Evolution of strategies for biodegradation of synthetic organic pollutants relies on recruitment of genes from catabolic pathways for natural compounds. Investigation of metabolic diversity in nature can provide insight into biochemical strategies that could be recruited for bioremediation of pollutants. As part of a search for novel metabolic diversity we isolated soil bacteria able to degrade 2-nitroimidazole (2NI) and 5-nitroanthranilic acid (5NAA), and determined the biochemistry and molecular biology of their biodegradation pathways. 2NI and its analogs are increasingly used as prodrugs for the treatment of both tuberculosis and cancer. The biodegradation of 2NI by a soil Mycobacterium sp. is initiated by an unusual hydrolytic denitration. The reaction is catalyzed by a novel nitrohydrolase with a divergent sequence and represents the discovery of a previously unreported drug resistance mechanism in soil prior to its identification in clinical situations. 5NAA is the starting material for various nitroaromatic compounds and dyes. The biodegradation pathway of 5NAA is initiated by an unusual hydrolytic deamination. The corresponding gene is very distantly related to biochemically characterized genes in the NCBI database. The nitro group of 5NAA is eliminated as nitrite during the spontaneous formation of lactones from a ring fission product, a previously unreported mechanism. Degradation pathways of 5NAA and 2NI serve as precedents for those of nitroaniline and nitroimidazole pollutants. The work supports the hypothesis that the study of the metabolism of natural organic compounds selected on the basis of unusual structural features and ecological roles can reveal new metabolic diversity.

A cana-de-açúcar (S. officinarum) é uma gramínea perene de grande importância na economia brasileira. Devido a isto, estudos relativos ao aprimoramento das condições de cultivo são de grande interesse, podendo ser uma destas bases um melhor conhecimento sobre as comunidades microbianas associadas à cana-de-açúcar. Sabe-se que a rizosfera é um ambiente de íntima interação entre as plantas e seus respectivos microbiomas, sendo esta relação intermediada pela exsudação radicular. Este estudo teve como objetivo realizar o cultivo de bactérias da rizosfera e do solo de cana-de-açúcar, sendo os isolados obtidos posteriormente caracterizados genética (BOX-PCR e sequenciamento parcial do gene 16S rRNA) e metabolicamente (BIOLOG®). Os resultados demonstraram que o número de bactérias cultiváveis na rizosfera é maior comparado ao solo, sendo que dentre os isolados destaca-se a maior afiliação taxonômica ao filo Proteobacteria (principalmente as classes γ-proteobacteria e β- proteobacteria). A análise de BOX-PCR mostrou uma grande diversidade genética, mesmo quando comparados isolados obtidos a partir do mesmo meio de cultivo, ou pertencentes ao mesmo grupo taxonômico. Em contrapartida, a análise de sequenciamento parcial do gene 16S rRNA mostrou que muitos isolados preservam grande similaridade do gene ribossomal analisado. A análise de perfil metabólico corroborou com os dados de BOX-PCR, onde isolados com afiliação taxonômica bastante correlata apresentaram capacidades distintas de utilizar as diversas fontes de carbono avaliadas. Por fim, esta diversidade metabólica se traduziu na obtenção de respostas distintas de isolados pertencentes aos mesmos gêneros bacterianos, isolados do solo ou da rizosfera, quando estes foram cultivados na presença de exsudatos radiculares. De maneira geral, este estudo demonstrou que as plantas de cana-de-açúcar podem influenciar o comportamento das comunidades bacterianas presentes no solo, e indicaram que existe uma grande diversificação dos organismos presentes na rizosfera, sendo a resposta a este ambiente diferenciada de forma independente da afiliação taxonômica dos isolados. / The sugarcane plant (S. officinarum) is a perennial gamineous with a great importance for the Brazilian economy. Due to this, studies related to the improvement of cultivation conditions are of great importance, indicating that one of these bases must contribute with the better knowledge about the microbial communities associated with sugarcane. It is known that the rhizosphere is an environment that hosts an intimate interaction between plants and their respective microbiomes, being it mediated by the roots exudates. This study aimed to cultivate bacterial isolates