Tez (Yüksek Lisans) - Süleyman Demirel Üniversitesi, Sağlık Bilimleri Enstitüsü, Tıbbi Biyoloji ve Genetik Anabilim Dalı, 2005. / Bibliyografya var.

The phylogenetic reconstruction has noted great development in recent decades. The development of computers and device for sequencing biopolymers have been an enormous amount od phylogenetic data from different sources and different types. The scientists are trying to reconstruct a comlet tree of life from these data. The phylogenetic supertree are theoretically this option because a supertree alow a combination of all information gathered so far – in contras to the phylogenetic trees. This thesis present the method of reconstruction supertrees using average konsensus method.

The Phylogenetic reconstruction has seen great development in the last 30 years. Computers have become more powerful and more generally accessible, and computer algorithms more sophisticated. It comes the effort of scientists to reconstruct the entire tree of life from a large amount of phylogenetic data. Just for this purpose are formed phylogenetic supertrees that allow the combination of all information gathered so far. The aim of this work is to find a method to construct supertree that will give correct results.

Microbial Source Tracking (MST) is a field of study that attempts to identify the source of fecal contamination in waterways in order to assist with development of remediation strategies. Biologists at Cal Poly Center for Applications in Biotechnology (CAB) are developing a new MST method using microbes from the genus Bacteroides. Bacteroides species are host-specific microorganisms that can theoretically be used to trace back to a single host species. After fecal samples are collected, biologists use Next-Generation Sequencing (NGS) techniques to obtain only the genetic sequences of microorganisms belonging to the phylum Bacteroidetes. Investigators hypothesize that similar sequences belong to the same phlyogenetic group (i.e., the same genus) and can therefore be computationally clustered. Each cluster of related sequences, typically 97% similar, is called an Operational Taxonomic Unit (OTU). Theoretically, an OTU acts as a molecular signature that can be traced back to a specific host genus. This thesis presents LOTUS, the Library of OTUs, a web-based computational tool for the preliminary investigation of the use of the Bacteroides OTU library as an MST method. This work discusses the four contributions of LOTUS: a database design which accurately models OTUs and the underlying relationships necessary for source tracking, a pipeline to create OTUs from raw sequencing reads, a method of assigning taxonomy to OTUs, and a web-based user interface. In preliminary testing for a reference library of twelve samples, LOTUS produced 1,431 OTUs, of which 891 were single-source (OTUs derived from sequences from a single host species). Using these OTUs, LOTUS was able to accurately taxonomically match four of five unknown test samples, showing promise for using OTUs as an MST method.

The topic of my diploma thesis is the statistical evaluation of biological sequences with the help of phylogenic trees. In the theoretical part we will create a literary recherche of estimation methodology concerning the course of phylogeny on the basis of the similarity of biological sequences (DNA and proteins) and we will focus on the inaccuracies of the estimation, their causes and the possibilities of their elimination. Afterwards, we will compare the methods for the statistical evaluation of the correctness of the course of phylogeny. In the practical part of the thesis we will suggest algorithms that will be used for testing the correctness of the phylogenic trees on the basis of bootstrapping, jackknifing, OTU jackknifing and PTP test which are able to the capture phylogenic tree with the method neighbor joining from the biological sequences in FASTA code. It is also possible to change the distance model and the substitution matrix. To be able to use these algorithms for the statistical support of phylogenic trees we have to verify their right function. This verification will be evaluated on the theoretical sequences of the amino acids. For the verification of the correct function of the algorithms, we will carry out single statistical tests on real 10 sequences of mammalian ubiquitin. These results will be analysed and appropriately discussed.

Microorganisms are all around us. Some of them even live in our body and are essential for our healthy being. Study of microbial communities based on their genetic content has become very popular with the development of new technologies, which enable easy reading of DNA or RNA. The key role of these studies is usually to characterize significant microbial patterns of an environment. However, currently used visualization tools have many drawbacks for such analyses. The subject of this thesis is to design a R/Bioconductor package for simple creation of bipartite graphs from microbial data. This type of visualization brings many advantages for microbiome analysis. Benefits of bipartite graphs are further demonstrated by analysis of main parameters affecting computer processing of microbial data.

