun sensing was significant for Abarema cochiliacarpos (Gomes) B. & JW Grimes (barbatimão), Anacardium occidentale Linn (cajueiro), Psidium guajava Linn (goiabeira) and the fruit of Hymenaea courbaril L. (Jatobá). The results support the hypothesis that extracts and partitions of some medicinal plants have anti-quorum sensing activity as a mechanism of antimicrobial activity. / O objetivo desta pesquisa foi investigar a existência de compostos vegetais que apresentem atividade antimicrobiana e anti-quorum sensing em micro-organismos formadores de biofilmes bucais. As plantas utilizadas nesta pesquisa foram: Schinus terebinthifolius Raddi (aroeira da praia); Lippia sidoides Scham (alecrim pimenta); Abarema cochiliacarpos (Gomes) B. & J.W. Grimes (barbatimão); Anacardium occidentale Linn (cajueiro); Psidium guajava Linn (goiabeira); Tabebuia avellanedae Lorentz ex Griseb (ipê-roxo); Hymenaea courbaril L. (jatobá) e a Bowdichia virgiloides Kunth (sucupira), Anadenanthera macrocarpa Benth (angico), Abarema cochiliacarpos (Gomes) B. & J.W. Grimes (barbatimão), Ziziphus joazeiro Mart (joazeiro), Sida cordifolia L. (malva branca), Chenopodium ambrosioides L. (mastruz), Punica granatum L. (romã) e Zingiber officinale (gengibre). Os extratos de Schinus terebinthifolius Raddi (aroeira da praia); Lippia sidoides Scham (alecrim pimenta); Abarema cochiliacarpos (Gomes) B. & J.W. Grimes (barbatimão); Anacardium occidentale Linn (cajueiro); Psidium guajava Linn (goiabeira); Tabebuia avellanedae Lorentz ex Griseb (ipê-roxo); Hymenaea courbaril L. (jatobá) e a Bowdichia virgiloides Kunth (sucupira), Abarema cochiliacarpos (Gomes) B. & J.W. Grimes (barbatimão), Ziziphus joazeiro Mart (joazeiro) foram submetidos à partição com solventes em ordem crescente de polaridade, hexano, diclorometano e acetato de etila. A atividade antimicrobiana foi determinada pela técnica da microdiluição em caldo e utilizou o modelo de QSIs (sistema Inibidor de Quorum Sensing) proposto por Rasmussen e colaboradores (2005). Os resultados obtidos revelaram-se promissores para o desenvolvimento de fitomedicamentos. A atividade anti-quorun sensing foi significante para Abarema cochiliacarpos (Gomes) B. & J.W. Grimes (barbatimão); Anacardium occidentale Linn (cajueiro); Psidium guajava Linn (goiabeira); e o fruto do Hymenaea courbaril L. (jatobá). Os resultados suportam a hipótese que extratos e partições de algumas plantas medicinais possuem atividade anti-quorum sensing como mecanismo de atividade antimicrobiana.

With the difficulty in treating recalcitrant infections and the growing resistance to antibiotics, new therapeutic modalities are becoming increasingly necessary. The interruption of bacterial quorum sensing (QS), or cell-cell communication is known to attenuate virulence, while limiting selective pressure toward resistance. This study initiates an ethnobotanically-directed search for QS inhibiting agents in south Florida medicinal plants. Fifty plants were screened for anti-QS activity using two biomonitor strains, Chromobacterium violaceum and Agrobacterium tumefaciens. Of these plants, six showed QS inhibition: Conocarpus erectus L. (Combretaceae), Chamaecyce hypericifolia (L.) Millsp. (Euphorbiaceae), Callistemon viminalis (Sol.ex Gaertn.) G. Don (Myrtaceae), Bucida burceras L. (Combretaceae), Tetrazygia bicolor (Mill.) Cogn. (Melastomataceae), and Quercus virginiana Mill. (Fagaceae). These plants were further examined for their effects on the QS system and virulence of Pseudomonas aeruginosa, an intractable opportunistic pathogen responsible for morbidity and mortality in the immunocompromised patient. C. erectus, B. buceras, and C. viminalis were found to significantly inhibit multiple virulence factors and biofilm formation in this organism. Each plant presented a distinct profile of effect on QS genes and signaling molecules, suggesting varying modes of action. Virulence attenuation was observed with marginal reduction of bacterial growth, suggesting quorum quenching mechanisms unrelated to static or cidal effects. Extracts of these plants were also investigated for their effects on P. aeruginosa killing of the nematode Caenorhabditis elegans. Results were evaluated in both toxin-based and infection-based assays with P. aeruginosa strains PA01 and PA14. Overall nematode mortality was reduced 50-90%. There was no indication of host toxicity, suggesting the potential for further development as anti-infectives. Using low-pressure chromatography and HPLC, two stereoisomeric ellagitannins, vescalagin and castalagin were isolated from an aqueous extract of C. erectus. Structures were confirmed via mass spectrometry and NMR spectroscopy. Both ellagitannins were shown to decrease signal production, QS gene expression, and virulence factor production in P. aeruginosa. This study introduces a potentially new therapeutic direction for the treatment of bacterial infections. In addition, this is the first report of vescalagin and castalagin being isolated from C. erectus, and the first report of ellagitannin activity on the QS system.

