Magister Scientiae (Biodiversity and Conservation Biology) - MSc (Biodiv & Cons Biol) / Fire as a form of disturbance is unique in the way it impacts upon the environment, acting like a herbivore with a ubiquitous appetite. Consuming both dead and living material, converting complex organic molecules into organic and mineral products, which return to the soil. The role of disturbance has long been considered a driver of diversity within Mediterranean type ecosystems. Recently the interest in soil microbes has been piqued, as the importance thereof has been emphasised, particularly their role in nutrient cycling and the chelation of essential plant nutrients. The occurrence of fire results in several environmental and ecological impacts on soil, as well as the dynamics of the microbial populations present. This study explores the impact of fire as a disturbance on the plant and bulk soil microbial communities of the Succulent Karoo. This was achieved through two sub-studies, in which three different states were studied: unburnt, 7-year and 2-year post-fire. Today microbial profiles are also used as indicators of disturbance, thus many techniques exploring microbial community composition are available.

Les virus sont ubiquitaires et abondants dans l’environnement. Ils influent fondamentalement sur l’écologie de l’ensemble des écosystèmes et du microbiote humain. Dès 2002, avec la découverte de virus géants d’amibes, la virologie s’est complexifiée. Les Megavirales (nouvel ordre au sein des grands virus nucléocytoplasmiques) ont 10% de gènes homologues aux cellules eucaryotes, et ont la caractéristique singulière d’être infectés par des virophages.Avec l’avènement de la métagénomique, le nombre de métagénomes produits ne cesse de croître de manière exponentielle. C’est ainsi que la virologie a connu un nouvel essor et a pu mieux être étudiée en s’affranchissant des difficultés de culture et d’isolement des virus dans les conditions artificielles de laboratoire. La métagénomique permet d’étudier les communautés microbiennes mais également de découvrir de nouveaux microbes. La bioinformatique est devenue incontournable dans le domaine de la biologie et essentielle pour les biologistes afin de traiter les masses de données et en extraire toute la richesse de l’information biologique nécessaire. La première partie de cette thèse consiste en une revue de la littérature décrivant la bioinformatique au service de la métagénomique virale. La deuxième partie présente la création d’un nouvel outil « MG-Digger » dédié à l’analyse rapide et automatisée de séquences d’intérêts spécifiques dans les métagénomes. La dernière partie se concentre sur l’utilisation de cet outil sur des données issues de projets métagénomiques afin de répondre à des questions biologiques précises, notamment sur les données de l’expédition scientifique TARA à travers les océans. / Viruses are ubiquitous and abundant in the environment and can influence all ecosystems ecology and the human microbiota.Since 2002, with the discovery of giant viruses of amoeba, virology has become more complex and the definition of virus has been called into question, not only because of their phenotypic sizes similar to those of bacteria but also their genomic content exceeding thousand genes. Megavirales, also known as nucleocytoplasmic large DNA viruses, have 10% homologous genes to eukaryotic cells and interestingly can be infected by virophages. With the advent of metagenomic, the number of metagenomes produced is exponentially increasing as well as our understanding of virology which has been studied. Metagenomics studies showed an efficient way to study microbial communities and identify novel viruses without the difficulties of culture and isolation of viruses in artificial laboratory conditions.Metagenomic requires considerable computational and storage resources (Big data processing). Therefore, bioinformatics has become an integral part of research and development in the biomedical sciences by providing tools that handle complex datasets and finally giving the necessary biological information. The first part of this thesis consists of an exhaustive review of the literature describing bioinformatics and viral metagenomics. The second part presents a new "MG-Digger" tool dedicated to the rapid and automated analysis of specific interest sequences in metagenomes. The third part focuses on the use of this tool on metagenomic data to answer to specific biological questions, including data from the TARA scientific expedition across the oceans.

