The cold saline springs at Gypsum Hill (GH) and Colour Peak (CP) in the Canadian high Arctic are rare examples of perennial flows in thick continuous permafrost. We surveyed the microbial diversity of the springs by analyzing clone libraries of the small subunit rRNA gene. Half of the bacterial clones from the GH library classified as Delta- and Gammaproteobacteria. Clones related to Proteobacteria (82%), Firmicutes (9%) and Bacteroidetes (6%) constituted 97% of the bacterial library from CP. At least 56 and 76% of the bacterial clones from GH and CP, respectively, were from putative sulfur-metabolizers, and clones related to the sulfur-oxidizing bacterium Thiomicrospira psychrophila dominated in both springs. Archaeal clones were affiliated with uncultured Crenarchaeota, methanogens and haloarchaea. The eukaryotic clones were related to known Fungi, Viridiplantae (green algae), Strametopila (e.g. diatoms), Cercozoa and Alveolata in the CP sediment. Eukaryotic rRNA genes could not be amplified from the GH sediments. Forty-nine phylotypes of heterotrophic and sulfur-oxidizing bacteria (Thiomicrospira, Halothiobacillus ) were isolated from the GH springs. The strains were predominantly psychrotrophic and halotolerant, and were most related to bacteria also isolated from permanently cold environments. Some heterotrophic strains possessed genes for photosynthesis and thiosulfate oxidation, possibly enabling them to better compete in these sulfur-rich ecosystems exposed to continuous light in the summer. Assays of leucine and CO2 incorporation showed a low heterotrophic activity in the GH spring water, but significant autotrophic activity in complete darkness (chemoautotrophy). Microbial sulfur metabolism was demonstrated in microcosms of the GH sediment. Sulfur oxidation and chemoautotrophy were also demonstrated in microbial filaments that thrive in the GH spring channels. Fluorescence in situ hybridization (FISH) showed that the filaments were almost exclusively composed of rod-shaped Gammaproteobacteria that were further identified as Thiomicrospira. This work describes active microbial communities capable of sustainability in extreme environments that combine low temperature, moderate salinity, and prolonged periods of continuous light or darkness. Sulfur oxidation seems to be a major energetic process and chemolithoautotrophy appears to contribute significantly to primary production. Similar hydrosystems, formed by brines in the cold permafrost, are hypothesized to exist on Mars.

Inflammatory bowel disease (IBD), comprises Crohn’s disease (CD) and ulcerative colitis (UC), and is a chronic relapsing inflammation of gastrointestinal tract without any known cause or cure. Currently, it is accepted that IBD is a result of a dysfunctional immune response to commensal bacteria in a genetically susceptible host, and that environmental factors can trigger the onset or reactivation of the disease. This thesis considers the possibility of a specific pathogenic agent as well as an imbalance in the composition of the normal microflora in the pathogenesis of IBD. Gut biopsy tissues were taken from a population-based case-control tissue bank held at the University of Manitoba. Automated ribosomal intergenic spacer analysis (ARISA) and terminal restriction fragment length polymorphisms (T-RFLP) were employed to assess the diversity of gut microbiota. The phylogenetic, virulence and biochemical characteristics of Escherichia coli isolated from IBD biopsies were examined using multi-locus sequence typing (MLST), DNA microarray technology and API 20E system. Utilizing ARISA and T-RFLP, a remarkable increase in the order of unclassified Clostridia was detected in inflamed tissues, particularly in CD patients (P < 0.05). Moreover, species richness and diversity were the highest in non-inflamed IBD biopsies. Culture-based quantification detected a significantly higher number of E. coli in IBD tissues (P < 0.05). Phylogenetic analysis revealed the tendency of E. coli isolated from IBD patients to be grouped into separate clonal clusters based on their allelic profiles (P = 0.02). A link was detected between uropathogenic E. coli (UPEC) CFT073 and strains isolated from IBD, with regards to gene distribution and virulence, using microarray technology. Amino acid substitutions N91S and S99N in FimH, the adhesive subunit of E. coli type I fimbria, were significantly associated to IBD (P < 0.05). This study demonstrated an increase in the microbial diversity of non-inflamed IBD tissues and suggested a recruitment phase of bacterial adherence and colonization, before the inflammation sets in. Furthermore, E. coli isolated from IBD tissues were distinct from commensal strains in both clonal and virulence characteristics and shared remarkable traits with extraintestinal pathogenic E. coli. Features involved in bacterial adhesion to epithelial cells may hold the key to E. coli pathogenesis in IBD.

