La caractérisation exaustive des variations de l'ADN peut aider à progresser dans de nombreux champs liés à la génomique du cancer. Le séquençage nouvelle génération (NGS en anglais pour Next Generation Sequencing) est actuellement la technique la plus efficace pour déterminer une séquence ADN, du aux faibles coûts et durées des expériences comparé à la méthode de séquençage traditionnelle de Sanger. Cependant, la détection de mutations à partir de données NGS reste encore un problème difficile, en particulier pour les mutations somatiques présentes en très faible abondance comme lorsque l'on essaye d'identifier des mutations sous-clonales d'une tumeur, des mutations dérivées de la tumeur dans l'ADN circulant libre, ou des mutations somatiques dans des tissus normaux. La difficulté principale est de précisement distinguer les vraies mutations des artefacts de séquençage du au fait qu'ils atteignent des niveaux similaires. Dans cette thèse nous avons étudié la nature systématique des erreurs dans les données NGS afin de proposer des méthodologies efficaces capables d'identifier des mutations potentiellement en faible abondance. Dans un premier chapitre, nous decrivons needlestack, un nouvel outil d'appel de variants basé sur la modélisation des erreurs systématiques sur plusieurs échantillons pour extraire des mutations candidates. Dans un deuxième chapitre, nous proposons deux méthodes de filtrage des variants basées sur des résumés statistiques et sur de l'apprentissage automatique, dans le but de d'améliorer la précision de la détection des mutations par l'identification des erreurs non-systématiques. Finalement, dans un dernier chapitre nous appliquons ces approches pour développer des biomarqueurs de détection précoce du cancer en utilisant l'ADN circulant tumoral / Comprehensive characterization of DNA variations can help to progress in multiple cancer genomics fields. Next Generation Sequencing (NGS) is currently the most efficient technique to determine a DNA sequence, due to low experiment cost and time compared to the traditional Sanger sequencing. Nevertheless, detection of mutations from NGS data is still a difficult problem, in particular for somatic mutations present in very low abundance like when trying to identify tumor subclonal mutations, tumor-derived mutations in cell free DNA, or somatic mutations from histological normal tissue. The main difficulty is to precisely distinguish between true mutations from sequencing artifacts as they reach similar levels. In this thesis we have studied the systematic nature of errors in NGS data to propose efficient methodologies in order to accurately identify mutations potentially in low proportion. In a first chapter, we describe needlestack, a new variant caller based on the modelling of systematic errors across multiple samples to extract candidate mutations. In a second chapter, we propose two post-calling variant filtering methods based on new summary statistics and on machine learning, with the aim of boosting the precision of mutation detection through the identification of non-systematic errors. Finally, in a last chapter we apply these approaches to develop cancer early detection biomarkers using circulating tumor DNA

Les leucémies aiguës myéloïdes (LAM) du sujet âgé sont les plus fréquentes des leucémies aiguës. Bien que de physiopathologie hétérogène, elles partagent un pronostic défavorable. L’azacitidine est devenue un des traitements de référence pour les patients jugés inéligibles pour une chimiothérapie intensive mais les critères de sélection des patients entre ces deux approches sont controversés. L’identification de biomarqueur prédictif de réponse à l’azacitidine doit permettre de rationnaliser ce choix thérapeutique. Les facteurs pronostiques classiques d’une cohorte de 334 patients atteints de LAM manquent de précision pour guider la meilleure stratégie pour un patient donné. A partir du séquençage de 224 patients traités par azacitidine, nous montrons un impact défavorable des mutations de TP53 sur la survie globale, quel que soit leur caractérisation fonctionnelle. Le séquençage des exomes de 49 patients selon leur réponse à l’azacitidine (26 répondeurs et 23 non répondeurs), suivi du re-séquençage ciblé de 4 polymorphismes chez 175 patients a montré un impact positif du polymorphisme rs7622799 de MECOM sur la survie globale sous azacitidine. / Elderly patients with acute myeloid leukemias (AML) represent the most frequent acute leukemias. Although they differ in their pathophysiology, they all share an adverse prognosis. Azacitidine has become one of the reference low-intensity frontline therapy for patients deemed unfit for intensive chemotherapy. Patients selection between these 2 options remains controversial. Predictive biomarkers of response to azacitidine should allowed to rationalize this decision making. Classical prognosis factors of a cohort of 334 newly diagnosed AML lack of precision to determine the optimum strategy for any individual patient. By sequencing of a 224-patients series of azacitidine-treated AML patients, we demonstrate an adverse impact of TP53 mutation on overall survival, irrespective of the functional characterization of p53 mutants. Exome sequencing of 49 patients with extreme phenotype as defined by their response under azacitidine (26 responders versus 23 non-responders), followed by targeted sequencing of 4 common polymorphisms in a validation set of 175 patients, showed a positif impact of MECOM rs7622799 on overall survival.