Made available in DSpace on 2014-06-11T19:27:21Z (GMT). No. of bitstreams: 0 Previous issue date: 2004-04-07Bitstream added on 2014-06-13T19:14:40Z : No. of bitstreams: 1 silveira_el_me_jabo.pdf: 655008 bytes, checksum: ac46de019a2703894b3c1bc37633b43c (MD5) / Coordenação de Aperfeiçoamento de Pessoal de Nível Superior (CAPES) / Métodos tradicionais de isolamento e cultivo limitam análises da diversidade microbiana no meio ambiente, pois acredita - se que aproximadamente 10% desses microrganismos possam ser cultivados. A ecologia microbiana molecular teve recentes progressos através da construção de bibliotecas metagenômicas, o que constitui uma poderosa abordagem para explorar a diversidade microbiana de solo fornecendo inclusive dados sobre os microrganismos não cultiváveis desse habitat. Este trabalho teve por objetivo comparar e estimar a diversidade de comunidades bacterianas, em solos de duas áreas, sendo solo de Floresta Nativa (SFN) e a outra sob arboreto com eucaliptos (SAE) de uma mesma região. Utilizando oligonucleotídeos iniciadores específicos, o gene 16S rRNA foi amplificado por PCR, os amplicons foram clonados em pGEMR-T e os clones obtidos seqüenciados parcialmente. No solo SFN foram analisados 231 clones e no solo SRE 248 clones. As seqüências obtidas foram submetidas à análise de similaridade de nucleotídeos com o banco de dados GenBank. Os filos bacterianos que mais se destacaram nos dois tipos de solo foram Acidobacteria e Proteobacteria. No solo SFN destacaram-se também as bactérias pertencentes ao filo Bacteroidetes e no solo SAE observou-se alta freqüência das bactérias Actinobacteria. Análises filogenéticas revelaram diferenças em ambos os solos, verificando através de índice de diversidade bacteriana observou-se que o solo sob eucalipto apresentou maior diversidade quando comparado ao solo sob de Floresta Nativa. / Traditional methods of isolation and culture limits the analyses of the microbian diversity in the environment, because it is believed - that approximately 10% of these microrganisms can be cultivated. The molecular microbian ecology have had recent progress through the construction of metagenomics Iibraries, what constitutes a powerful boarding to explore the microbian diversity of soil, also supplying information on the microrganisms that cannot be cultivated on this habitat. This work had the objective of stimate the diversity of bacterial communities, in two areas, an area of Native Forest (SFN) and the another one under eucalypts (SAE) of the same region. Using specific oligonucleotides starters, the gene 16S rRNA was amplified by PCR, amplicons had been cloned in pGEMR-T and the obtained clones were partial/y sequenced. In the soil SFN, 231 clones had been analyzed and 248 clones in the soil SAE. The sequences obtained were submitted to the nucleotides similarity analysis with the GenBank database. The bacterial phylum that was more evident in the two types de soil were Acidobacteria and Proteobacteria. In the soi! SFN the bacteria of the phylum Bacteroidetes were evident and in the soil SRE high frequency of the Actinobacteria bacteria was observed. Phylogenetic analyses reveled an extensive diversity in both soils, verified through diversity index differences in the bacterial communities in both areas, and that the area under eucalypt presented more diversity in relation to the area of Native Forest.