Beim Bergbau werden bestehende Ökosysteme in großem Maße beeinflusst. Im ehemaligen Uranbergwerk Königstein (Sachsen) wurde die Umwelt durch den Einsatz von chemischen Säuren zur Lösung des Urans aus dem Erz (Laugung) in Folge der Verschiebung des pH-Wertes zusätzlich belastet. Durch diesen Prozess entstand eine Umgebung, die einen niedrigen pH-Wert und hohe Konzentrationen an gelösten Schwermetall-Ionen aufweist. Die komplexe mikrobielle Lebensgemeinschaft verschob sich daraufhin, indem sich bevorzugt säuretolerante und Schwermetall-tolerante Mikroorganismen durchsetzten. Diese Mikroorganismen wurden durch die Flutung der unter Tage Schächte im Jahr 2010 in ihrer Zusammensetzung erneut beeinflusst. In dieser Arbeit wurde die mikrobielle Biozönose nach Flutung der unter Tage Schächte des ehemaligen Uranbergwerkes Königstein charakterisiert und mit den Ergebnissen der mikrobiellen Diversität vor dem Flutungsprozess verglichen. Hierfür kam ein breites Spektrum an Methoden zum Einsatz, das klassische mikrobiologische Methoden und molekularbiologische Techniken umfasste. Die Analysen erfolgten dabei über mehrere Jahre hinweg, um die Variabilität der mikrobiellen Population im Grubenwasser planktonisch und im Biofilm zu erfassen.

Inflammatory bowel disease (IBD), comprises Crohn’s disease (CD) and ulcerative colitis (UC), and is a chronic relapsing inflammation of gastrointestinal tract without any known cause or cure. Currently, it is accepted that IBD is a result of a dysfunctional immune response to commensal bacteria in a genetically susceptible host, and that environmental factors can trigger the onset or reactivation of the disease. This thesis considers the possibility of a specific pathogenic agent as well as an imbalance in the composition of the normal microflora in the pathogenesis of IBD. Gut biopsy tissues were taken from a population-based case-control tissue bank held at the University of Manitoba. Automated ribosomal intergenic spacer analysis (ARISA) and terminal restriction fragment length polymorphisms (T-RFLP) were employed to assess the diversity of gut microbiota. The phylogenetic, virulence and biochemical characteristics of Escherichia coli isolated from IBD biopsies were examined using multi-locus sequence typing (MLST), DNA microarray technology and API 20E system. Utilizing ARISA and T-RFLP, a remarkable increase in the order of unclassified Clostridia was detected in inflamed tissues, particularly in CD patients (P < 0.05). Moreover, species richness and diversity were the highest in non-inflamed IBD biopsies. Culture-based quantification detected a significantly higher number of E. coli in IBD tissues (P < 0.05). Phylogenetic analysis revealed the tendency of E. coli isolated from IBD patients to be grouped into separate clonal clusters based on their allelic profiles (P = 0.02). A link was detected between uropathogenic E. coli (UPEC) CFT073 and strains isolated from IBD, with regards to gene distribution and virulence, using microarray technology. Amino acid substitutions N91S and S99N in FimH, the adhesive subunit of E. coli type I fimbria, were significantly associated to IBD (P < 0.05). This study demonstrated an increase in the microbial diversity of non-inflamed IBD tissues and suggested a recruitment phase of bacterial adherence and colonization, before the inflammation sets in. Furthermore, E. coli isolated from IBD tissues were distinct from commensal strains in both clonal and virulence characteristics and shared remarkable traits with extraintestinal pathogenic E. coli. Features involved in bacterial adhesion to epithelial cells may hold the key to E. coli pathogenesis in IBD.

