The objective of this study was to characterize the mitochondrial genomes of the species within the genus Ceratocystis and investigate the evolutionary process of the ribosomal RNA cistron found within these fungi. Ceratocystis incorporates a number of pathogenic species affecting a variety of hosts, making the study of these fungi economically significant. The fortuitous identification of a Ceratocystis species, C. manginecans, which contained two different internally transcribed spacer sequence types within the ribosomal rRNA cistron, enabled a study of concerted evolution in this fungus. Using this non-model organism we were able to show empirical evidence for unequal crossing over and gene conversion as the ultimate forces acting on this gene region dictating a concerted evolutionary effect. We suggest that this process is true for all eukaryotes. Using the knowledge drawn from previously characterized and annotated mitochondrial genomes of other eukaryotes, the genomes of three Ceratocystis species, namely Ceratocystis fimbriata, Ceratocystis albifundus and Ceratocystis moniliformis were fully assembled and annotated for comparative analysis. This comparative study addressed the genome size, gene content, tRNA presence as well as intron types and their homing endonucleases found among these three mitochondrial genomes. An interspecies characterization was then undertaken using the mitochondrial genomes of six different C. albifundus isolates from different geographical locations in Africa. Genetic variation and similarities among these isolates supports the previous hypothesis that the origin of this fungus is Southern Africa. It is hoped that the research presented in this thesis will contribute to the improved understanding of the mitochondrial genomes in Ceratocystis species. / Thesis (PhD)--University of Pretoria, 2013. / gm2013 / Genetics / Unrestricted

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Mitochondrial DNA encodes thirteen protein subunits in the oxidative phosphorylation system (OXPHOS) that is responsible for cellular energy production. Mitochondrial disorders have been identified to be associated with mtDNA mutations. However, the molecular mechanisms of specific mtDNA mutations are still being explored in order to establish causative links. This study tries to elucidate the mutational effects of mtDNA on OXPHOS complex activities and cell growths. Using mouse 3T3 fibroblasts as a cell model, single-cell clones with different growth rates were isolated. The entire mtDNA genome was sequenced for mutations. The enzymatic activities of OXPHOS complex I to V were analysed. Three growth patterns represented by five clones were identified. Three clones (clone #2, #3, and #6) had the shortest doubling times (11.5 - 14.9 hours). Clone #1 had a medium growth rate (19.2 hous); and clone #5 had a significantly slow growth rate (22 hours). MtDNA sequencing results revealed that clone #5 had several heteroplasmic mutations (one in 16S rRNA, two in tRNAser (UCN), three in tRNAasp, one in tRNAlys, one in COI, five in COII, and one in ATPase8) while the other four clones showed sequence homology. Enzymatic analyses showed that on average clone #5 had significantly low complex III, IV, and V activities (p < 0.05). Changes in biochemical properties and protein structure were analyzed to deduct possible mechanisms for reduced respiration. In conclusion, the slow growth rate is associated with reduced OXPHOS enzyme functions. It is most likely that the combination of COI and COII mutations resulted in the reduction of complex IV function. It is still unclear whether the ATPase8 mutation (T7869A) in the non-conserved region alone can have such a pronounced phenotypic effect. A reduction in complex III also cannot be explained since there were no mutations in the only mtDNA-encoded complex III gene, but it is possible that there are mutations in the nDNA-encoded complex III genes. Mutations in tRNA and rRNA genes may also be responsible for reduced protein syntheses and consequently reduced OXPHOS activities. It is unclear why complex I activity was not affected. Although the mutational effect of individual mtDNA mutation observed cannot be clearly identified, this study establishes a correlation between mtDNA mutation and cell energy production and growth.