Made available in DSpace on 2015-04-01T14:16:05Z (GMT). No. of bitstreams: 1 arquivototal.pdf: 1357815 bytes, checksum: a8f6776ded44373a63e29cc1331535eb (MD5) Previous issue date: 2013-04-19 / Coordenação de Aperfeiçoamento de Pessoal de Nível Superior - CAPES / Reproductive disorders in bovines can be caused by various pathogens, which might already be present in the reproductive tract. The microbial community of the reproductive apparatus, when known, can provide information about the health of the host. The metagenomics has been used to characterize and obtain genetics information about microbial communities in various environments and can relate certain diseases with changes in community composition. In this study, samples were collected from the secretion of mucosa of vaginal surface of 05 healthy cows and 05 cows that showed symptoms of reproductive disorders. Metagenomics DNA samples were extracted of secretion samples and amplified with primers for the V5-V6 regions of the 16S rRNA and sequenced on the platform "Ion Torrent Personal Genome Machine - PGM". The data were processed to remove low quality sequences and chimeras, with the Mothur program, then the data were released in the Ribosomal Database Project for classification of OTU's. Local blastn was performed and this results was loaded in MEGAN program, for viewing profiles and taxonomic microbial attributes. The control profile showed a total of 15 taxa, being the taxa Bacteroides, Enterobacteriaceae and Victivallis, with the highest representation of OTU's; the positive profile showed 68 taxa, and Bacteroides, Enterobacteriaceae, Histophilus, Victivallis, Alistipes and Coriobacteriaceae, the taxa with more OTU's representation. The genus Histophilus have pathogenics species in reproductive tract of cattle. There was a change in the community composition, well as in microbial attributes of profiles there may be a relationship between the pathogen and of other representatives taxa, in production of metabolites to disease progression. / Distúrbios reprodutivos em bovinos podem ser causados por diversos patógenos, que poderiam já estar presentes no trato reprodutivos. A comunidade microbiana dos aparelhos reprodutivos, quando conhecida, pode fornecer informações a respeito da saúde do hospedeiro. A metagenômica tem sido utilizada para caracterizar e obter informações genéticas a respeito de comunidades microbianas de diversos ambientes, podendo relacionar determinadas doenças com alterações na composição das comunidades. Nesse trabalho, foram coletadas amostras de secreção de mucosa da superfície vaginal de 05 vacas sadias e 05 vacas que apresentaram sintomas de distúrbios reprodutivos. Amostras de DNA metagenômico foram extraídas das amostras de secreção e amplificadas com primers para as regiões V5-V6 do gene 16S rRNA e sequenciadas na plataforma Ion Torrent Personal Genome Machine PGM . Os dados foram tratados para retirada de sequencias de baixa qualidade e quimeras, com o programa Mothur; em seguida, os dados foram lançados no Ribosomal Database Project para classificação das OTU´s. Blastn local foi efetuado e seus resultados foram carregados no programa MEGAN, para visualização de perfis taxonômicos e atributos microbianos. O perfil controle apresentou um total de 15 taxa, sendo os taxa Bacteroides, Enterobacteriaceae e Victivallis, os de maior representação de OTU´s; o perfil com distúrbios apresentou 68 taxa, sendo Bacteroides, Enterobacteriaceae, Histophilus, Victivallis, Alistipes, e Coriobacteriaceae, os taxa com maior representação de OTU´s. O gênero Histophilus possui espécies patogênicas em trato reprodutivo de bovinos. Observou-se uma alteração na composição das comunidades estudadas, bem como nos atributos microbianos dos perfis podendo haver uma relação entre patógenos e representantes de outros taxa, na produção de metabólitos para progressão da doença.

La métagénomique ciblée, étude de la composition et de la diversité des communautés microbiennes présentes dans différents échantillon biologiques sur la base d'un marqueur génomique, a connu un véritable essor lors de cette dernière décennie grâce à l'arrivée du séquençage haut-débit. Faisant appel à des outils de biologie moléculaire et de bioinformatique, elle a été à l’origine de substantiels progrès dans les domaines de l’évolution et de la diversité microbienne. Cependant, de nouvelles problématiques sont apparues avec le séquençage haut-débit : la génération exponentielle de données soulève des problèmes d'analyse bioinformatique, qui doit être adaptée aux plans d'expérience et aux questions biologiques associées. Cette thèse propose des solutions d'amélioration des études de métagénomique ciblée par le développement d'outils et de méthodes innovantes, apportant une meilleure compréhension des biais d'analyse inhérents à de telles études, et une