Brazil occupies a place of prominence in the productive sector, as well as in the exploitation of agricultural waste, from sugar cane. This use is a fairly widespread practice, both for effluents, mainly vinasse, but also for solid residues, such as filter-cake and bagasse from sugar cane. Thus, at the start of the 2005/06 harvest, samples were collected from residual waters of the S.T.E. (Station for the treatment of effluents), bagasse and from the filtrate liquid (percolate) from bagasse that was accumulated since the 2004/05 harvest, from the industrial processing of sugar cane in S.A. Usina Coruripe Açúcar e Álcool . The target was the isolation of microorganisms that produce extracellular enzymes able to degrade cellulose, hemicellulose, and lignin phenols. There were originally 42 microorganisms isolated, of which 31 were screened for the verification of the production of these and other enzymes, as well as for their morphobiochemistry identification. From these, 29 were identified, with the predominance of the genera Flavobacterium, Chromobaterium and Achromobacter. The other bacterial isolates belong to the genus Corynebacterium, Aeromonas., Bacillus, Clostridium, Citrobacter, Nocardia, Kurthia, Mycobacterium, Serratia, Pseudomonas, e Actinomyces. Among the fungal isolates, the genera detected were: Penicillium, Rhodotorula, Gonatobotrys and Gliocladium. The actynomicete Nocardia (PB4), isolated from the percolate of bagasse, presented a broad spectrum of enzymatic activities, being selected for evaluation of its cellulolitic activity in liquid medium containing carboxymethylcellulose and bagasse from sugar cane as a source of carbon. The production of cellulase in these substrates was evaluated according to the concentration of reducing sugars and of total protein, and the content of total phenols has also been determined. In addition to this isolated, another 5 microorganisms were chosen - because of their cellulolytic, xylanolytic, and phenolytic activities in a solid medium, and used for implementing a process of composting of the sugar cane bagasse (from 2005/06 and 2006/07 harvests) with filter-cake (2006/07 crop), in scale of the greenhouse for 48 days. These were divided into 2 consortia (CM1 and CM2), formed by 3 microorganisms each. Factors such as temperature, moisture, organic matter content, nitrate, nitrite, phosphate and organic carbon were evaluated in order to monitor the process of composting substrates, without the addition of other macro and micronutrients. From the physico-chemical analyses, it was found that there were not wide variations in the performance of both consortia, and that both led to a reduction of the concentration of organic matter and contents of nitrate and phosphate. Besides this, in period (48 days) and in the conditions (trays with large diameter and small height, which favors changes of temperature of the substrate with the environment; non addition of other sources of nitrogen and phosphorus) of this study, the substrates do not reached stabilization. However, the microorganisms presented here, have broad potential for exploitation in cases of composting the solid and liquid agri-industrial residues, provided that relations C:N be corrected, and also in bioremediation, because of its broad spectrum of enzymatic activities. It is suggested to combine the solid waste with a percentage of vinasse, because this is rich in nutrients that influence the action of microorganisms studied. / Fundação de Amparo a Pesquisa do Estado de Alagoas / O Brasil ocupa um lugar de destaque tanto no setor produtivo quanto no aproveitamento agrícola dos resíduos da cana-de-açúcar. Este aproveitamento constitui-se numa prática bastante generalizada, tanto para os efluentes, principalmente a vinhaça, como também para os descartes sólidos, como a torta de filtro e o bagaço de cana. Deste modo, no início da safra 2005/06, foram coletadas amostras de águas resíduárias da E.T.E. (estação de tratamento de efluentes), bagaço de cana e percolado desse bagaço (acumulado desde a safra 2004/05) oriundos do processamento industrial da cana-deaçúcar na S.A. Usina Coruripe Açúcar e Álcool, visando o isolamento de microrganismos produtores de enzimas extracelulares capazes de degradar celulose, hemicelulose, lignina e fenóis. Foram inicialmente isolados 42 microrganismos, dos quais 31 foram triados para a verificação da produção dessas e outras enzimas, bem como para sua identificação morfo-bioquímica. Destes, 29 microrganismos foram identificados, havendo a predominância dos gêneros Flavobacterium, Chromobaterium e Achromobacter. Os demais isolados bacterianos pertencem aos gêneros Corynebacterium, Aeromonas, Bacillus, Clostridium, Citrobacter, Nocardia, Kurthia, Mycobacterium, Serratia, Pseudomonas, e Actinomyces. Entre os isolados fúngicos, os gêneros detectados foram: Penicillium, Rhodotorula, Gonatobotrys e Gliocladium. O actinomiceto Nocardia (PB4), oriundo do percolado do bagaço, apresentou um amplo espectro de atividades enzimáticas, sendo selecionado para avaliação de sua atividade celulolítica em meios líquidos contendo carboximetilcelulose e bagaço de cana como fonte de carbono. Avaliou-se a produção de celulase a partir destes substratos, bem como a liberação de glicídios redutores, proteínas totais e fenóis totais. Além deste isolado, outros 5 microrganismos foram escolhidos, devido ao seu potencial em degradar celulose, xilana, lignina e fenóis em meio sólido, para execução de um processo de compostagem de bagaço de cana (oriundo da moagem das safras 2005/06 e 2006/07) e torta de filtro (safra 2006/07), em escala de casa