<p>While the many completely sequenced genomes of bacterial pathogens contain all the determinants of the host-pathogen interaction, and also every possible drug target and recombinant vaccine candidate, computational tools for selecting suitable candidates for further experimental analyses are limited to date. The overall objective of my PhD project was to attempt to design reusable systems that employ the two most important features of bacterial evolution, horizontal gene transfer and adaptive mutation, for the identification of potentially novel virulence-associated factors and possible drug targets. In this dissertation, I report the development of two novel technologies that uncover novel virulence-associated factors and mechanisms employed by bacterial pathogens to effectively inhabit the host niche. More importantly, I illustrate that these technologies may present a reliable starting point for the development of screens for novel drug targets and vaccine candidates, significantly reducing the time for the development of novel therapeutic strategies. Our initial analyses of proteins predicted from the preliminary genomic sequences released by the Sanger Center indicated that a significant number appeared to be more similar to eukaryotic proteins than to their bacterial orthologs. In order determine whether acquisition of genetic material from eukaryotes has played a role in the evolution of pathogenic bacteria, we developed a system that detects genes in a bacterial genome that have been acquired by interkingdom horizontal gene transfer.. Initially, 19 eukaryotic genes were identified in the genome of Mycobacterium tuberculosis of which 2 were later found in the genome of Pseudomonas aeruginosa, along with two novel eukaryotic genes.</p> <p>Surprisingly, six of the M. tuberculosis genes and all four eukaryotic genes in P. aeruginosa may be involved in modulating the host immune response through altering the steroid balance and the production of pro-inflammatory lipids. We also compared the genome of the H37Rv M. tuberculosis strain to that of the CDC- 1551 strain that was sequenced by TIGR and found that the organisms were virtually identical with respect to their gene content, and hypothesized that the differences in virulence may be due to evolved differences in shared genes, rather than the absence/presence of unique genes. Using this observation as rationale, we developed a system that compares the orthologous gene complements of two strains of a bacterial species and mines for genes that have undergone adaptive evolution as a means to identify possibly novel virulence &ndash / associated genes. By applying this system to the genome sequences of two strains of Helicobacter pylori and Neisseria meningitidis, we identified 41 and 44 genes that are under positive selection in these organisms, respectively. As approximately 50% of the genes encode known or potential virulence factors, the remaining genes may also be implicated in virulence or pathoadaptation. Furthermore, 21 H. pylori genes, none of which are classic virulence factors or associated with a pathogenicity island, were tested for a role in colonization by gene knockout experiments. Of these, 61% were found to be either essential, or involved in effective stomach colonization in a mouse infection model. A significant amount of strong circumstantial and empirical evidence is thus presented that finding genes under positive selection is a reliable method of identifying novel virulence-associated genes and promising leads for drug targets.</p>

<p>Despite the fact that the sequence of the human genome is now complete it has become clear that the elucidation of the transcriptome is more complicated than previously expected. There is mounting evidence for unexpected and previously underestimated phenomena such as alternative splicing in the transcriptome. As a result, the identification of novel transcripts arising from the genome continues. Furthermore, as the volume of transcript data grows it is becoming increasingly difficult to integrate expression information which is from different sources, is stored in disparate locations, and is described using differing terminologies. Determining the function of translated transcripts also remains a complex task. Information about the expression profile &ndash / the location and timing of transcript expression &ndash / provides evidence that can be used in understanding the role of the expressed transcript in the organ or tissue under study, or in developmental pathways or disease phenotype observed.<br /> <br /> In this dissertation I present novel computational approaches with direct biological applications to two distinct but increasingly important areas of research in gene expression research. The first addresses detection and characterisation of alternatively spliced transcripts. The second is the construction of an hierarchical controlled vocabulary for gene expression data and the annotation of expression libraries with controlled terms from the hierarchies. In the final chapter the biological questions that can be approached, and the discoveries that can be made using these systems are illustrated with a view to demonstrating how the application of informatics can both enable and accelerate biological insight into the human transcriptome.</p>

Viral metagenomics of feces collected from 58 Peruvian children with unexplained diarrhea revealed several small circular ssDNA genomes. Two genomes related to sequences previously reported in feces from chimpanzees and other mammals and recently named smacoviruses were characterized and then detected by PCR in 1.7 % (1/58) and 19 % (11/58) of diarrheal samples, respectively. Another three genomes from a distinct small circular ssDNA viral group provisionally called pecoviruses encoded Cap and Rep proteins with <35 % identity to those in related genomes reported in human, seal, porcine and dromedary feces. Pecovirus DNA was detected in 15.5 % (9/58), 5.9 % (3/51) and 3 % (3/100) of fecal samples from unexplained diarrhea in Peru, Nicaragua and Chile, respectively. Feces containing these ssDNA genomes also contained known human enteric viral pathogens. The cellular origins of these circular ssDNA viruses, whether human cells, ingested plants, animals or fungal foods, or residents of the gut microbiome, are currently unknown.