L’objectif principal de ce travail était de réaliser une étude écotoxicologique du complexe lac Ichkeul-lagune de Bizerte. Un recensement des pesticides agricoles les plus utilisés dans le bassin versant agricole a été effectué afin de déterminer les pesticides les plus problématiques. Le recensement effectué auprès des agriculteurs, des revendeurs de produits phytosanitaires et le commissariat régional de développement agricole de Bizerte a permis d’identifier les molécules les plus utilisées au niveau des zones agricoles du bassin versant du complexe lac Ichkeul – lagune de Bizerte: l’iodosulfurone, le mésosulfurone, le 2,4D, le glyphosate et le fénoxaprop comme matières actives des herbicides ; le tébuconazole et l’époxiconazole comme fongicide et la deltaméthrine matières active des insecticides. Ensuite, les concentrations en pesticides, HAP et PCB ont été déterminées. Les résultats ont montré une contamination au niveau de certains sites du complexe. Les stations situées dans la lagune de Bizerte ont des teneurs en HAP dépassant les limites tolérées. Les concentrations en pesticides dépassent les normes au niveau de quelques sites du lac Ichkeul. Enfin, l’impact de ces polluants sur les communautés microbiennes et la densité de nématodes a été déterminé. La CCA combinant les résultats des analyses chimiques et les données T-RFLP des communautés bactériennes met en évidence que les communautés bactériennes du lac sont influencées parles pesticides alors que celles de la lagune sont influencées par les HAP. La répartition des T-RFs montre la présence de T-RFs spécifiques dans les sites contaminés : le T-RF 114 bp caractérise le site le plus contaminé en pesticide, les microorganismes associés à ce T-RF correspondent probablement à des microorganismes résistants aux pesticides et capables de les dégrader. La composition des communautés bactériennes a été déterminée par pyroséquençage 454 sur les sites les plus représentatifs : le moins contaminé du complexe, le plus contaminé en HAP, le plus contaminé en pesticides et un site présentant une contamination multiple HAP-Pesticides. L’impact de la présence des pesticides sur la densité des nématodes libres marins a été étudié. L'analyse taxinomique des données de pyroséquençage regroupe les séquences dans 44 embranchements différents. En général, Proteobacteria étaient l’embranchement le plus dominant avec une prédominance de Gammaproteobacteria, Alphaproteobacteria et deltaproteobacteria dans les sédiments. Les communautés bactériennes des quatre sites ont 211 OTU commun. Le lac Ichkeul a été caractérisé par 219 OTU spécifique et la lagune de Bizerte 235 OTU. Le Site 2, le plus contaminé en pesticides présente l’abondance la plus élevée en nématodes libres marins. / The main objective of this work was to conduct an ecotoxicological study of the complex Ichkeul Lake-Bizerta lagoon. A survey of the most used agricultural pesticides in the watershed of the complex was conducted to determine the most problematic compounds. The survey of pesticides use with farmers, dealers of pesticides and the Regional Commissioner for Agricultural Development Bizerte has helped to identify the most used pesticides in agricultural areas of the watershed of the complex Ichkeul Lake-Bizerta lagoon: iodosulfuron, the mesosulfurone, 2,4D, glyphosate and fenoxaprop as herbicides active materials, tebuconazol epoxiconazol as fungicide active materials and deltamethrin as insecticides. Then, the concentrations of pesticides, PAHs and PCBs were determined in the sediment of the complex. The results showed a contamination of some complex sites. Stations located in the lagoon of Bizerte have a level of PAH in excess of permissible limits. Pesticide concentrations exceed the standards at some sites from the Ichkeul Lake. Finally, the impact of these pollutants on microbial communities and nematode density was determined. CCA combines the results of chemical analyzes and T-RFLP data of bacterial communities shows that the bacterial communities of the lake are influenced by pesticides while those of the lagoon are influenced by PAHs. The distribution of T-RFs showed the presence of specific T-RFs in contaminated sites: the T-RF 114 bp characterizes the most contaminated site in pesticides, microorganisms associated with the T-RF are probably microorganisms resistant to pesticides and capable of degrading this pollutant. The composition of bacterial communities was determined by 454 pyrosequencing of the most representative sites: the least contaminated of the complex, the most contaminated by PAH, the most contaminated by pesticides and a site with multiple PAH-pesticides contamination. The impact of the presence of pesticides on the density of free living marine nematodes was studied. The taxonomic analysis of the pyrosequencing data grouped the sequences into 44 different phyla. In general, Proteobacteria were the most dominant phyla with predominance of Gammaproteobacteria, Alphaproteobacteria and Deltaproteobacteria within the sediments. Besides Proteobacteria, there are a number of sequences affiliated to the following major phyla detected in all four sites: Chloroflexi, Bacteroidetes, Nitrospirae, Planctomycetes, Actinobacteria, Gemmatimonadetes, Firmicutes, Cyanobacteria, Spirochaetes, Acidobacteria. The bacterial communities of the four sites shared 211 common OTUs. The Lake Ichkeul was characterized by 219 specific OTUs and the Bizerta lagoon by 235 OTUs. Site 2, the most contaminated by pesticides has the highest abundance of free living marine nematodes.