meilleure conception des plans d'expérience. Tout d'abord, une expertise du pipeline d'analyse utilisé en production sur la plate-forme PEGASE-biosciences a été menée. Cette évaluation a révélé la nécessité de mettre en place une méthode d'évaluation formelle de pipelines d'analyses de données de métagénomique ciblée, qui a été développée sur la base de données simulées et réelles, et de métriques d'évaluation adaptées. Cette méthode a été utilisée sur plusieurs pipelines d'analyse couramment utilisés par la communauté, tout comme sur de nouvelles approches d'analyse jamais utilisées dans un tel contexte. Cette évaluation a permis de mieux comprendre les biais du plan d'expérience qui peuvent affecter les résultats et les conclusions biologiques associées. Un de ces biais majeurs est le choix des amorces d'amplification de la cible ; un logiciel de design d'amorces adaptées au plan d'expérience a été spécifiquement développé pour minimiser ce biais. Enfin, des recommandations de montage de plan d'expérience et d'analyse ont été émises afin d'améliorer la robustesse des études de métagénomique ciblée. / Targeted metagenomics is the study of the composition of microbial communities in diverse biological samples, based on the sequencing of a genomic locus. This application has boomed over the last decade thanks to the democratisation of high-throughput sequencing, and has allowed substantial progress in the study of microbial evolution and diversity. However, new problems have emerged with high-throughput sequencing : the exponential generation of data must be properly analyzed with bioinformatics tools fitted to the experimental designs and associated biological questions. This dissertation provides solutions to improve targeted metagenomics studies, by the development of new tools and methods allowing a better understanding of analytical biases, and a better design of experiments. Firstly, an expert assessment of the analytical pipeline used on the PEGASE-biosciences plateform has been performed. This assessment revealed the need of a formal evaluation method of analytical pipelines used for targeted metagenomics analyses. This method has been developed with simulated and real datasets, and adequate evaluation metrics. It has been used on several analytical pipelines commonly used by the scientific community, as well as on new analytical methods which have never been used in such a context before. This evaluation allowed to better understand experimental design biases, which can affect the results and biological conclusions. One of those major biases is the design of amplification primers to target the genomic locus of interest. A primer design software, adaptable to different experimental designs, has been specifically developed to minimize this bias. Finally, analytical guidelines and experimental design recommendations have been formulated to improve targeted metagenomics studies.

Resumo: Métodos tradicionais de isolamento e cultivo limitam análises da diversidade microbiana no meio ambiente, pois acredita - se que aproximadamente 10% desses microrganismos possam ser cultivados. A ecologia microbiana molecular teve recentes progressos através da construção de bibliotecas metagenômicas, o que constitui uma poderosa abordagem para explorar a diversidade microbiana de solo fornecendo inclusive dados sobre os microrganismos não cultiváveis desse habitat. Este trabalho teve por objetivo comparar e estimar a diversidade de comunidades bacterianas, em solos de duas áreas, sendo solo de Floresta Nativa (SFN) e a outra sob arboreto com eucaliptos (SAE) de uma mesma região. Utilizando oligonucleotídeos iniciadores específicos, o gene 16S rRNA foi amplificado por PCR, os amplicons foram clonados em pGEMR-T e os clones obtidos seqüenciados parcialmente. No solo SFN foram analisados 231 clones e no solo SRE 248 clones. As seqüências obtidas foram submetidas à análise de similaridade de nucleotídeos com o banco de dados GenBank. Os filos bacterianos que mais se destacaram nos dois tipos de solo foram Acidobacteria e Proteobacteria. No solo SFN destacaram-se também as bactérias pertencentes ao filo Bacteroidetes e no solo SAE observou-se alta freqüência das bactérias Actinobacteria. Análises filogenéticas revelaram diferenças em ambos os solos, verificando através de índice de diversidade bacteriana observou-se que o solo sob eucalipto apresentou maior diversidade quando comparado ao solo sob de Floresta Nativa. / Abstract: Traditional methods of isolation and culture limits the analyses of the microbian diversity in the environment, because it is believed - that approximately 10% of these microrganisms can be cultivated. The molecular microbian ecology have had recent progress through the construction of metagenomics Iibraries, what constitutes a powerful boarding to explore the microbian diversity