Orientador: Fernando Sebastián Baldi Rey / Coorientador: Gregório Miguel Ferreira de Camargo / Banca: Danisio Prado Munari / Banca: Saulo da Luz e Silva / Banca: Ana Fabrícia Braga Magalhães / Resumo: Este estudo propôs avaliar uma outra forma de detecção de variações estruturais no genoma bovino, as variações no número de cópias CNVs, e sua associação com características de qualidade de carne. O capitulo 1 aborda considerações gerais e revisão de literatura das características de qualidade da carne e de sua inclusão em estudos genômicos, bem como a utilização das variações do número de cópias em estudos de associação com características de interesse econômico para a agropecuária brasileira. O capítulo 2 teve como objetivo explorar diferentes softwares para avaliar diferenças na detecção de CNVs em uma população de bovinos da raça Nelore. A diferença metodológica entre os softwares pode ser a causa da diferença de CNVR detectados em ambos os métodos. Os grupos funcionais enriquecidos significativos foram relacionados a funções de ligações de ATP, bem como estruturas das membranas celulares e resposta do sistema imunológico. A presença de genes do tipo LOC também é bastante interessante, pois estes são genes os quais ainda não foram confirmadas sua função biológica, necessitando de mais estudos para a determinação da mesma. O estudo de diferentes algoritmos na detecção de CNVRs pode divergir dependendo das plataformas e metodologias aplicadas em cada software, como foi observado no presente estudo. A sobreposição dos mesmos nos fornece maior confiabilidade na detecção de regiões de CNVs. O capítulo 3 propôs identificar regiões no genoma de bovinos da raça Nelore que apresen... (Resumo completo, clicar acesso eletrônico abaixo) / Abstract: The aim of this study was to evaluate another way to detect structural variations in bovine genome, the variations in CNVs number of copies and its association with meat quality traits. Chapter 1 addresses general considerations and literature review of meat quality traits and their inclusion in genomic studies, as well as the use of variations in the number of copies in association studies with characteristics of economic interest for Brazilian agribusiness. Chapter 2 aimed to explore different softwares to evaluate differences in the detection of CNVs in a population of Nellore cattle. The difference in methodologies between the softwares can be the cause of the difference in CNVR detected in both methods. Significant enriched functional groups were related to ATP binding functions, as well as cell membrane structures and immune system response. The presence of genes of the LOC type is also very interesting, since the biological function of these genes have not been confirmed yet, requiring further studies to determine the same. The study of different algorithms in CNVRs detection can differ depending on the applied platforms and methodologies in each software, as it was observed in this study. The overlaps can provide more reliability in the detection of CNVs regions. Chapter 3 proposed to identify genome regions of Nellore cattle that presented variations in the number of copies (CNV: copy number variation) and to study the genetic association of CNVs with quantitative an... (Complete abstract click electronic access below) / Doutor

Orientador: Rogério Abdallah Curi / Banca: Luciana Correia de Almeida Regitano / Banca: Mauricio de Alvarenga Mudadu / Banca: Henrique Nunes de Oliveira / Banca: Josineudson Augusto II de Vasconcelos Silva / Resumo: Caracterizada por sua grande versatilidade em várias modalidades equestres, a raça Quarto de Milha se destaca mundialmente, representando 52% de toda população equina no mundo. Dentro da raça há subdivisão em diferentes segmentos de aptidão, provenientes de distintos objetivos de seleção, consideradas linhagens, entre elas corrida e trabalho. Objetivou-se com este estudo avaliar as diferenças morfológicas e genômicas que ocorrem entre as linhagens de corrida e trabalho como resultado da seleção para diferentes objetivos, a identificação de regiões genômicas que tenham sido alteradas pela seleção na formação da linhagem de corrida em relação à linhagem de trabalho e realizar dois estudos, independentes, de associação ampla do genoma (GWAS) para identificar regiões cromossômicas e genes candidatos posicionais associados com características de desempenho na linhagem de corrida e com características morfométricas na raça como um todo. Para as análises foram utilizados 120 animais de corrida e 64 animais de trabalho de ambos os sexos registrados na Associação Brasileira de Criadores de Cavalo Quarto de Milha. As características morfométricas avaliadas foram: peso, altura à cernelha, comprimentos corporal, da canela, da quartela, da garupa, da cabeça, e do pescoço, perímetros torácico, da canela e do casco e a característica de desempenho índice de velocidade. A análise das diferenças morfológicas foi realizada por meio do procedimento GLM do programa SAS utilizando-se modelo que