Orientador: Semíramis Guimarães Ferraz Viana / Resumo: Giardia duodenalis é um dos principais agentes etiológicos de diarreia infecciosa, sobretudo em crianças em idade pré-escolar que vivem em comunidades de baixa renda. Estudos da diversidade genética de G. duodenalis ampliaram o conhecimento da epidemiologia nas infecções humanas, entretanto um dos temas mais interessantes e menos conhecidos é a possível interação de Giardia com o microbioma do hospedeiro e com patógenos concomitantes. No presente estudo, avaliou-se a composição e a diversidade da comunidade bacteriana de crianças saudáveis e crianças com diarreia, parasitadas por Giardia e outros protozoários intestinais. Os isolados de Giardia obtidos nessa população foram caracterizados geneticamente. Amostras de fezes foram obtidas de 181 crianças de 0 a 6 anos de idade, das quais 156 crianças hígidas atendidas em centros de educação infantil e 25 crianças com diarreia atendidas no PS Infantil Municipal. Cada amostra de fezes foi processada para o exame microscópico e submetida à extração de DNA a ser empregado em duas etapas distintas: (1) amplificação e sequenciamento Sanger para a caracterização genética de Giardia e o diagnóstico de Blastocystis sp, Dientamoeba fragilis, Enterocytozoon bieneusi e Cryptosporidium spp. e (2) amplificação do gene 16s RNA ribossomal e sequenciamento de nova geração (plataforma Illumina MiSeq) para a caracterização da microbiota intestinal. Giardia (36,5%) e Blastocystis (41,7%) foram os parasitas mais prevalentes. A caracterização genética... (Resumo completo, clicar acesso eletrônico abaixo) / Abstract: Giardia duodenalis is one of the major etiological agents of infectious diarrhea, especially in preschool children living in low-income settings. Studies focused on genetic diversity of G. duodenalis have provided insights for a better understanding of epidemiology in human infections. However, one of the most interesting and least known issues is the possible interplay between Giardia and the host microbiome and concomitant pathogens. In this work, we evaluated the diversity and composition of bacterial community of healthy children and children presenting with diarrhea infected by Giardia and/or other intestinal protozoa. In addition, Giardia isolates infecting this population were genotyping. A total of 181 stool samples from children aged 0 to 6 years old (156 from daycare children and 25 from diarrheic children attending in an emergence pediatric center) were tested by microscopic examination and submitted to DNA extraction for the following steps: (1) conventional PCR/sequencing for Giardia genotyping and the diagnosis of Blastocystis sp, Dientamoeba fragilis, Enterocytozoon bieneusi and Cryptosporidium spp and (2) next-generation sequencing (Illumina MiSeq) based analysis of intestinal microbiota. Giardia (36.5%) and Blastocystis (41.7%) were the most prevalent parasites. Analysis of Giardia sequences retrieved from 61 isolates revealed infections by assemblages A (31%), B (69%) and mixed infections A+B (3%). Metagenomic analyzes revealed similarity of bacterial microb... (Complete abstract click electronic access below) / Doutor

The Brigham Young University (BYU) Radio Astronomy Systems group, in collaboration with the National Radio Astronomy Observatory (NRAO), the Center for Astrophysics at West Virginia University (WVU), and the Green Bank Observatory (GBO) have developed, and commissioned, a broadband real-time digital back end processing system for a 38-element phased array feed (PAF) with 150 MHz of instantaneous bandwidth. This system is capable of producing coarse and fine channel correlations, and implements a real-time beamformer that forms 7 simultaneous dual-polarized beams. This thesis outlines the hardware and software development for the digital back end and presents on-telescope commissioning results. This system has been measured to provide an unprecedented low Tsys/η noise level of 28 K and can perform maps of galactic hydrogen observations in a fraction of the time of a conventional single horn feed. The National Radio Astronomy Observatory (NRAO) has recently announced the concept and development of the next generation Very Large Array (ngVLA), a large interferometric array consisting of 300 radio telescopes and longest baseline (distance between a pair of antennas) of 300 km. Large interferometric arrays have been shown to attenuate radio frequency interference (RFI) because it is decorrelated as it propagates across long baselines. This is not always sufficient, especially with dense core array geometries and with the ever-increasing amount of strong RFI sources. Conventional RFI projection-based mitigation techniques have performed poorly on large interferometers because of covariance matrix estimation error due to decorrelation when identifying interference subspace parameters. This thesis presents an algorithm that overcomes the challenge of decorrelation by applying subspace projection via subarray processing (SP-SAP). Each subarray is designed to have a set of elements with high mutual correlation in the interferer for better estimation of subspace parameters. In simulation, compared to the former approach of applying subspace projection on the full array, SP-SAP improves mitigation of the RFI on the order of 9 dB. A signal of interest is shown then to be observable through the RFI in a full synthetic image.