de vegetação, durante 48 dias. Estes foram distribuídos em 2 consórcios (CM1 e CM2) compostos por 3 microrganismos cada. Fatores como temperatura, umidade, teor de matéria orgânica, nitrato, nitrito, carbono orgânico e fosfato foram avaliados a fim de acompanhar o processo de compostagem desses substratos sem adição de outros macro ou micronutrientes. A partir das análises físico-químicas, verificou-se que não ocorreram grandes variações no desempenho de ambos os consórcios, e que ambos conduziram a uma redução da concentração de matéria orgânica e disponibilização de nitrato e fosfato. Constatou-se que nas condições e período do estudo (48 dias, bandejas largas com uma superfície de exposição do volume de substrato utilizado suscetível a trocas de temperatura com o ambiente, e não adição de outras fontes de nitrogênio e fósforo), o material não atingiu a estabilização. Contudo, os microrganismos aqui apresentados, apresentam amplo potencial para exploração em processos de compostagem de resíduos agroindustriais sólidos/líquidos, desde que se corrijam as relações C:N, e biorremediativos, devido ao seu amplo espectro de atividades enzimáticas. Sugere-se uma combinação dos resíduos sólidos com um percentual de vinhaça, visto esta ser rica nos nutrientes que influenciaram a ação dos microrganismos estudados.

Le sol possède un capital naturel qui lui confère la capacité à produire des services écosystémiques aussi bien culturel que de régulation ou d’approvisionnement, il est indispensable à la Vie telle que nous la connaissons et au développement des activités humaines. Cependant les activités anthropiques et les pollutions, notamment par les éléments traces métalliques (ETMs) tel que le mercure (Hg), perturbent les sols et modifient en profondeur l’organisation des écosystèmes. Face à ces enjeux, des projets de remédiation et de gestion des sites et sols pollués se sont multipliés durant les dernières décennies en vue de futures ré-exploitations de ces sols. Cette thèse s’inscrit dans le cadre des projets ANR-BIOFILTREE et EC2CO FREIDI-Hg gérés par le laboratoire Chrono-Environnement. Mes travaux ont permis l’exploration de la diversité des communautés de microorganismes associées à une plantation de peuplier sur un site contaminé par le Hg et géré par phytomanagement, via les approches combinées de séquençage à très haut débit et par l’approche culture dépendante. Ces méthodes combinées ont permis de révéler i) la diversité des communautés bactériennes et fongiques de la peupleraie ; ii) les groupes de microorganismes particulièrement résistant au Hg (Trichoderma et Pseudomonas) ; et iii) des bactéries promotrices de croissance des plantes (PGPB). Par ailleurs, la compréhension des mécanismes cellulaires liés à l’accumulation de Hg par les microorganismes a été un de mes sujets d’étude en partenariat avec le LIEC (Université de Lorraine). Les modèles eucaryotes Saccharomyces cerevisiae et Podospora anserina ont été utilisés pour tester le rôle potentiel de certains transporteurs d’ions dans l’entrée du Hg dans les cellules fongiques. Les résultats ont montré que le transporteur de magnésium Alr1 situé sur la membrane plasmique pourrait participer au transport du Hg. En outre, une approche de transcriptomique chez Saccharomyces cerevisiae après une courte exposition au Hg des souches mutantes et sauvages a été mise en œuvre. Pour conclure, ce travail de thèse ambitionne d’être un travail de référence pour les futurs projets de phytomanagement en milieux contaminé par le Hg, qui met en avant les communautés de microorganismes et leurs rôles fondamentaux. / Soil has a natural capital that gives it the capacity to produce ecosystem services, cultural as well as regulation or supply, it is essential to the Life as we know it and the development of human activities. However, anthropogenic activities and pollution, in particular by trace elements (ETs) such as mercury (Hg), disrupt the soil and modify in depth the organization of ecosystems. Facing these challenges, remediation and management projects for polluted sites and soils have emerged during the last decades with a view to future re-exploitation of these soils. This thesis is part of the ANR-BIOFILTREE and EC2CO FREIDI-Hg projects managed by the Chrono-Environnement laboratory. My Ph-D work explored the diversity of microorganism communities associated with a poplar plantation at a Hg-contaminated site managed by phytomanagement, combining approaches such as very high-throughput sequencing and conventional culture-based techniques. These combined methods revealed i) the diversity of the bacterial and fungal communities of the poplar plantation; ii) the groups of microorganisms particularly resistant to Hg (Trichoderma and Pseudomonas); and iii) plant growth promoting bacteria (PGPB). In addition, understanding the cellular mechanisms related to the accumulation of Hg by microorganisms was one of my objectives carried out in collaboration with the LIEC (University of Lorraine). The eukaryotic models Saccharomyces cerevisiae and Podospora anserina were used to test the potential role of some ion transporters in the entry of Hg into fungal cells. The results showed that the magnesium transporter Alr1 located on the plasma membrane could participate in the transport of Hg. In addition, a transcriptomic approach in Saccharomyces cerevisiae after a short exposure to Hg of mutant and wild strains has been implemented. To conclude, this work aims to be a reference work for future phytomanagement projects in Hg-contaminated environments, which highlights micro-organism communities and their fundamental roles.