from soil and rhizosphere of sugarcane, followed by the genetic (BOX-PCR and partial sequencing of the 16S rRNA gene) and metabolic (BIOLOG®) characterization of them. Results demonstrated higher numbers of cultivable bacteria in rhizosphere when compared to soil samples, with the prevalent affiliation of the isolates to the phylum Proteobacteria (specially with the classes γ-proteobacteria and β- proteobacteria). The BOX-PCR results showed a great genetic diversity, even when isolates obtained in the same culture medium or affiliated to the same taxa are compared. In counterpart, the analysis of the 16S rRNA gene sequence indicated that several isolates preserve high similarities in the ribosomal gene. The metabolic profiling results corroborated with the BOX-PCR data, which isolates highly correlated in the taxonomical analyses presented distinct capacities to use the carbon sources that were tested. At the end, this metabolic diversity was evidenced by the distinct behavior of isolates belonging to the same genera, isolated from soils or rhizosphere samples as well, when cultivated in the presence of roots exudates. In general, this study demonstrated that sugarcane plants can influence the behavior of bacterial communities present in soils, and indicated a great diversification of organisms present in the rhizosphere being the response to this environment very particular, regardless of the taxonomical affiliation of the isolates.

A cana-de-açúcar (S. officinarum) é uma gramínea perene de grande importância na economia brasileira. Devido a isto, estudos relativos ao aprimoramento das condições de cultivo são de grande interesse, podendo ser uma destas bases um melhor conhecimento sobre as comunidades microbianas associadas à cana-de-açúcar. Sabe-se que a rizosfera é um ambiente de íntima interação entre as plantas e seus respectivos microbiomas, sendo esta relação intermediada pela exsudação radicular. Este estudo teve como objetivo realizar o cultivo de bactérias da rizosfera e do solo de cana-de-açúcar, sendo os isolados obtidos posteriormente caracterizados genética (BOX-PCR e sequenciamento parcial do gene 16S rRNA) e metabolicamente (BIOLOG®). Os resultados demonstraram que o número de bactérias cultiváveis na rizosfera é maior comparado ao solo, sendo que dentre os isolados destaca-se a maior afiliação taxonômica ao filo Proteobacteria (principalmente as classes γ-proteobacteria e β- proteobacteria). A análise de BOX-PCR mostrou uma grande diversidade genética, mesmo quando comparados isolados obtidos a partir do mesmo meio de cultivo, ou pertencentes ao mesmo grupo taxonômico. Em contrapartida, a análise de sequenciamento parcial do gene 16S rRNA mostrou que muitos isolados preservam grande similaridade do gene ribossomal analisado. A análise de perfil metabólico corroborou com os dados de BOX-PCR, onde isolados com afiliação taxonômica bastante correlata apresentaram capacidades distintas de utilizar as diversas fontes de carbono avaliadas. Por fim, esta diversidade metabólica se traduziu na obtenção de respostas distintas de isolados pertencentes aos mesmos gêneros bacterianos, isolados do solo ou da rizosfera, quando estes foram cultivados na presença de exsudatos radiculares. De maneira geral, este estudo demonstrou que as plantas de cana-de-açúcar podem influenciar o comportamento das comunidades bacterianas presentes no solo, e indicaram que existe uma grande diversificação dos organismos presentes na rizosfera, sendo a resposta a este ambiente diferenciada de forma independente da afiliação taxonômica dos isolados. / The sugarcane plant (S. officinarum) is a perennial gamineous with a great importance for the Brazilian economy. Due to this, studies related to the improvement of cultivation conditions are of great importance, indicating that one of these bases must contribute with the better knowledge about the microbial communities associated with sugarcane. It is known that the rhizosphere is an environment that hosts an intimate interaction between plants and their respective microbiomes, being it mediated by the roots exudates. This study aimed to cultivate bacterial isolates from soil and rhizosphere of sugarcane, followed by the genetic (BOX-PCR and partial sequencing of the 16S rRNA gene) and metabolic (BIOLOG®) characterization of them. Results demonstrated higher numbers of cultivable bacteria in rhizosphere when compared to soil