Orientadores: Carmen Silvia Passos Lima, Maria de Lourdes Setsuko Ayrizono / Tese (doutorado) - Universidade Estadual de Campinas, Faculdade de Ciências Médicas / Made available in DSpace on 2018-08-21T01:40:41Z (GMT). No. of bitstreams: 1 Guimaraes_JoseLuizMiranda_D.pdf: 4302732 bytes, checksum: 2d6604590176d72460b4a606d6ba2dd6 (MD5) Previous issue date: 2012 / Resumo: O desenvolvimento de câncer colorretal (CCR) é resultado de uma complexa interação de variáveis, incluindo elementos externos, como a exposição a agentes ambientais e dietéticos, e fatores internos, de natureza somática ou hereditária. Não está estabelecido se genótipos de polimorfismos de baixa penetrância em genes relacionados com o metabolismo do ácido fólico, como o metilenotetrahidrofolato redutase (MTHFR C677T e MTHFR A1298C), o metionina sintase (MTR A2756G), o metionina sintase redutase (MTRR A66G) e o timidilato sintase (TS 2R3R), estão associados com o risco de ocorrência da doença ou com suas manifestações clínicas. Portanto, o objetivo deste estudo foi verificar se esses polimorfismos gênicos influenciam o risco de ocorrência do adenocarcinoma colorretal esporádico (ACRE) e suas manifestações clínicas e biológicas em pacientes da região sudeste do Brasil. Foram avaliados 113 pacientes com ACRE e 188 controles, considerando os aspectos clínicos como a idade, o sexo, a raça, a localização, o grau de diferenciação do tumor, o estágio e os genótipos de cada gene. Os genótipos dos polimorfismos dos genes MTHFR, MTR, MTRR e TS foram avaliados por meio da reação em cadeia da polimerase (PCR) seguida ou não por digestão enzimática. O significado estatístico das diferenças entre grupos foi calculado por meio do teste da probabilidade exata de Fisher ou qui-quadrado. As determinações dos riscos de ocorrência do ACRE, a que pacientes e controles foram submetidos, foram obtidas por meio das razões das chances (ORs) e calculadas considerando um intervalo de confiança de 95%. Portadores dos genótipos MTRR 66AG+GG, do MTHFR 1298AC+CC+677CT+TT, do MTHFR 677CT+TT+MTR 2756AG+GG, do MTHFR 1298AC+CC + 677CT+TT + MTR 2756AG+GG e MTHFR 1298AC+CC + 677CT+TT + MTRR 66AG+GG apresentaram riscos 1,99, 3,26, 2,22, 10,92 e 14,88 vezes maiores, respectivamente, de desenvolver ACRE do que os outros. Além disso, os indivíduos com o genótipo MTHFR 677CT+TT e os genótipos MTR 2756AG+GG tiveram um risco de 2,12 e 1,42 vezes maior de desenvolver ACRE com idade menor do que 50 anos. Afro-Brasileiros com o genótipo GG do polimorfismo MTRR A66G tiveram risco 1,98 vezes maior de desenvolver ACRE, e indivíduos com o genótipo MTR 2756AG+GG e os genótipos MTHFR 677CT+TT estiveram sob risco 2,11 e risco 1,62 vezes maiores de ocorrência de tumores indiferenciados e avançados, respectivamente, do que os demais. Portadores dos genótipos MTHFR 1298AC+CC e MTHFR 1298AC+CC + MTRR 66AG+GG estiveram sob riscos 1,42 e 3,07 vezes maiores de tumor no reto, respectivamente, enquanto que portadores dos genótipos MTHFR 677CT+TT e MTHFR 677CT+TT + TS 2R3R+3R3R estiveram sob riscos 1,55 e 5,39 vezes maiores de tumor de cólon, respectivamente, do que portadores dos genótipos selvagens. Estes dados sugerem que polimorfismos dos genes MTHFR, MTR, MTRR e TS, que codificam enzimas que participam do metabolismo do ácido fólico, especialmente em combinação, têm papéis consistentes para o risco de desenvolver ACRE em indivíduos da região sudeste do Brasil / Abstract: The development of colorectal cancer (CRC) is the result of a complex interaction of variables, including external factors such as exposure to environmental agents and dietary factors and internal factors, whether somatic or hereditary. Is not been established genotypes with low penetrance polymorphisms in genes related to metabolism of folic acid such as methylenetetrahydrofolate reductase (MTHFR C677T and A1298C), methionine synthase (MTR A2756G), methionine synthase reductase (MTRR A66G) and thymidylate synthase (TS 2R3R), are associated with the risk of the disease or its clinical manifestations. Therefore, the aim of this study was to determine whether these genetic polymorphisms influence the risk of sporadic colorectal adenocarcinoma (SCA) and their clinical and biological manifestations in patients from southeast Brazil. For this, we analyzed 113 patients with SCA and 188 