Vulcões ativos na Antártica contrastam com a paisagem predominantemente gelada do continente. Eles fornecem condições únicas capazes de selecionar uma grande variedade de adaptações microbianas. A Ilha Deception localiza-se na região da Península Antártica e difere de outros vulcões antárticos especialmente pela influência marinha e temperaturas mais elevadas. Foram coletadas amostras de sedimentos associados a fumarolas e geleiras em dois sítios geotermais de Deception, com temperaturas variando entre 0°C a 98°C. Diferentes técnicas independentes de cultivo foram empregadas com o intuito de entender como as comunidades microbianas respondem as variações ambientais extremas produzidas pela atividade vulcânica. Os resultados indicaram que a co-ocorrência de arqueias hipertermófilas e suas adaptações com micro-organismos metabolicamente diversos adaptados a regiões geladas representa uma estrutura de comunidades única para ecossistemas antárticos. Este trabalho forneceu dados ineditos sobre questoes centrais de diversidade e adaptacao microbiana a ambientes geotermais polares. / Active volcanoes in Antarctica contrast with the predominately icy landscape. They harbor unique conditions capable to select an extreme range of microbial adaptations. Deception Island is located in the Antarctic Peninsula region and differs from other Antarctic volcanoes specially by its higher temperatures and marine influence. We collected sediment samples associated to active fumaroles and glaciers on two geothermal sites of Deception Island, with temperatures ranging from 0°C to 98°C. Different cultivation-indepedent techniques were used to understand how microbial communities respond to extreme environmental variations produced by volcanic activity. The results indicate that co-occurrence of hyperthermophiles and their specific adaptations with metabolically diverse cold-adapted micro-organisms represents a unique community structure for antarctic ecosystems. This study provided primordial data on central questions about microbial diversity and adaptation to polar geothermal environments.