samples, with the prevalent affiliation of the isolates to the phylum Proteobacteria (specially with the classes γ-proteobacteria and β- proteobacteria). The BOX-PCR results showed a great genetic diversity, even when isolates obtained in the same culture medium or affiliated to the same taxa are compared. In counterpart, the analysis of the 16S rRNA gene sequence indicated that several isolates preserve high similarities in the ribosomal gene. The metabolic profiling results corroborated with the BOX-PCR data, which isolates highly correlated in the taxonomical analyses presented distinct capacities to use the carbon sources that were tested. At the end, this metabolic diversity was evidenced by the distinct behavior of isolates belonging to the same genera, isolated from soils or rhizosphere samples as well, when cultivated in the presence of roots exudates. In general, this study demonstrated that sugarcane plants can influence the behavior of bacterial communities present in soils, and indicated a great diversification of organisms present in the rhizosphere being the response to this environment very particular, regardless of the taxonomical affiliation of the isolates.

Les Streptomyces possédent un large arsenal enzymatique ayant des rôles importants dans le sol. Au cours de cette thèse, nous avons exploré leur diversité génétique, fonctionnelle et écologique à partir de collections provenant de sols forestiers. Ainsi, l’exploration du potentiel cellulolytique et la capacité à détecter des sucres libérés lors de l’attaque du bois par des champignons lignivores a permis de créer un biosenseur dont l’exploitation pourrait constituer un nouvel outil normatif pour la détection de la dégradation du bois. Suite à une approche de génomique comparative réalisée entre des isolats sympatriques, nos résultats ont permis de démontrer que des souches phylogénétiquement très apparentées présentaient de grandes différences en termes de présence/absence de gènes, suggérant une vitesse d’évolution rapide du génome accessoire au sein de la population. Ces gènes, souvent associés à des éléments potentiellement transférables, a souligné un rôle important du transfert horizontal pour la diversification de la population. Par une approche d’écologie réverse, la fonction prédite de certains de ces gènes a également pu être corrélée avec un rôle écologique potentiel. Ainsi, l’un des clusters de gènes variables identifié était impliqué dans la production de métabolites secondaires et pourrait constituer un bien commun pour la population. Nos résultats ont confirmé la grande diversité métabolique des Streptomyces (et leur utilité à des fins appliquées), mais indique également qu’une diversification rapide entre souches proches, aurait un rôle écologique important au niveau des populations naturelles de Streptomyces / Streptomyces are known to possess a large enzymatic arsenal which can have important roles in the soil. During this thesis, we explored their genetic, functional and ecological diversity using collections from forest soils. Thus, the exploration of their cellulolytic potential and their ability to detect complex sugars released by wood during lignivorous fungi attacks has led to the creation of a biosensor whose exploitation could constitute a new normative tool for the detection of the degradation of wood. Subsequent to comparative genomic approach carried out between sympatric isolates, our results also demonstrated that phylogenetically highly related strains exhibited large differences in the presence / absence of genes, suggesting a rapid rate of evolution of the population accessory genome. These genes, often associated with potentially transferable elements, underlined important role of horizontal transfer for population diversification. Using a reverse ecology approach, the predicted function of some of these genes could also be correlated with a potential ecological role. Thus, one of the variable gene clusters identified by genome analysis was involved in the production of secondary metabolites and would constitute a common good for the population. All of our results confirm the wide metabolic diversity of Streptomyces (and their utility for applied purposes), but also indicates that this diversification would be rapid between nearby strains and would have an important ecological role in the natural populations of Streptomyces

No description available.