of soil, also supplying information on the microrganisms that cannot be cultivated on this habitat. This work had the objective of stimate the diversity of bacterial communities, in two areas, an area of Native Forest (SFN) and the another one under eucalypts (SAE) of the same region. Using specific oligonucleotides starters, the gene 16S rRNA was amplified by PCR, amplicons had been cloned in pGEMR-T and the obtained clones were partial/y sequenced. In the soil SFN, 231 clones had been analyzed and 248 clones in the soil SAE. The sequences obtained were submitted to the nucleotides similarity analysis with the GenBank database. The bacterial phylum that was more evident in the two types de soil were Acidobacteria and Proteobacteria. In the soi! SFN the bacteria of the phylum Bacteroidetes were evident and in the soil SRE high frequency of the Actinobacteria bacteria was observed. Phylogenetic analyses reveled an extensive diversity in both soils, verified through diversity index differences in the bacterial communities in both areas, and that the area under eucalypt presented more diversity in relation to the area of Native Forest. / Orientadora: Lúcia Maria Carareto Alves / Coorientadora: Eliana Gertrudes Macedo Lemos / Banca: Janete Apparecida Desiderio Sena / Banca: Uderlei Doniseti Silveira Covissi / Mestre

Orientador: Jackson Antônio Marcondes de Souza / Banca: Alessandro de Mello Varani / Banca: Maria de Lourdes Teixeira de Moraes Polizeli / Resumo: Indústrias e governos buscam pela obtenção de biocombustível celulósico, entretanto a desconstrução do arranjo lignocelulósico para obtenção de açúcares livres de forma eficiente e economicamente viável ainda é um grande desafio. Avanços conquistados através da metagenômica ressaltam sua aplicação como alternativa para compreender e desvendar o grande potencial metabólico presente nos ambientes a fim de superar tais desafios. Contudo, ainda são poucos os trabalhos voltados à cana-de-açúcar. O solo trata-se da maior fonte microbiana de todo o planeta, sendo essa biodiversidade ainda desconhecida, esta comunidade microbiana fornece um ponto de partida para a exploração de novas enzimas responsáveis para a degradação de biomassa vegetal. Este trabalho buscou acessar o panorama do perfil taxonômico e funcional do metagenoma da comunidade microbiana presente em solo sob cultivo de cana-de-açúcar contendo palhada sobre o mesmo, possuindo como foco enzimas envolvidas na hidrólise de materiais lignocelulóscicos. Os dados encontrados demonstraram uma variedade de famílias enzimáticas envolvidas na degradação do complexo lignocelulósico. Verificou-se a presença de 27 famílias de Glycoside Hydrolases (GH), 21 famílias de Carbohydrate-Binding Module (CBM), 11 famílias de Carbohydrate Esterases (CE) e 4 famílias de Auxiliary Activities (AA). Os filos mais abundantes neste ambiente foram Proteobacteria e Actinobacteria, possuindo o último reconhecida importância industrial dadas suas enzim... (Resumo completo, clicar acesso eletrônico abaixo) / Abstract: Industries and governments seek for obtaining cellulosic biofuel, but the deconstruction of lignocellulosic arrangement to obtain free sugars efficiently and economically viable is still a big challenge. Advances made by metagenomics emphasize their application as an alternative to understand and unravel the great metabolic potential present in the environment in order to overcome such challenges. However, there are few studies related to sugarcane. The soil is the largest source of microbial whole planet, and its biodiversity still unknown, this microbial community provides a starting point for the exploration of new enzymes responsible for the degradation of plant biomass. This study aimed to access the overview of taxonomic and functional profile of the metagenome of soil under sugarcane cultivation containing straw on it, having focused on enzymes involved in the hydrolysis of lignocellulosic materials. The soil metagenome under sugarcane cultivation showed a variety of enzyme families involved in the degradation of the lignocellulosic complex. The presence of 27 Glycoside Hydrolases (GH) families, 21 Carbohydrate-Binding Module (CBM) families, 11 Carbohydrate Esterases (CE) families and 4 Auxiliary Activities (AA) families has been verified. Proteobacteria and Actinobacteria were the abundest phyla in this environment, which the last has recognized industrial importance given their enzymes with resistence to changes in pH and temperature. Such microbial community showed ... (Complete abstract click electronic access below) / Mestre