Les champignons mycorhiziens arbusculaires (CMA) appartiennent au phylum Glomeromycota. Ces champignons forment une association symbiotique (obligatoire pour les champignons) avec plus de 80 % des espèces de plantes vasculaires, incluant la plupart des plantes cultivées. Cette symbiose mycorhizienne arbusculaire, très répandue dans la nature, a été retrouvée dans presque tous les écosystèmes terrestres aussi bien dans des déserts chauds et froids que dans des forêts tropicales ou tempérées. L’emploi d’outils de biologie moléculaire et des fossiles ont permis d’estimer la date de l’origine de la symbiose mycorhizienne arbusculaire à environ 450 millions d’années avant notre ère. Cette date coïncide avec l’apparition des premières plantes terrestres primitives. Il existe plusieurs types de mycorhizes impliquant différents champignons et différentes plantes hôtes. Ces différents faits suggèrent que les plantes vasculaires et les champignons mycorhiziens à arbuscules ont co-évolué, et que les Glomeromycota ont influencé la capacité des plantes à coloniser le milieu terrestre. Plusieurs études ont montré que les contaminants organiques peuvent influer négativement cette relation mutuellement bénéfique qu'entretiennent la plante et le champignon conduisant à une diminution de la croissance du CMA mais aussi à une modification des interactions au sein de la communauté de CMA elle-même. Ainsi, mon projet de maîtrise vise à quantifier la diversité mycorhizienne arbusculaire de deux espèces de plante Eleocharis obtusa et Panicum capillare qui poussent dans un bassin hautement contaminé par des hydrocarbures pétroliers de façon très hétérogène et étudier les structures des communautés des CMA dans des échantillons de sol rhizosphérique et de racines de celles-ci. L'un des résultats important de mon étude est la découverte d’une très grande diversité des CMA associés à E. obtusa et P. capillare dans ce milieu contaminé. En effet, pas moins de 26 unités taxonomiques opérationnelles (OTU) de champignons mycorhiziens à arbuscules ont été recensés. La quasi-totalité des familles de Glomeroycota sont représentées à l'exception de Ambisporaceae, Geosiphonaceae et Pacisporaceae. Ainsi, le genre Diversispora domine la communauté des CMA par son abondance relative, cependant le genre Claroideoglomus est le plus abondant des genres des CMA. Ensuite, mon projet a déterminé l'influence de la plante hôte sur la composition de la communauté de CMA. D'après l'analyse de la diversité alpha, les plantes présentent une structure de diversité différente. Les valeurs de Local Contributions to beta Diveristy (LCBD) élevées étaient souvent liées à un site avec un grand nombre de taxons de CMA. En utilisant une analyse en composante principale (ACP), j’ai constaté que les échantillons récoltés en 2011 auraient sensiblement la même composition en OTU. J’ai pu observer également une variation de la communauté des OTU de CMA d'une année à l'autre pour les plantes. La rhizosphère des plantes récoltées la même année avait une communauté d'OTU similaire globalement, pourtant au niveau racinaire, j’ai observé une spécificité chez des certains OTU. Cette étude fournit une vue d’ensemble de la diversité des CMA associés à des végétaux localisés dans les sédiments très polluées. Elle suggère aussi que les CMA peuvent être des candidats microbiens potentiellement importants dans la restauration des sols contaminés par le pétrole. / The arbuscular mycorrhizal fungi (AMF) belong to phylum Glomeromycota, a group of early divergent fungal lineage. These fungi form a mutualistic symbiotic association with more than 80% of vascular plant species, including most important crops. The arbuscular mycorrhizal symbiosis is widespread and has been found in almost all terestrial ecosystems on earth such as hot and cold deserts, as well as tropical and temperate forests. The use of molecular biology tools and fossil records dated back the origin of arbuscular mycorrhizal symbiosis to 460 million years ago, which coincides with the emergence of terrestrial plants. There are several types of mycorrhizas depending on fungal groups and host plants. It has been suggested that vascular plants and AMF have co-evolved and that Glomeromycota helped plant ancestors to colonize the land. It has been reported that organic contaminants can negatively impact this mutually beneficial symbiosis between plant and fungi reducing the growth of the AMF and changing the structure of AMF communities. The objective of my master project was to determine the diversity of arbuscular mycorrhizal fungi associated with two plant species, Eleocharis obtusa and Panicum capillare, which spontaneously growing a highly petroleum hydrocarbons contaminated basin with an heterogeneous pattern of contamination, and to describe the structures of their communities in their rhizosphere soils and roots. My results revealed an unexpected high diversity of AMF OTUs associated with two plant species. Indeed, we identified 26 OTUs of AMF. This diversity covered the most important families of Glomeroycota except Ambisporaceae, Geosiphonaceae and Pacisporaceae. Genus Diversispora dominated the AMF community by its relative abundance, however the genus Claroideoglomus was the most represented in the samples. My study determined the influence of the host plant on the composition of the AMF community. Based on the analysis of the α diversity, we found that the rhizospheric soil and root samples showed different community structures between the two plants species. β diversity analysis showed that high Local Contributions to β Diveristy (LCBD) values (high uniqueness of species composition) were often linked to the sample with a large number of AMF taxa. Using a principal component analysis (PCA), we found that the samples collected in 2011 have approximately the same composition of OTUs. It is likely that a turnover of AMF OTU and 4 plants may occur from one year to another. This study provides an overview of the diversity of AMF associated with plants located in heavily polluted sediments. It also suggests that AMF may be potentially important microbial candidates in the remediation of oil-contaminated soils.