Este trabalho investigou a diversidade microbiana e a filogenia de arquéias e bactérias em consórcios anaeróbios metanogênicos, originados de sedimentos estuarinos enriquecidos com pentaclorofenol (PCP) e 2,6-diclorofenol (2,6-DCP). Para tanto foram construídas bibliotecas genômicas e utilizados métodos moleculares independentes de cultivo como a Eletroforese em Gel de Gradiente Desnaturante (DGGE) e o seqüenciamento de segmentos específicos do DNAr 16S microbiano. Os resultados da DGGE permitiram verificar alterações na estrutura das comunidades microbianas, as quais provavelmente ocorreram devido às adversidades ocorridas nos sistemas durante o período de incubação como a entrada de oxigênio nos frascos e o acúmulo de compostos clorados no meio de cultivo, principalmente o 2,6-DCP. A estimativa da diversidade beta, realizada pela comparação dos padrões de bandas da DGGE, também permitiu inferir que as alterações nas composições das comunidades de arquéias e bactérias foram devidas às duas estratégias empregadas para o enriquecimento da microbiota autóctone do estuário estudado, ou seja, a pasteurização/não pasteurização das amostras de sedimentos estuarinos. Os resultados das análises filogenéticas revelaram que as seqüências analisadas dos clones bacterianos foram relacionadas ao grupo das bactérias Gram-positivas com baixo conteúdo de G+C pertencentes à Ordem Clostridiales (100%) do Filo Firmicutes e as das arquéias relacionadas ao grupo das metanogênicas pertencentes às Ordens Methanobacteriales (7,1%), Methanosarcinales (14,3%), Methanomicrobiales (57,1%) e arquéias não identificadas (21,4%) do Filo Euryarchaeota. Em relação às bactérias, alguns clones foram identificados como pertencentes aos gêneros Sedimentibacter, Clostridium e Alkalibacter, os quais são representados por microrganismos que apresentam metabolismo fermentativo e requerem a presença de extrato de levedura para o crescimento. Provavelmente as bactérias analisadas neste trabalho fermentaram a glicose e o piruvato, os quais foram utilizados como doadores de elétrons, com conseqüente produção de lactato, etanol, butirato, acetato, formiato e hidrogênio/gás carbônico, que podem ter sido utilizados por outros grupos de microrganismos no processo global da digestão anaeróbia. Tais bactérias foram importantes para o processo global de degradação dos clorofenóis, pois também utilizaram como doadores de elétrons os produtos parcialmente degradados por outras bactérias fazendo com que não houvesse acúmulo desses compostos no sistema. Enquanto que algumas arquéias foram relacionadas a organismos hidrogenotróficos pertencentes aos gêneros Methanoculleus e Methanocalculus, Methanobacterium, e acetotróficos do gênero Methanosaeta, as quais utilizaram como substratos para a metanogênese os subprodutos da fermentação bacteriana, ou seja, o \'H IND.2\'/\'CO IND.2\', o formiato e o acetato, contribuindo assim para a manutenção do equilíbrio das demais reações ocorridas no sistema. Desta forma, os dados obtidos neste trabalho poderão auxiliar na compreensão do funcionamento de ecossistemas contaminados por compostos clorados, bem como contribuir para o desenvolvimento de novos processos biotecnológicos aplicados aos problemas ambientais. / This work aimed to investigate microbial diversity and phylogeny of archaea and bacteria microrganisms methanogenic in estuarine sediment samples enriched with organic sources under methanogenic and halophlic conditions. The samples were obtained from previous study on anaerobic degradation of pentachlorophenol (PCP) and 2,6-dichlorophenol (2,6-DCP). Microbial studies were done by the molecular methods without cultivation requirement, Denaturing Gradient Del Electrophoresis (DGGE) and specific segments of 16S rDNA sequencing. DGGE-profile showed structure changes in microbial community. Probably, as a consequence of \'O IND.2\' intake and accumulation of chlorinated compounds, mainly 2,6-DCP, in the culture medium. The beta diversity estimation showed that changes in arquaea and bacteria communities probably occurred as a consequence of the two enrichment strategies used for estuarine indigenous microorganisms, pasteurization and non-pasteurization of the estuarine sediments samples. Phylogenetic analysis indicated the presence of gram-positive bacterium with low G+C content related to Clostridiales Order (100%) belonging to Firmicutes Phylum, as well as related to methanogenic archaea from Methanobacteriales (7,1%), Methanosarcinales (14,3%) and Methanomicrobiales (57,1%) Order. Non-identified archaeas from Euryarchaeota Phylum were also found (21,4%). Concerning to bacterias, it was identified clones related to genera Sedimentibacter, Clostridium and Alkalibacter, which have fermentative metabolism and require yeast extract to grow. Probably, bacterial cells analised in this work fermented glucose and pyruvate producing lactate, ethanol, butirate, acetate, formiate and hydrogen/carbon dioxide. All these products could be used for other microbial groups in the global process of anaerobic digestion. The bacterial microorganisms were important to global process of chlorophenol degradation because contributed in the overall process utilizing the products partially degraded by other bacterias and preventing its accumulation in the system. Identified archaeal cells were related to hydrogenotrophs microorganisms of the genera Methanosaeta, Methanoculleus, Methanocalculus and Methanobacterium, which utilized the bacterial fermentation sub-products as acetate, \'H IND.2\'/\'CO IND.2\' and formiate as substrate for the methanogenesis, contributing for the maintenance of the balance of other reactions occurred in the system. The results of this work give advanced knowledge about understanding biotechnological process of contaminated ecosystems by chlorinated compounds. Therefore, it could contribute to development of new biotechnological processes applied to environmental problems.