controls, considering the clinical aspects such as age, sex, race, location, stage, degree of tumor differentiation and the genotypes of each gene described above. The genotypes of the polymorphisms of the MTHFR, MTR, MTRR and TS were assessed by polymerase chain reaction (PCR) and enzyme digestion. The statistical significance of differences between groups was calculated using the probability test of Fisher's exact or chi-square. Determination of the risks of SCA, the patients and controls were submitted, was obtained through the odds ratios (ORs) and calculated assuming a range of 95%. Carriers of the MTRR 66AG + GG, the MTHFR 1298AC+CC + 677CT+TT, the MTHFR 677CT+TT + MTR 2756AG+GG, the MTHFR 1298AC+CC + 677CT+TT + MTR 2756AG+GG, and the MTHFR 1298AC+CC + 677CT+TT + MTRR 66AG+GG genotypes had a 1.99, a 3.26, a 2.22, a 10.92 and a 14.88-fold increased risks for SCA than others, respectively. In addition, individuals with the MTHFR 677CT+TT and the MTR 2756AG+GG genotypes had a 2.12 and a 1.42-fold increased risks for SCA diagnosed under 50 years. African-Brazilians with the MTRR 66GG genotype had a 1.98-fold increased risk for SCA, and individuals with the MTR 2756AG+GG and the MTHFR 677CT+TT genotypes were under a 2.11 and a 1.62-fold increased risks for undifferentiated and advanced tumors, respectively, than others. Carriers of the MTHFR 1298AC+CC and the MTHFR 1298AC+CC + MTRR 66AG+GG genotypes had a 1.42 and a 3.07-fold increased risks for rectal tumor, respectively, while carriers of the MTHFR 677CT+TT and the MTHFR 677CT+TT + TS 2R3R+3R3R genotypes had a 1.55 and a 5.39-fold increased risks for colon tumor, respectively, than carriers of the wild genotypes. This data suggest that polymorphisms of genes MTHFR, MTR, MTRR and TS, which encode folate-dependent enzymes, particularly in combination, have consistent roles for SCA risk in southeastern Brazil / Doutorado / Fisiopatologia Cirúrgica / Doutor em Ciências

>Magister Scientiae - MSc / Recent advances in the field of plant genetics and application of molecular technologies has lead to greater understanding of various crop genomes and their organization.The applications of these techniques include molecular markers which have been used to examine DNA variation within crop species. This allows for the creation of further genetic variation for new and favourable traits.Molecular markers or DNA markers are short fragments of DNA that can be used to locate desirable genetic traits in the genome or show specific genetic differences. The Maloideae subfamily includes fruit species such as pear. Pears (Pyrus communis L.) are large edible fruit that are grown in cool climates, native to coastal regions in Africa, Asia and Europe. The external appearance of this fruit plays a vital role on its rate of sale potential. Thus it is important for the appearances of the pear to meet the expectations of the consumer.External factors affecting the appearance of fruit, such as shape and colour, can have a large influence on the consumer’s first impression and opinion of what the fruit may taste like(Jaeger and MacFie, et al., 2001). The South African pear industry is the fourth largest in the fruit industry after apple, citrus and grape, exporting 3.8% to Europe (Ferrandi, et al., 2005).Increase in production and export of the pear is dependant on the variety of cultivars with desired traits. New cultivars, especially ranges of new cultivars, with harvest dates from early to late in the season, can fill gaps in the marketing strategy of exporters and in the local markets (Human, et al., 2005) Therefore, development of molecular markers allows for their possible use in maker-assisted selection and for the construction of a genetic linkage map thus leading to the location of favourable traits and ultimately the improvement of the quality of the pear.In this study high throughput genomic DNA extractions were performed. The Cetyltrimethyl ammonium bromide (CTAB) method was employed as the results proved to be most promising. Furthermore the screenings of molecular markers were conducted in order to obtain DNA variation. Molecular markers were used to locate specific genetic differences.Multiplexing PCR was conducted using fluorescent primers for further screening and results proved to be useful as many variations could be observed.