Orientadores: Valéria Maia de Oliveira, Suzan Pantaroto de Vasconcellos / Dissertação (mestrado) - Universidade Estadual de Campinas, Instituto de Biologia / Made available in DSpace on 2018-08-16T10:57:30Z (GMT). No. of bitstreams: 1 Dellagnezze_BrunaMartins_M.pdf: 2605800 bytes, checksum: 9f658e8eb7ad92524b8042afa7233e71 (MD5) Previous issue date: 2010 / Resumo: Bioemulsificantes ou biossurfactantes são compostos sintetizados por micro-organismos que aumentam a biodisponibilidade de hidrocarbonetos, promovendo a estabilização de substâncias com diferentes polaridades, como por exemplo, emulsões água-óleo existentes nos reservatórios de petróleo. As amostras utilizadas neste trabalho foram coletadas na Bacia Potiguar (RN), e empregadas como inóculo para a realização de enriquecimentos anaeróbios e aeróbios da microbiota do petróleo. A amostra de óleo selecionada (GMR 75) foi obtida a partir de um poço com nível de biodegradação moderado. Após o período de incubação dos enriquecimentos, foi realizada a extração de DNA dos mesmos e a construção das bibliotecas de fosmídios, utilizando-se kit de clonagem comercial. A avaliação da produção de emulsificantes pelos clones fosmidias provenientes das bibliotecas metagenômicas foi realizada através de dois tipos de ensaios de triagem, um teste baseado em atividade biológica e outro em similaridade de sequência. O teste funcional, denominado "colapso da gota", apresenta caráter qualitativo e foi utilizado para a triagem preliminar de 2.880 clones, gerando 2.104 resultados positivos. Para o teste baseado em seqüência, os clones que se mostraram positivos no teste funcional foram avaliados através de reações de PCR utilizando primers homólogos a regiões conservadas de genes para a síntese de biossurfactantes. Entretanto, nenhum sinal positivo de amplificação foi observado com os primers utilizados. Ensaios de tensiometria, utilizando meio mineral e hexadecano como única fonte de carbono e energia, foram realizados para a confirmação da produção dos emulsificantes pelos clones. Nove clones capazes de reduzir a tensão interfacial de 45 mN\m para 30 mN/m foram selecionados e submetidos aos ensaios de emulsificação utilizando petróleo, óleo diesel e hexadecano. Os resultados demonstraram a formação de emulsão, principalmente em óleo diesel, sugerindo poder emulsificante sobre este composto. As análises colorimétricas dos biossurfactantes produzidos pelos nove clones revelaram proporções similares de carboidratos, proteínas e ácidos urônicos, não permitindo resultados conclusivos sobre a composição destes compostos. / Abstract: Bioemulsifiers or biosurfactants are compounds produced by microorganisms which enhance the bioavailability of hydrophobic substances promoting the stabilization of substances with different polarities, like oil-in-water emulsion occurring in petroleum reservoirs. The samples used in this work were collected in the Potiguar Basin (RN, Brazil), and employed as inoculum for aerobic and anaerobic enrichments of the petroleum microbiota. The selected oil sample (GMR 75) was obtained from a reservoir presenting a moderate biodegradation level. After the incubation period of the microbial enrichments, total DNA was extracted and used for the assembly of the metagenomic libraries by using a comercial kit. The evaluation of the potential for emulsifier production by the fosmid clones derived from the metagenomic libraries was performed by means of two screening assays, one based on biological activity and another based on sequence similarity. The functional assay, named "drop collapse", has a qualitative character and was used for the preliminary screening of 2,880 clones, yielding 2,104 positive results. For the sequence-based assay, clones shown to be positive in the functional assay were evaluated through PCR reactions using primers homologous to conserved regions of genes for biosurfactant synthesis. However, no positive amplification signal was observed with the primers used. Tensitometric assays, using mineral medium and hexadecane as sole carbon and energy source, were performed to confirm the production of the biosurfactants by the clones. Nine clones able to reduce the interfacial tension from 45 mN\m to 30 mN/m were selected and subsequently submitted to emulsification assays using petroleum, diesel oil and hexadecane. Results demonstrated the formation of an emulsion, mainly in diesel oil, suggesting the emulsifier power of the biosurfactants over this substance. Colorimetric analyses of the biosurfactants produced by the clones revealed similar proportion of carbohydrates, proteins and uronic acids, not allowing conclusive characterization of their composition. / Mestrado / Genetica de Microorganismos / Mestre em Genética e Biologia Molecular