Le développement de salissures marines (ou biofouling) est à l’origine de nombreux problèmes économiques et écologiques. Ces salissures marines sont constituées de microorganismes (bactéries, microalgues…) formant le microfouling sur lequel va se développer le macrofouling constitué de macroorganismes tels que les algues, coquillages et éponges. La formation de ces salissures va induire un ralentissement des navires provoquant une surconsommation de carburant. De plus, l’utilisation de revêtements antisalissures ou peintures antifouling à base de métaux lourds et de biocides dans le passé a conduit à des problèmes environnementaux. L’objectif de la thèse est de rechercher une alternative aux composés actuellement utilisés (cuivre) en respectant le milieu marin. L’environnement est une source d’inspiration, une approche biomimétique pourrait être une stratégie de lutte efficace contre le biofouling. Dans une première partie, un criblage d’une centaine de composés est réalisé contre des souches bactériennes. Huit composés issus des deux familles : les batatasins et les hemibastadins sont étudiées en vue de comprendre leurs modes d’action. Parmi les composés sélectionnés, la famille des hemibastadins comprenant le DiBromoHemiBastadin-1 (DBHB) a montré des propriétés antifouling prometteuses. En effet, cette molécule est capable d’inhiber 50 % la formation du biofilm avec une IC50= 6,44 µg/mL pour la bactérie Pseudomonas aeruginosa PAO1 et une IC50 = 12,8 µg/mL pour la bactérie marine Paracoccus sp. 4M6. Afin de comprendre le mode d’action de cette molécule, son impact sur la communication bactérienne, le quorum sensing est étudié et le composé DBHB est capable de l’inhiber. Dans une seconde partie, un autre groupe d’organismes participant au microfouling est étudié : les microalgues. Afin d’évaluer l’impact de composés de la famille des hemibastadins et notamment du DBHB, l’adhésion et la formation de biofilm de microalgues sont étudiées. Le DBHB montre des inhibitions de l’adhésion et de la formation du biofilm uniquement envers la diatomée Cylindrotheca closterium. Une dernière partie, s’est intéressée à l’évaluation de revêtements contenant six composés de la famille des batatasins et des hemibastadins sur le microfouling naturel. Une méthode d’immersion de revêtements en conditions contrôlées (photobioréacteur) est mise au point afin de pallier aux contraintes environnementales. L’ensemble de cette étude a permis de mettre en évidence les propriétés antifouling du DBHB et de caractériser son mode d’action. Ce composé offre d’intéressantes voies d’étude dans la lutte contre le biofouling. De plus, l’approfondissement des connaissances sur les procédés d’adhésion et de formation de biofilm de microalgues permet de définir de nouvelles stratégies de lutte. / Biofouling induces important economic and ecological problems. This phenomenon includes microorganisms (bacteria, microalgae…) giving the microfouling which allows the macrofouling development with algae, invertebrates and sponges. These organisms colonize every immersed surface as boat hull. The colonization induces reduced speed of ships and fuel overconsumption. In the past, the utilization of AF coatings with heavy metals or pesticides caused environmental problems. The purpose of the study is to find an alternative to AF compounds (copper) respecting the marine medium. The marine environment is an inspiration; a biomimetic approach could be an interesting strategy to inhibit biofouling. In a first part, a screening of one hundred compounds is realized against marine and terrestrial bacteria. Eight molecules from two families (batatasins and hemibastadins) are studied to understand the way of action. Among selected compounds, Dibromohemibastadin-1 (DBHB) from hemibastadin family shows promising AF activities. This molecule is able to inhibit the biofilm formation with an IC50 of 6,44 µg/mL against the bacterium Pseudomonas aeruginosa PAO1 and 12,8 µg/mL for the marine bacterium Paracoccus sp. 4M6. To identify the way of action of DBHB, the impact on the bacterial communication named quorum sensing is investigated. The molecule shows an anti-quorum sensing property. In a second part, another group participating at microfouling is studied: microalgae. In order to evaluate the impact of hemibastadin family in particular DBHB, microalgae adhesion and biofilm formation are characterized. DBHB induces inhibition only on the adhesion and the biofilm formation of the diatom Cylindrotheca closterium. The last part presents the formulation of coatings containing six compounds from the batatasin and hemibastadin families. These coatings have been immersed in a harbor to evaluate the impact on natural microfouling. Furthermore, a new method for the evaluation of AF coatings is developed in controlled conditions, in a photobioreactor. This method allows the evaluation of coatings on the formation of a mixed biofilm (bacteria and microalgae). This method has been established to avoid environmental constraints by immersion in natural condition. This study allows the characterization of the AF property of DBHB. This compound provides promising research path to limit biofouling. Moreover, the development of a test allowing adhesion and microalgae biofilm formation in dynamic condition improves the characterization of compounds activities.