Orientadores: Maria Imaculada Zucchi, Ricardo Ribeiro Rodrigues / Tese (doutorado) - Universidade Estadual de Campinas, Instituto de Biologia / Made available in DSpace on 2018-08-26T12:21:20Z (GMT). No. of bitstreams: 1 Schwarcz_KaiserDias_D.pdf: 5972760 bytes, checksum: 6089d106796adfd5fab684651bfb298e (MD5) Previous issue date: 2014 / Resumo: A degradação ecológica e o desflorestamento são processos que se iniciaram há muito tempo e cuja história confunde-se com a da agricultura. A Mata Atlântica é a segunda maior floresta tropical em ocorrência e importância na América do Sul, possuindo grande diversidade biológica e altos níveis de endemismo. A ocupação desordenada da Mata Atlântica causou sua redução a 11,26% de sua área original, com distribuição de forma fragmentada pelo território brasileiro. A destruição da Mata Atlântica tem resultado na eliminação de muitas populações e, potencialmente, na erosão da diversidade genética de diversas espécies. Essa combinação de alto endemismo e forte ameaça de extinção, faz com que a Mata Atlântica seja considerada um hotspot para a conservação. Nas últimas décadas a recuperação de ecossistemas degradados recebeu a atenção da comunidade científica, dando origem ao campo do conhecimento chamado Ecologia da Restauração, que se dedica aos estudos teóricos dos princípios, práticas, resultados e conseqüências de projetos de restauração. O estudo e monitoramento de áreas de restauração florestal é essencial para melhorar as técnicas de restauração em ecossistemas tropicais e subtropicais. Para que uma determinada espécie se perpetue em uma área em processo de restauração, é preciso que a mesma desenvolva todo o seu ciclo de vida e que gerem descendentes capazes de se desenvolver a ponto de substituir as árvores mães quando as mesmas entrarem em senescência. Por isso há a necessidade de se estudar a variabilidade genética de populações arbóreas dentro de áreas de floresta restaurada, assim como a ocorrência e efetividade do fluxo gênico entre estas áreas e os fragmentos de seu entorno. Neste trabalho, estudamos a variabilidade genética de Myroxylon peruiferum L. f., em duas diferentes áreas de restauração florestal e em duas áreas de remanescentes naturais de Floresta Estacional Semidecidual. Nossos resultados indicam que as restaurações florestais de Cosmópolis e Iracemápolis conservam diversidades genéticas HE e alélicas semelhantes às de remanescentes naturais. A principal diferença entre áreas naturais e restauradas foi a menor riqueza de alelos endêmicos nestas últimas o que é um efeito de amostragem que favorece a perda de alelos raros. A área de restauração florestal mais antiga em Cosmópolis apresentou uma estruturação genética espacial compatível com a de áreas naturais. O mesmo não ocorreu com a restauração mais recente de Iracemápolis. Observou-se a ocorrência de estruturação genética local nas áreas naturais e na área de restauração mais antiga e indícios de fluxo gênico entre os áreas nativas e restauradas. Um estudo adicional do efeito de amostragem sobre as freqüências alélicas demonstrou o fenômeno de perda de alelos com baixa freqüência em eventos de amostragem. O mesmo trabalho indicou que uma amostra de cerca de 30 indivíduos é capaz de representar adequadamente alelos com freqüências acima de 0,05; sendo este um bom número a se considerar na seleção de matrizes para fornecimento de mudas para restauração florestal / Abstract: Ecological degradation and deforestation are processes that started long ago and whose history is intertwined with that of agriculture. Atlantic Forest is the second largest rainforest in occurrence and importance in South America, having great biological diversity and high levels of endemism. Disordered occupation of Atlantic Forest caused its reduction to 11.26% of the original area, with distribution in forest fragments poorly conected across the Brazilian territory. Destruction of the Atlantic Forest has resulted in the elimination of many populations and potentially the erosion of genetic diversity of several species. This combination of high endemism and strong threat of extinction