Obsessive-compulsive disorder (OCD) and the spectrum of associated conditions, affect 2-4% of the population worldwide. Although heritability studies in OCD have shown a 3 - 12 times increased risk for first degree relatives, the identification of the underlying risk-conferring genetic variation using classic genetic association studies has proven to be difficult. The possibility of a larger contribution of rare genetic variants to the risk of psychiatric disorder has been suggested by several successful studies. We expect that a spectrum of risk allele frequencies exists, which includes not only common variation but also a substantial amount of rare genetic variants that contribute to OCD. This thesis is aimed at identifying and functionally characterizing rare genetic variation in the OCD spectrum. Identified statistically significant variants were scrutinized for changes related to synaptic function using high content screening and subsequent functional analyses. Identifying the genetic profile of rare variants found in the OCD spectrum cohort combined with the functional impact that these variants have has provided insight into the etiology of the OCD spectrum. With these approaches a foundation can be laid for the development of a predictive model of the OCD spectrum.

The chimpanzee is humankind’s closest living relative and the two species diverged ~6 million years ago. Comparative studies of the human and chimpanzee genomes and transcriptomes are of great interest to understand the molecular mechanisms of speciation and the development of species-specific traits. The aim of this thesis is to characterize differences between the two species with regard to their genome sequences and the resulting transcript profiles. The first two papers focus on indel divergence and in particular, indels causing premature termination codons (PTCs) in 8% of the chimpanzee genes. The density of PTC genes is correlated with both the distance to the telomere and the indel divergence. Many PTC genes have several associated transcripts and since not all are affected by the PTC we propose that PTCs may affect the pattern of expressed isoforms. In the third paper, we investigate the transcriptome divergence in cerebellum, heart and liver, using high-density exon arrays. The results show that gene expression differs more between tissues than between species. Approximately 15% of the genes are differentially expressed between species, and half of the genes show different splicing patterns. We identify 28 cassette exons which are only included in one of the species, often in a tissue-specific manner. In the fourth paper, we use massive parallel sequencing to study the chimpanzee transcriptome in frontal cortex and liver. We estimate gene expression and search for novel transcribed regions (TRs). The majority of TRs are located close to genes and possibly extend the annotations. A subset of TRs are not found in the human genome. The brain transcriptome differs substantially from that of the liver and we identify a subset of genes enriched with TRs in frontal cortex. In conclusion, this thesis provides evidence of extensive genomic and transcriptomic variability between human and chimpanzee. The findings provide a basis for further studies of the underlying differences affecting phenotypic divergence between human and chimpanzee.

<p>Providing carrier grade characteristics for Internet Protocol (IP) communication applications is a significant problem for IP application providers in order to offer integrated services that span IP&nbsp / and telecommunication networks. This thesis addresses the provision of life-cycle management, which is only one carrier grade characteristic, for a SoftBridge application, which is an example of IP communication applications. A SoftBridge provides semi-synchronous multi-modal IP-based communication. The work related to IP-Telecommunication integrated services and the SoftBridge is analyzed with respect to life-cycle management in a literature review. It is suggested to use an Application Server in a Next Generation Network (NGN) to provide life-cyclemanagement functionality for IP-Telecommunication applications. In this thesis, the Application Server is represented by a JAIN Service Logic Execution Environment(JSLEE), in which&nbsp / a SoftBridge application can be deployed, activated, deactivated, uninstalled and upgraded online.Two methodologies are applied in this research: exploratory prototyping, which evolves the development of a SoftBridge application, and empirical comparison, which is concerned with the empirical evaluation of a SoftBridge application in terms of carriergrade capabilities. A SoftBridge application called SIMBA&nbsp / provides a Deaf Telephony service similar to aprevious Deaf Telephony SoftBridge, However, SIMBA&rsquo / s SoftBridge design and implementation are unique to this thesis. In order to test the life-cycle&nbsp / management ability of SIMBA, an empirical evaluation is carried out including the experiments oflife-cycle management and call-processing performance. The final experimental results of the evaluation show that a JSLEE is able to provide life-cycle management for SIMBA without causing a significant decrease in performance. In conclusion, the life-cycle management can be provided&nbsp / or a SoftBridge application by using an Application Server such as a JSLEE. Futhermore, the results indicate that&nbsp / approach of using Application Server (JSLEE) integration should be&nbsp / sufficiently general to provide life cycle management, and indeed other carrier grade capabilities, for other IP communication applications. This allows IP communication applications to be&nbsp / &nbsp / &nbsp / integrated into an NGN.</p>