Polihidroxialcanoatos (PHA) são poliésteres acumulados por diversas bactérias a partir de fontes renováveis e são termoplásticos, biodegradáveis e biocompatíveis. A variabilidade da composição monomérica de PHA determina suas propriedades mecânicas e permite seu uso em diversas aplicações. A PHA sintase é a enzima responsável pela polimerização do PHA. O objetivo deste trabalho foi à busca por genes codificadores desta enzima, construção e avaliação de recombinantes portando tais genes. Inicialmente buscaram-se novos genes de PHA sintase a partir de clones de uma biblioteca metagenômica previamente detectados por PCR como positivos para algum tipo de PHA sintase. Posteriormente buscou-se PHA sintases de classe III e construiram-se recombinantes de Pseudomonas sp e B. sacchari pela introdução de genes de C. vinosum (phaECCv). Nas duas recombinantes, os genes inseridos foram capazes de aumentar a fração de 3HHx em relação a linhagem selvagem, quando se utilizou glicose e hexanoato como fontes de carbono. / Polyhydroxyalkanoates (PHA) are polyesters accumulated from renewable sources by several bacteria and are thermoplastic, biodegradable and biocompatible. The variability of the PHA monomer composition determines its mechanical properties and allows their use in many applications. PHA synthase is the enzyme responsible for the polymerization of PHA. The objective was to search for genes encoding this enzyme, construction and the assessment of recombinant bacteria carrying such genes. Initially a screening of PHA synthase genes was made from a metagenomic library clones previously identified as positive by PCR for any type of PHA synthase. Later a search for class III PHA synthases was made as a construction of a recombinant Pseudomonas sp and B. sacchari by introducing genes of C. vinosum (phaECCv). In the two recombinant strains, the genes inserted were able to increase the fraction of 3HHx compared to the wild strain when glucose and hexanoate was used as carbon sources.

The availability of high-throughput sequencing technologies has enabled metagenomicinvestigations into complex bacterial communities with unprecedented resolution andthroughput. The production of dedicated data sets for metagenomic analyses is, however, acostly process and, frequently, the first research questions focus on the study species itself. Ifthe source material is represented by fecal samples, target capture of host-specific sequencesis applied to enrich the complex DNA mixtures contained within a typical fecal DNA extract.Yet, even after this enrichment, the samples still contain a large amount of environmentalDNA that is usually left unanalysed. In my study I investigate the possibility of using shotgunsequencing data that has been subjected to target enrichment for mtDNA from the hostspecies, Grauer’s gorilla (Gorilla beringei graueri), for further analysis of the microbialcommunity present in these samples. The purpose of these analyses is to study the differencesin the bacterial communities present within a high-altitude Grauer’s gorilla, low-altitudeGrauer’s gorilla, and a sympatric chimpanzee population. Additionally, I explore the adaptivepotential of the gut microbiota within these great ape populations.I evaluated the impact that the enrichment process had on the microbial community by usingpre- and post-capture museum preserved samples. In addition to this, I also analysed the effectof two different extraction methods on the bacterial communities.My results show that the relative abundances of the bacterial taxa remain relatively unaffectedby the enrichment process and the extraction methods. The overall number of taxa is,however, reduced by each additional capture round and is not consistent between theextraction methods. This means that both the enrichment and extraction processes introducebiases that require the usage of abundance-based distance measures for biological inferences.Additionally, even if the data cannot be used to study the bacterial communities in anunbiased manner, it provides useful comparative insights for samples that were treated in thesame fashion.With this background, I used museum and fecal samples to perform cluster analysis to explorethe relationships between the gut microbiota of the three great ape populations. I found thatpopulations cluster by species first, and only then group according to habitat. I further foundthat a bacterial taxon that degrades plant matter is enriched in the gut microbiota of all threegreat ape species, where it could help with the digestion of vegetative foods. Another bacterialtaxon that consumes glucose is enriched in the gut microbiota of the low-altitude gorilla andchimpanzee populations, where it could help with the modulation of the host’s mucosalimmune system, and could point to the availability of fruit in the animals diet. In addition, Ifound a bacterial taxon that is linked with diarrhea in humans to be part of the gut microbiotaof the habituated high-altitude gorilla population, which could indicate that this pathogen hasbeen transmitted to the gorillas from their interaction with humans, or it could be indicative ofthe presence of a contaminated water source.