causes the Atlantic Forest to be considered a hotspot for conservation. In the last decades recovery of degraded ecosystems has received attention from the scientific community giving birth to an new area of knowledge called the Restoration Ecology. The study and monitoring of areas of forest restoration is essential to improve restoration techniques in tropical and subtropical ecosystems. For a given species to perpetuate itself in an area undergoing a restoration process, it needs to develop its whole life cycle and generates progeny capable of developing to the point of replacing mothers trees when they die. Therefore there is a need to study the genetic variability of tree populations within areas of restored forest, as well as the occurrence and effectiveness of gene flow between these areas and surrounding fragments. We studied the genetic variability of Myroxylon peruiferum L. f., in two different areas of forest restoration and in two areas of natural remnants of semideciduous forest. Our results indicates that restorations in Cosmopolis and Iracemápolis conserve genetic and allelic diversity HE similar to that of natural remnants. The main difference between natural and restored areas was the lowest richness of endemic alleles which is the result of a sampling effect that favors the loss of rare alleles. The area of older forest restoration in Cosmopolis presented a spatial genetic structure consistent with natural areas. This did not occur with the newer restoration in Iracemápolis. We observed the occurrence of local genetic structure in natural areas and in the area of older restoration and evidence of gene flow between native and restored areas. An additional study about the effect of sampling size on allele frequencies showed the phenomenon of loss of low frequency alleles in sampling events. The same study found that a sample of about 30 individuals are able to adequately represent alleles with frequencies above 0.05; this is a good number to consider in selecting matrix trees to supply seedlings for forest restoration / Doutorado / Genetica Vegetal e Melhoramento / Doutor em Genetica e Biologia Molecular

Para estudar a epidemiologia e evolução do novo genótipo HRSVB denominado BA, caracterizado pela duplicação de 60 nt na proteína G, analisamos 4274 amostras clínicas coletadas de crianças hospitalizadas no Hospital Universitário/USP e Hospital da Santa Casa de Misericórdia, cidade de São Paulo entre os anos de 2001 e 2009. As amostras foram submetidas a RT-PCR seguido do sequenciamento da região G2 do gene G. A duplicação de 60 nt foi detectada em 104 (28.3%) das 367 amostras analisadas. De 2001 a 2004 a circulação do genótipo BA foi baixa, seguido de 85.4% (2005), 57.6% (2006), sem circulação (2007), 10% (2008) e 75% (2009) do total de amostras sequenciadas. As sequências foram comparadas com outras BA de diversos países do mundo. A análise filogenética preliminar dividiu as amostras brasileiras em 5 grupos (BA-I, BAII, BAIII, BAIV e BAVI), sendo que a maioria das amostras de 2005 a 2009 agruparam juntas na linhagem BA-IV, estabelecendo um grupo temporal e geográfico. / In order to study the epidemiology and evolution of the new genotype of HRSVB named BA characterized with a 60-nt duplication in the G protein we analyzed 4274 clinical samples collected from children hospitalized in University Hospital/USP and Santa Casa de Misericórdia Hospital, in São Paulo city, during 2001 to 2009. The samples were subject to RT-PCR followed by sequencing of the G2 region of the G gene. The 60 nt-duplication were detected in 104 (28.3%) of 367 sequencing samples. From 2001 to 2004 the circulation of the BA genotype was low, followed by 85.4% (2005), 57.6% (2006), no circulation (2007), 10% (2008) and 75% (2009) of total sequencing samples. Sequences were compared with G sequences with the 60 nt-duplication globally sampled. Preliminary phylogenetic analysis divided Brazilian samples into five clusters (BA-I, BAII, BAIII and BAVI and BAIV), and almost all samples from 2005 to 2009 were clustered together in BA-IV lineage, establishing temporal and geographical cluster.