Understanding the genetic basis of biological processes is a fundamental component of modern ecology and evolutionary biology studies. With the recent advent of next generation sequencing (NGS) technologies, it is now possible to perform large genome and transcriptome projects for ecologically important non-model species. In this thesis, I focused on the development and application of genomic resources of two non-model bird species, the black grouse (Tetrao tetrix) and the great snipe (Gallinago media). Using the chicken genome as a reference, I developed a reference guided NGS pipeline to assemble the complete draft genome of black grouse. The draft genome has a good coverage of the main 29 chromosomes of the chicken genome. The genome was used to develop a vast number of genetic markers. Comparing this genome with that of other species, I identified the genomic regions which were important for the lineage specific evolution of black grouse. I also sequenced and characterised the spleen transcriptome of the black grouse. I identified and validated a large number of gene-based microsatellite markers from the transcriptome and identified and confirmed the expression of immune related genes. Using a similar RNA-Seq approach, I also sequenced the blood transcriptomes of 14 great snipe males with different mating success. I identified genes and single nucleotide polymorphisms (SNPs) which might be related to male mating success in this species, both in terms of gene expression levels and genetic variation structure. For the immunologically important major histocompatibility complex (MHC) gene region of black grouse, I constructed a fosmid library and used it to sequence the complete core MHC region of this species. This resource allowed me to perform a comprehensive comparative genomics analysis of the galliform MHC, by which I found that some genes in this region were affected by selective forces. I was also able to develop a single locus genotyping protocol for the duplicated MHC BLB (class IIB) genes and found that the two black grouse BLB loci followed different evolutionary trajectories. This thesis set an example of developing genomic resources in non-model species and applying them in addressing questions relevant to ecology and evolutionary biology.

Esta tesis doctoral aborda el problema de la definición de movimientos rápidos sin interrupciones (seamless handoffs) en redes heterogéneas de próxima generación (NGNs) mediante definición de un proceso de distribución de claves seguro, que habilite un proceso de re-autenticación rápida a la vez que un acceso autenticado anónimo y que no se pueda trazar. Concretamente, el sistema de control de acceso desarrollado ofrece un conjunto de características que, hasta la fecha, no han confluido en una misma solución: (1) aplicable a las futuras redes NGN basadas en EAP; (2) reducción de la latencia introducida por el proceso de autenticación en entornos móviles, con independencia del tipo de handoff realizado por el usuario; (3) que el proceso cumpla fuertes requisitos de seguridad; (4) fácil despliegue en redes existentes; (5) compatibilidad con las actuales tecnologías estandarizadas; y (6) soporte de protección de privacidad del usuario. / This PhD thesis deals with the problem of defining fast movements without interruptions (seamless handoffs) in the next generation of heterogeneous networks. This objective is achieved through a secure key distribution process, which enables a fast re-authentication process providing both user anonymity and untraceability. The developed access control system offers a set of features not covered so far by a single solution: (1) applicable for EAP-based NGNs; (2) reduction of the authentication latency in mobile environments irrespective of the type of handoff performed by the user; (3) provision of strong security properties; (4) easy deployment in current networks; (5) compatibility with current standardized technologies; and (6) user privacy support.

As a result of rapid progress in research and development, today's wireless world exhibits several heterogeneous communication networks, such as cellular networks, satellite networks, wireless local area networks (WLAN), mobile ad hoc networks (MANET), and sensor networks. These networks are complementary to each other. Hence, their integration can realize a unified wireless system that has the best features of the individual networks. This has spurred much research interest in designing integrated next-generation of wireless systems (NGWS). While existing wireless networks have been extensively studied individually, the integrated wireless system brings new challenges in architecture design, system management, and protocol design. The different wireless networks use different communication technologies and are based on different networking paradigms. Therefore, it is challenging to integrate these networks such that their heterogeneities are hidden from each other and a harmonious inter-operation among them is achieved. The objective of this research is to design a scalable, secure, and robust architecture and to develop seamless mobility management protocols for NGWS. More specifically, an architecture that integrates the heterogeneous wireless systems is first proposed for NGWS. Next, a cross-layer (Layer 2 + 3) handoff management protocol is developed for NGWS. Afterward, analytical modeling is developed to investigate the handoff performance of the existing mobility management protocols for different types of applications. Finally, a framework for multi-layer mobility management is developed to support the seamless handoff support to all types of applications in NGWS.