incluiu os efeitos de sexo e linhagem e a covariável idade à mensuração. Para a análise das diferenças genômicas, 54.602 SNPs foram genotipados utilizando-se o painel de marcadores Illumina Equine SNP50 BeadChip. Os resultados obtidos revelaram mudanças significativas entre as linhagens para todas as características morfométricas avaliadas. Animais de corrida apresentaram maiores pesos, alturas, comprimentos ... / Abstract: Characterized by its versatility in several sporting activities, the Quarter Horse breed excels globally, representing 52% of the entire equine population in the world. The Quarter Horse breed is subdivided into different lines according to skills resulting from distinct selection objectives, including cutting and racing horses. The aims of the present study were to investigate the morphological and genomic difference, between cutting and racing lines as a result of selection for different objectives; identification of the genomic regions divergently selected in racing line in relation to cutting line and to perform two genome-wide association studies, independently, to identify chromosomal regions and positional candidate genes associated with performance trait in the racing line and morphometric traits in the breed as a whole. To perform the analyses 120 racing animals and 64 cutting animals of both sexes and registered at the Brazilian Association of Quarter Horse Breeders were used. The evaluated morphometric traits were: weight; height at withers; length of the body, shank, pastern, rump, head, and neck; circumference of the chest, shank, and hoof and speed index performance trait. Morphological differences analysis was performed using a model that included the fixed effects of sex and line, and age at recording as covariate. The GLM procedure of the SAS program was used for statistical analysis. To perform genomic differences analysis, 54,602 SNPs were genotyped by the Illumina Equine SNP50 BeadChip array. The results showed significant changes in the morphometric traits of the animals. Racing animals were heavier and taller and presented greater body lengths and perimeters than cutting horses. The number of informative SNPs and the SNP density found in the genome of cutting and racing animals suggest that the Equine SNP50 BeadChip can be used for different purposes in the Quarter Horse breed. To identify genomic regions ... / Doutor

Orientador: João Meidanis / Dissertação (mestrado) - Universidade Estadual de Campinas, Instituto de Computação / Made available in DSpace on 2018-09-11T21:07:22Z (GMT). No. of bitstreams: 1 Fortuna_ViniciusJose_M.pdf: 2317520 bytes, checksum: 52e18a1fcb2b671c1296276814c65290 (MD5) Previous issue date: 2005 / Resumo: Uma das principais formas de se medir a distância evolutiva entre espécies é avaliando-se quão transformado um genoma foi em relação a outro. Tais transformações são conhecidas como rearranjos de genoma. Neste trabalho estaremos analisando o rearranjo chamado de transposição, evento que troca de posição dois blocos consecutivos de genes de um mesmo cromossomo. Mais especificamente, buscamos encontrar o número mínimo de transposições que transforma um cromos somo em outro, valor conhecido como distância de transposição. Matematicamente, consideramos os cromossomos como permutações e o problema de se transformar uma permutação em outra pode ser visto como uma ordenação. Em nosso estudo, introduzimos uma operação de remoção de elementos, ferramenta ainda pouco explorada no estudo da distância de transposição, mas que nos possibilitou obter um limite superior para a distância de transposição. Também sugerimos novas formas de se utilizar a remoção de elementos e a análise de subseqüências de permutações. Como forma de se tentar obter novos conhecimentos sobre o problema da distância de transposição, consideramos a variação do problema em que somente transposições de prefixo são permitidas. Zanoni Dias propôs um algoritmo polinomial que transforma qualquer permutação de comprimento n em sua reversa em f3n/41 passos, porém sem uma prova completa. Modificamos esse algoritmo, mantendo o número de passos, e apresentamos uma prova completa da correção do algoritmo modificado. Ainda no problema de distância de transposição de prefixo, analisamos as permutações cujas distâncias se igualam ao limite inferior de distância de pontos de quebra. Tais permutações são fáceis de serem ordenadas por transposições de prefixo em tempo polinomial, pois possuem ordenação ótima única e bem definida. Ao final chegamos à conclusão que as permutações que são fáceis de se ordenar no problema de transposições de prefixo também são fáceis no problema de transposições, o que prova que a variação do problema auxilia no estudo do problema original / Abstract: One of the main ways of measuring the evolution distance among species is to evaluate how large chunks have moved when comparing two genomes. Such changes are know as genome rearrangements. In this work we analyze a rearrangement event called transposition that