Western redcedar (Thuja plicata Donn ex D. Don; redcedar), an indeterminate conifer in the Cupressaceae family, is vulnerable to maladaptation in the face of climate change. Assisted gene flow is one mitigation strategy and involves human-mediated migration of populations, where the projected climate of the area of deployment matches the source climate of the population. Despite the overall projections of warmer temperatures globally, in British Columbia (B.C.), the risk of seasonal frost events will remain and therefore the potential for cold damage and mortality of redcedar exists if the newly migrated populations cannot withstand these freezing events. Knowledge of redcedar's ability to withstand freezing temperatures (cold hardiness) is therefore crucial. Redcedar, like many Cupressaceae species, produces and accumulates the purple-coloured carotenoid rhodoxanthin during the winter. This was hypothesized to be correlated with cold hardiness. Assessment of variation in overall, fall and spring cold hardiness and associated rhodoxanthin concentrations were done through repeated, seasonal freeze testing of clonal grafts originating from across the range of redcedar, and seedling progeny from a subset of these clones. Cold damage was quantified using electrolyte leakage and rhodoxanthin concentrations were quantified using high performance liquid chromatography. Cold hardiness and rhodoxanthin were individually modelled using univariate and bivariate mixed effect models with clone/family as a random effect. Model outputs were compared to climatic variables associated with clonal origin to test for climatic relationships. This study found genetic variation in cold hardiness of redcedar with weak climatic clines. This indicates that assisted gene flow of redcedar should be done on a case-by-case basis, with no need for a climatic threshold. Overall heritability of cold hardiness was 0.17 ± 0.03. Novel findings included the positive genetic correlation between fall and spring cold hardiness (0.55 ± 0.33); lack of reciprocal or parental effect for overall cold hardiness; and weak climatic relationships between cold hardiness and predominantly temperature, with the strongest correlation between number of frost-free days in January (0.38, p < 0.01) in the location of origin and cold hardiness. All findings related to rhodoxanthin were novel. Rhodoxanthin varied with family/provenance and season with heritabilities of 0.30 ± 0.09 in fall, 0.42 ± 0.09 in winter and 0.28 ± 0.09 in spring. Winter and spring rhodoxanthin concentrations were phenotypically correlated (0.50, p < 0.01) and genetically correlated (0.76 ± 0.14). Surprisingly, rhodoxanthin was not detected in clonal grafts of redcedar in any season. Results also indicate that rhodoxanthin cannot be used to estimate cold hardiness. The absence of rhodoxanthin in the clonal grafts compared to the seedlings suggests that plant age impacts rhodoxanthin accumulation. / Graduate / 2021-12-14

As urban areas expand, agricultural lands become increasingly important habitat for many species. Compared to some types of agricultural land-use, ranchlands provide vast expanses of minimally modified habitat that support many threatened and endangered species. Conservation biologists can promote ecologically sound management approaches by quantifying the effects of agricultural practices on resident species. I examined the effects of pasture management, cattle grazing, and landscape characteristics on both adult and larval treefrogs in a ranchland in south-central Florida. I experimentally determined optimal deployment of artificial treefrog shelters constructed of polyvinylchloride (PVC) pipe to efficiently sample adult treefrogs (Chapter 1). Seventy-two shelters were hung on oak trees (Quercus virginiana) and cabbage palm trees (Sabal palmetto) with smooth trunks or boots (residual palm fronds), at all possible combinations of three heights (2, 3, and 4 m), four compass directions (N, S, E, and W) and two water levels (with or without 10 cm). Shelter residence was completely dominated by the Squirrel Treefrog, Hyla squirella (N = 65). Significantly fewer H. squirella were found in shelters on palms with boots than on smooth palms or oak trees (0.29 ± 0.21 [mean ± 1 SE hereinafter] versus 