As bactérias fototróficas freqüentemente apresentam florescimentos em lagoas de estabilização utilizadas no tratamento de esgoto sanitário, formando uma camada de cor púrpura na sua superfície. Portanto, o estudo das condições que propiciam tais florescimentos, a diversidade microbiana, o potencial de remoção da matéria orgânica e o estabelecimento das relações entre tais conhecimentos, permitem compreender o metabolismo do sistema. Nesse sentido, o objetivo deste trabalho foi avaliar a diversidade de bactérias (domínio Bacteria), bactérias fototróficas púrpuras e bactérias redutoras de sulfato (BRS) em lagoas de estabilização do Vale do Ribeira (Cajati, SP). Para tal, foram realizadas coletas sazonais (primavera, verão, outono e inverno) na sub-superfície, camada intermediária e interface água-sedimento, em dois horários (14:00 h e 02:00 h), nas lagoas anaeróbia e facultativa. Para analisar os diferentes grupos de microrganismos, utilizou-se a técnica de PCR/DGGE, com primers específicos. Nas análises de filogenia realizou-se o seqüenciamento parcial do gene RNAr 16S e da subunidade M do centro de reação fotossintético das bactérias fototróficas púrpuras. Análises físico-químicas, tais como sulfato, DQO, sólidos, nitrogênio e fósforo foram realizadas, além da determinação da concentração de oxigênio dissolvido, pH, temperatura e radiação solar fotossinteticamente ativa incidente. No outono observou-se maior diversidade de microrganismos do domínio Bacteria, bactérias fototróficas púrpuras e BRS, enquanto na primavera foi verificada a menor diversidade desses microrganismos para as duas lagoas. Na lagoa facultativa foi observada maior diversidade do domínio Bacteria e das BRS em relação à lagoa anaeróbia. Verificou-se maior diversidade de bactérias fototróficas púrpuras na lagoa anaeróbia, caracterizada por duas populações predominantes nas quatro estações e nas diferentes profundidades. A concentração de matéria orgânica (DQO) variou de 60,3 mg/L (inverno) a 298,0 mg/L (primavera) e a maior concentração de sulfato observada foi de 51,0 mg/L (inverno). Bacilo curvo Gram negativo, semelhante à bactéria fototrófica púrpura não sulfurosa, presente em amostra proveniente da sub-superfície da lagoa anaeróbia foi purificado e apresentou 92% de similaridade com Rhodopseudomonas palustris. Em ambas as lagoas foram identificadas bactérias semelhantes a Chromobacterium suttsuga (95%), Clostridium sp. (99%), Rhodobacter sphaeroides (99%), Rhodopseudomonas palustris (99%), Lampropedia hyalina (97%), Campylobacter fetus (99%), Desulfovibrio vulgaris (95%), Rhodospirillum rubrum (95%) e diferentes bactérias não cultivadas. / The phototrophic bacteria frequently blossom in the stabilization lagoons that are used in sanitary sewer treatment, forming a purple layer on its surface. Therefore, the study of the conditions that propitiate such blooms, the microbial diversity, the removal of the organic matter and the establishment of the relations between them permit to understand the metabolism of the system. The objective of this work was to evaluate the diversity of the bacteria (Bacteria domain), purple phototrophic bacteria and sulfate reducing bacteria (SRB) in stabilization lagoons of Vale do Ribeira (Cajati - SP). For this, it was made seasonal collects (spring, summer, autumn and winter) from the sub-surface, intermediate layer and interface water-sediment, at two times (14:00 h and 02:00 h) of the anaerobic and facultative lagoons. To analyze the different groups of microorganisms it was used the PCR/DGGE technique, with specific primers; for the phylogenic analysis it was realized the DNA partial sequencing of the 16S RNAr gene and of the subunit M of the photosynthetic center of reaction of the purple photosynthetic bacteria. It was determined: the concentration of dissolved oxygen, pH, temperature and photosynthetically active incident solar radiation, and the physical-chemistry analysis as: COD, solids, nitrogen and phosphorus. In the autumn it was observed greater diversity of microorganisms of the Bacteria domain, the group of the purples phototrophic bacteria and SRB, while in the spring it was verified minor diversity of these microorganisms in the two lagoons studied. In the facultative lagoon it was observed greater diversity of the Bacteria domain and of the SRB with respect to the anaerobic lagoon. It was verified greater diversity of the purple phototrophic bacteria in the anaerobic lagoon, of what in the facultative lagoon, which was characterized by the two predominant populations in the four seasons and in the different points of collect. The concentration of the organic matter (COD) varied from 60,3 mg/L (winter) to 298,0 mg/L (spring) and the greater concentration of sulfate observed was of 51,0 mg/L (winter). Arched bacillus Gram-negative similar to purple not sulfurous bacteria, from a sample of the sub-surface of the anaerobic lagoon was purified and presented 92% of similarity with Rhodopseudomonas palustris. In both lagoons it was identified bacteria similar to Chromobacterium suttsuga (95%), Clostridium sp. (99%), Rhodobacter sphaeroides (99%), Rhodopseudomonas palustris (99%), Lampropedia hyalina (97%), Campylobacter fetus (99%), Desulfovibrio vulgaris (95%), Rhodospirillum rubrum (95%).