The master's thesis provides a comprehensive overview of resampling methods for testing the correctness topology of the phylogenetic trees which estimate the process of phylogeny on the bases of biological sequences similarity. We focused on the possibility of errors creation in this estimate and the possibility of their removal and detection. These methods were implemented in Matlab for Bootstrapping, jackknifing, OTU jackknifing and PTP test (Permutation tail probability). The work aims to test their applicability to various biological sequences and also to assess the impact of the choice of input analysis parameters on the results of these statistical tests.

A thesis submitted to the School of Graduate Studies in partial fulfillment of the requirements for the degree Master of Science / The effect of changing environmental conditions on microbial population structure can be observed at both the species and community level. Within the Escherichia coli species, null mutations in the RpoS stationary phase regulator are commonly selected by growth on poor carbon sources. In contrast, mutations which restore RpoS function may provide a selective advantage for cells exposed to environmental stress. The loss and subsequent restoration of RpoS form a population-level switch for adaptation within poor carbon and high stress environments. To investigate selection for RpoS reversion, we exposed rpoS-deficient E. coli to high salt concentrations and assessed the phenotype of presumptive mutants. 3-9% of salt-resistant mutants contained reversion mutations within rpoS, while in 91-97% the loss of RpoS function was maintained and mutations at alternative gene loci were identified. These results show that RpoS function can be restored in deficient E. coli under selective pressure. At the community level, the application of next-generation sequencing (NGS) technology to characterize environmental microbial diversity can potentially augment traditional water quality monitoring methods. To investigate the use of NGS in identifying microbial taxa within the Niagara Region, we collected water samples from Lake Erie, Lake Ontario, and nearby areas and examined the metagenome of microbial communities. A QIIME (Quantitative Insights Into Microbial Ecology) analysis of sequence data identified significant differences in relative microbial abundance with respect to sample metadata (e.g. location and subtype), significant correlations between relative abundance and quantitative parameters (e.g. Escherichia coli counts and fecal DNA markers), and detected pathogen-containing taxa at a relative abundance of 0.1-1.5%. These results show that sequence-based analyses can be used in conjunction with traditional identification methods to profile the metagenomic community of environmental samples and predict water quality. Both within-species and community-wide analyses thus offer insight into how microbial populations respond and adapt to environmental fluctuations. / Thesis / Master of Science (MSc) / The effect of changing environmental conditions on microbial population structure can be observed at both the species and community level. Within the Escherichia coli species, we investigated reversion of loss of function mutations in the RpoS protein regulator in high salt conditions and identified RpoS restoration under selective pressure. At the community level, we examined the microbial DNA of water samples from the Niagara Region under select environmental conditions and assessed the viability of next-generation sequencing in augmenting traditional water quality monitoring methods. Both within-species and community-wide analyses offer insight into how microbial populations respond and adapt to environmental fluctuations.