1.3 ± 0.21 and 1.1 ± 0.21, respectively), and shelters with water had slightly more H. squirella than those without (1.5 ± 0.19 versus 0.88 ± 0.19, respectively). Orientation and height did not affect the number of treefrogs encountered; thus, the optimal protocol is to deploy shelters on either smooth palms or oak trees, with water, at 2 m height for easy sampling, and in random compass orientations. I used this protocol to sample H. squirella in woodlots surrounding twelve wetlands and examined how time, frog stage and sex, and landscape features influenced treefrog survival, recapture and site fidelity (Chapter 2). I deployed 15 shelters/ha of woodlot within a 100 m buffer around each wetland. I sampled shelters three times during the fall breeding season, removed all shelters to force frogs to overwinter in natural refugia, and replaced shelters for the final spring sampling. During sampling periods, I sexed, measured, and individually marked each frog using visual implant elastomer (VIE) tags. I used Program MARK to build linear models that included either gender group (female, male or juvenile) or life history stage (adult, juvenile) and either time (sampling interval 1, 2, or 3) or season (fall, spring). I used the most informative model as a null model to assess effects of landscape covariates on survival and recapture. Females had higher survival than either males or juveniles, for which estimates were similar (0.867 vs 0.741 and 0.783, respectively). Survival did not vary over time, although there was some support for an effect of season, with lower survival during the final over-wintering period than in the fall intervals (relative variable importance: group = 0.730; stage = 0.134; time = 0.200; season = 0.310). Adults had higher recapture rates than juveniles (average recapture 0.214 vs 0.102), and recapture for both stages varied over time, with highest recapture in sampling interval two (relative variable importance: group = 0.262; stage = 0.514; time = 0.513; season = 0.229). Hyla squirella was extremely site loyal; no individuals moved between sampling sites, and 95% of recaptured individuals were in their original shelter. Strong terrestrial site fidelity calls into question the traditional "ponds as patches" metapopulation view of treefrog population dynamics. Area of woodlot within 250 m was the most important landscape variable in explaining additional variation in both survival and recapture. Frogs had higher survival and lower recapture in larger woodlots, indicating that intact, contiguous woodlots are higher quality habitat than more fragmented woodlots. Neither survival nor recapture varied with wetland grazing treatments or between pasture types. Finally, I experimentally assessed the effects of cattle grazing and pasture management on larval H. squirella. I selected four wetlands: two in semi-natural pastures (SN) and two in intensively managed pastures (IM). One wetland in each pasture type was fenced so that it was released from cattle grazing (R). I collected three clutches of H. squirella eggs (Clutches A, B, and C) and reared tadpoles in the laboratory until Gosner stage 25. In each wetland, I deployed a total of 50 tadpoles from each clutch into 105 L pens constructed of plastic laundry baskets and mesh window screening. Clutch significantly affected tadpole survival, with Clutch A having the highest percent survival, followed by Clutch B and finally Clutch C (41.66, 32.11 - 53.95 [mean, 95% confidence limits hereinafter]; 9.00, 6.76 - 11.88; 2.89, 2.02 - 4.01, respectively). Wetland type also affected survival, with SN wetlands supporting significantly higher survival than IM wetlands (SN-R: 53.95, 32.88 - 88.13; SN-G: 18.95, 11.30 - 31.36 vs IM-R: 7.32, 4.13 - 12.49; IM-G: 1.09, 0.29 - 2.39). Genetic variation in survival confirms the potential for H. squirella to adapt to rangeland management, but extremely low survival of some clutches indicates that few clutches may be able to survive in low quality wetlands, such as IM-G wetlands. Higher survival in SN pasture wetlands suggest this is a superior habitat and future management objectives should conserve semi-natural pastures and limit further modification of intensively managed pastures, including removing woodlots near wetlands.