Les interactions isopodes terrestres-Wolbachia féminisantes constituent un excellent modèle d'étude des interactions multipartites complexes. A ce jour, trois Wolbachia féminisantes, issues d'histoires évolutives différentes, ont été identifiées chez l'hôte Armadillidium vulgare. L'objectif de cette thèse est d'avoir une vue plus complète du microbiote d'A. vulgare et de l'impact des souches de Wolbachia sur cette communauté bactérienne. Dans ce but, des approches quantitative et métagénomique ont été combinées afin de caractériser le microbiote d'A.vulgare à plusieurs niveaux d'intégration: (i) dans des populations naturelles et des lignées contrôlées de laboratoire, (ii) au sein de différents tissus hôtes et (iii) en relation avec le statut d'infection par Wolbachia. Aucun microbiote coeur n'a été mis en évidence malgré une relative homogénéité des communautés symbiotiques dans les tissus de l'hôte. De plus, les bactéries environnementales avaient un impact important sur le microbiote symbiotique. Chez les animaux infectés, Wolbachia représente la bactérie dominante et le facteur majeur de la structuration de la communauté symbiotique. Nous avons également mis en évidence la présence d'un autre constituant majeur du microbiote dans tous les tissus de l'hôte: Candidatus Hepatoplasma crinochetorum, un symbiote facultatif réputé jusque-là pour être seulement associé aux glandes digestives de l'intestin. Afin d'éclairer les interactions potentielles entre hôte et ce symbiote relativement peu connu, la séquence du génome de Ca. H. crinochetorum a été obtenue. Une analyse préliminaire des interactions fonctionnelles des voies métaboliques impliquées dans la digestion de la cellulose nous a permis d'identifier des enzymes cellulolytiques spécifiques de l'hôte et du symbiote. Cette étude ouvre de nouvelles perspectives dans l'étude de la symbiose chez les isopodes. / In recent years, there has been a shift of focus in symbiosis studies, away from the traditional 'one host-one symbiont' concept towards a more holistic, community-based approach. This concept takes into account that a host is not only associated with one bacterium, but harbours and interacts with a diverse bacterial community, the microbiome. Terrestrial isopods represent an excellent model system for the understanding of complex multipartite symbioses due to their well-characterised association with feminizing Wolbachia bacteria. To date, three different feminizing Wolbachia strains have been identified in Armadillidium vulgare, presumably representing different host-symbiont co-evolutionary histories. The aim of this PhD was to get a more complete picture of the terrestrial isopod microbiome and the role of Wolbachia within the bacterial community. In order to achieve this, quantitative and metagenomic techniques were combined to characterize the microbiome of A. vulgare on multiple levels: (i) Field vs. lab populations, (ii) in different host tissues, and (iii) in relation to Wolbachia infection status, i.e. presence/absence of Wolbachia as well as infection with different Wolbachia strains. Wolbachia represented the predominant member of the bacterial community in infected individuals and was identified as an important factor influencing bacterial community structure. Apart from Wolbachia, we detected a second highly abundant bacterium: Candidatus Hepatoplasma crinochetorum, a facultative symbiont previously reported from the midgut caeca, was for the first time observed in all tested host tissues. The potential interactions of Wolbachia and Ca. H. crinochetorum constitute an interesting example for symbiont-symbiont relationships between two highly abundant members of a diverse bacterial community.