changes the position of two consecutive blocks of genes in a chromosome. More specifically, we look for the minimum number of transpositions needed to transform a chromosome into another. This value is called the transposition distance. Mathematically, chromosomes are regarded as permutations and changing one genome into another can be seen as a sorting problem. In our study, we introduce an operation of element removal from permutations, which has not been fully explored, but allowed us to find an upper bound for the transposition distance. We also suggest new ways of making use of element removal and the analysis of permutation subsequences. In the hope of obtaining new knowledge about the problem of transposition distance, we considered the variation of the problem where we allow prefix transpositions only. Zanoni Dias developed a polynomial algorithm that changes any permutation into its reverse using f3n/41 steps, but without a proof of its correctness. We have modified this algorithm, keeping the number of steps, and presented a complete proof of the correction of the modified algorithm. Still about the prefix transposition distance, we have analyzed those permutations whose distance equals the breakpoint lower bound. Such permutations are easily sorted by prefix transpositions in polynomial time, since they have a unique and well-defined optimum sorting. Finally, we concluded that the permutations that are easily sorted in the prefix transposition problem are also easily sorted in the transposition problem, which proves that the variation of the problem helps the study of the original problem / Mestrado / Mestre em Ciência da Computação

Made available in DSpace on 2015-03-04T18:57:57Z (GMT). No. of bitstreams: 1 Dissertacao_Nayra.pdf: 2967499 bytes, checksum: a3fbc3f85195a8b2ca04057e9efda7af (MD5) Previous issue date: 2013-06-14 / The study of the diversity and microbial taxonomy is very important to better understand of the micro-organisms ecology. The microbial taxonomy has undergone important changes with the advent of genomic. Until recently the taxonomy exclusively depended of the DNA-DNA hybridization for define species and to describe new species. From the analysis of the genomic signatures (Average Amino Acid Identity (AAI) e Karlin signature) is possible to determine the similarity between microbial genomes. However, the calculations of those signatures requires time. The automation those calculations using a software was the objective of this dissertation. For develop the software was chosen, for its multiplatform characteristics, the Java language, which allows the use of the software in operational systems Windows, Linux and iOS. For develop was used Eclipse VE and batch files for integrate with Blast 2.2.25+ used for comparison of the identity between genomes. The new software performs calculations AAI and Karlin signature for multiple genomes and metagenomes in real time. It s allowed studies about the genus Mycoplasma and the species Phrochlorococcus marinus. This species comprises 10 species according to the analysis of Karlin and AAI and indicates the need for taxonomy review. The Mycoplasma genus is polyphyletic and possibly comprises different genera. This study advances the taxonomic knowledge about relevant groups in marine habitats, besides create a useful tool for analyses of genome and metagenome. / O estudo da diversidade e taxonomia microbiana é de suma importância para o melhor entendimento da ecologia de microrganismos. A taxonomia microbiana tem passado por importantes transformações com o advento da genômica. Até recentemente a taxonomia dependia exclusivamente da hibridização de DNA-DNA para determinação de espécies e descrição de espécies novas. A partir da análise de assinaturas genômicas (Identidade Média de Aminoácidos (AAI) e Assinatura de Karlin) é possível agora determinar a similaridade entre genomas microbianos. Porém, o cálculo destas assinaturas também consome tempo. A automatização destes cálculos por meio de um software foi o objetivo desta dissertação de mestrado. Para a construção do software foi escolhida a linguagem Java por sua característica multiplataforma, que permite utilização em sistemas operacionais Windows, Linux e iOS. Para o desenvolvimento foi utilizado o software Eclipse VE e arquivos batch para integração com o Blast versão 2.2.25+ utilizado para comparação de identidade entre genomas. O novo software realiza os cálculos de AAI e assinatura de Karlin para múltiplos genomas e metagenomas em tempo real. O novo software permitiu o estudo da taxonomia do gênero Mycoplasma e da espécie Phrochlorococcus marinus. De acordo com as análises da assinatura e AAI esta espécie compreenderia 10 outras espécies, o que indica a necessidade de revisão taxonômica. Já o gênero Mycoplasma é polifilético e compreende possivelmente diferentes gêneros. O presente estudo avança o conhecimento taxonômico de grupos relevantes no meio marinho, além de criar uma ferramenta útil para análise de genomas e metagenomas.