No description available.

No description available.

Primers for 10 polymorphic DNA microsatellite loci were developed and characterized for the endangered oyster mussel Epioblasma capsaeformis from the Clinch River, TN. Microsatellite loci also were amplified for individuals collected from the following additional populations or species: (1) E. capsaeformis from Duck River, TN; (2) E. florentina walkeri from Indian Creek, upper Clinch River, VA; (3) E. florentina walkeri from Big South Fork Cumberland River, TN; and (4) E. torulosa rangiana from Allegheny River, PA. Allelic diversity ranged from 9-20 alleles/locus, and averaged 13.6/locus for all 5 populations investigated. Average expected heterozygosity (HE) per locus ranged from 0.78-0.92, and averaged 0.86. A genetic characterization of extant populations of E. capsaeformis and E. florentina walkeri was conducted to assess taxonomic validity and to resolve conservation issues related to recovery planning. These mussel species exhibit pronounced phenotypic variation, and are difficult to characterize phylogenetically using DNA sequences. Monophyletic lineages, congruent with phenotypic variation among species, were obtained only after extensive analysis of combined mitochondrial (1378 bp of 16S, cytochrome-b, ND1) and nuclear (515 bp of ITS-1) DNA sequences. In contrast, analysis of variation at 10 hyper variable DNA microsatellite loci showed moderate to highly divergent populations based on FST values, which ranged from 0.12-0.39. Quantitative genetic variation was observed in fish host specificity, with transformation success of glochidia of E. capsaeformis significantly greater (p<0.05) on the greenside darter Etheostoma blennioides, and that of E. f. walkeri significantly greater (p<0.05) on the fantail darter E. flabellare. Lengths of glochidia differed significantly (p<0.001) between species, with sizes ranging from 241-272 μm. Underwater photographs of mantle-pads and micro-lures of female mussels documented fixed phenotypic variation between species. The texture and color of the mantle-pad of E. capsaeformis is smooth and bluish-white, while that of E. f. walkeri is pustuled and brown, with tan mottling. Based on extensive molecular, morphological, and life history data, a population of E. capsaeformis from the Duck River, TN is described and proposed as a separate species, and a population of E. f. walkeri from the upper Clinch River, VA is described and proposed as a separate subspecies. Genetic management guidelines were developed to assess taxonomic status, genetic variation of donor-recipient populations targeted for augmentation, and field and laboratory protocols to maximize genetically effective population size, minimize genetic changes in captive-reared progeny, and prevent the release of juvenile mussels into non-native drainages. A pragmatic approach to species recovery is advocated; one that incorporates the principles of conservation genetics into breeding programs, but prioritizes the immediate demographic needs of critically endangered mussel species. / Master of Science

La schizophrénie est une maladie psychiatrique grave qui affecte approximativement 1 % de la population. Il est clairement établi que la maladie possède une composante génétique très importante, mais jusqu’à présent, les études ont été limitées au niveau de l’identification de facteurs génétiques spécifiquement liés à la maladie. Avec l’avènement des nouvelles avancées technologiques dans le domaine du séquençage de l’ADN, il est maintenant possible d’effectuer des études sur un type de variation génétique jusqu’à présent laissé pour compte : les mutations de novo, c.-à-d. les nouvelles mutations non transmises de manière mendélienne par les parents. Ces mutations peuvent avoir deux origines distinctes : une origine germinale au niveau des gamètes chez les parents ou une origine somatique, donc au niveau embryonnaire directement chez l’individu. L’objectif général de la présente recherche est de mieux caractériser les mutations de novo dans la schizophrénie. Comme le rôle de ces variations est peu connu, il sera également nécessaire de les étudier dans un contexte global au niveau de la population humaine. La première partie du projet consiste en une analyse exhaustive des mutations de novo dans la partie codante (exome) de patients atteints de schizophrénie. Nous avons pu constater que non seulement le taux de mutations était plus élevé qu’attendu, mais nous avons également été en mesure de relever un nombre anormalement élevé de mutations non-sens, ce qui suggère un profil pathogénique. Ainsi, nous avons pu fortement suggérer que les mutations de novo sont des actrices importantes dans le mécanisme génétique de la schizophrénie. La deuxième partie du projet porte directement sur les gènes identifiés lors de la première partie. Nous avons séquencé ces gènes dans une plus grande cohorte de cas et de contrôles afin d’établir le profil des variations rares pour ces gènes. Nous avons ainsi conclu que l’ensemble des gènes identifiés par les études de mutations de novo possède un profil pathogénique, ce qui permet d’établir que la plupart de ces gènes ont un rôle réel dans la maladie et ne sont pas des artéfacts expérimentaux. De plus, nous avons pu établir une association directe avec quelques gènes qui montrent un profil aberrant de variations rares. La troisième partie du projet se concentre sur l’effet de l’âge paternel sur le taux de mutations de novo. En effet, pour la schizophrénie, il est démontré que l’âge du père est un facteur de risque important. Ainsi, nous avons tenté de caractériser l’effet de l’âge du père chez des patients en santé. Nous avons observé une grande corrélation entre l’âge du père et le taux de mutations germinales et nous avons ainsi pu répertorier certaines zones avec un grand nombre de mutations de novo, ce qui suggère l’existence de zone chaude pour les mutations. Nos résultats ont été parmi les premiers impliquant directement les mutations de novo dans le mécanisme génétique de la schizophrénie. Ils permettent de jeter un nouveau regard sur les réseaux biologiques à l’origine de la schizophrénie en mettant sous les projecteurs un type de variations génétiques longtemps laissé pour compte. / Schizophrenia is a severe psychiatric disorder that affects roughly 1% of the general population. It has been clearly demonstrated that the disease possesses a strong genetic component, but thus far, studies have had limited success in identifying key schizophrenia genes. With the advent of new DNA sequencing technologies it is now possible to study a type of genetic variation that has been previously looked over: de novo mutations (new mutations not transmitted by parents) The main aim of the present thesis is to better characterize de novo mutations in schizophrenia. As the role of these variations is not very well known, it was also necessary to study them in a global context in the human population. The first part of our project was to do a comprehensive study of de novo mutations found in the coding section (exome) of patients affected with schizophrenia. We found that the mutation rate was higher than expected. We also observed an aberrant number of nonsense mutations, which suggests a pathogenic profile of mutations. Thus, we strongly suggested that de novo mutations are key players in the genetic mechanism of schizophrenia. The second part of the work builds on the genes bearing mutations identified in the exome sequencing analysis. We sequenced these genes in a larger cohort of cases and controls in order to establish the profile of rare variants for these genes. We were able to conclude that the global mutational profile of the genes identified during de novo mutation studies are indeed pathogenic, which confirms that some of those genes are really involved in the disease and are not sequencing artefacts. Additionally, we were also able to identify some genes that had an aberrant rare variation profile. The third part of the work aimed to characterize the paternal age effect on the de novo mutation rate. Indeed, in schizophrenia, it has been shown numerous times that paternal age is a risk factor for the disease. Thus, we have chosen to characterize this effect in a cohort of healthy subjects. We were able to observe a high correlation between paternal age and an elevated germline mutation rate. We were also able to confirm the existence of genomic regions that present an elevated number of de novo mutations, supporting the notion of mutational hotspots. Our results were amongst the first to be published on the scientific area to directly involve de novo mutations in the genetic mechanism of schizophrenia. Those results bring new clues on the biological networks underlying schizophrenia by investigating a genetic variation type long overlooked.