The European rabbit, Oryctolagus cuniculus, was introduced to Australia in 1859 and quickly became a significant vertebrate pest species in the country across a wide distribution. In arid and semi-arid environments, rabbit populations exist as metapopulations - undergoing frequent extinction recolonisation cycles. Previous studies identified population genetic structuring at the regional level between arid and semi-arid environments, and habitat heterogeneity was suggested as a possible causal factor. For the most part, rabbit behaviour has been overlooked as a factor that could contribute to explaining population genetic structure in arid and semi-arid environments. This study utilised a combination of genetic sampling techniques and a simulated territorial intrusion approach to observing wild rabbit behaviour in arid and semi-arid environments. The genetic component of the study compared population samples from each region using four polymorphic microsatellite loci. The behavioural component examined variation in the level of territoriality exhibited by three study populations in the arid region towards rabbits of known versus unknown origins (resident vs transgressor (simulating dispersal)). A difference was observed in population genetic structure determined from nuclear markers between arid and semi-arid regions, which supports findings of previous research using mitochondrial DNA data in the same area. Additionally, differences in aggressive response to known vs unknown rabbits were identified in parts of the arid region, which together with the effects of habitat heterogeneity and connectivity may explain the observed differences in population genetic structure. Knowledge of behavioural plasticity and its effect on relative dispersal success and population genetic structure may contribute to improved management and control of feral rabbit populations at the regional level within Australia; and may assist with conservation efforts in the species' natural range in Europe.

Previous evidence has shown that the simple sequences microsatellites and poly-purine/poly-pyrimidine tracts (PPTs) could be both a cause, and an effect, of meiotic recombination. The causal link between simple sequences and recombination has not been much explored, however, probably because other evidence has cast doubt on its generality, though this evidence has never been conclusive. Several questions have remained unanswered in the literature, and I have addressed aspects of three of them in my thesis. First, what is the scale and magnitude of the association between simple sequences and recombination? I found that microsatellites and PPTs are strongly associated with meiotic double-strand break (DSB) hotspots in yeast, and that PPTs are generally more common in human recombination hotspots, particularly in close proximity to hotspot central regions, in which recombination events are markedly more frequent. I also showed that these associations can't be explained by coincidental mutual associations between simple sequences, recombination and other factors previously shown to correlate with both. A second question not conclusively answered in the literature is whether simple sequences, or their high levels of polymorphism, are an effect of recombination. I used three methods to address this question. Firstly, I investigated the distributions of two-copy tandem repeats and short PPTs in relation to yeast DSB hotspots in order to look for evidence of an involvement of recombination in simple sequence formation. I found no significant associations. Secondly, I compared the fraction of simple sequences containing polymorphic sites between human recombination hotspots and coldspots. The third method I used was generalized linear model analysis, with which I investigated the correlation between simple sequence variation and recombination rate, and the influence on the correlation of additional factors with potential relevance including GC-content and gene density. Both the direct comparison and correlation methods showed a very weak and inconsistent effect of recombination on simple sequence polymorphism in the human genome.Whether simple sequences are an important cause of recombination events is a third question that has received relatively little previous attention, and I have explored one aspect of it. Simple sequences of the types I studied have previously been shown to form non-B-DNA structures, which can be recombinagenic in model systems. Using a previously described sodium bisulphite modification assay, I tested for the presence of these structures in sequences amplified from the central regions of hotspots and cloned into supercoiled plasmids. I found significantly higher sensitivity to sodium bisulphite in humans in than in chimpanzees in three out of six genomic regions in which there is a hotspot in humans but none in chimpanzees. In the DNA2 hotspot, this correlated with a clear difference in numbers of molecules showing long contiguous strings of converted cytosines, which are present in previously described intramolecular quadruplex and triplex structures. Two out of the five other hotspots tested show evidence for secondary structure comparable to a known intramolecular triplex, though with similar patterns in humans and chimpanzees. In conclusion, my results clearly motivate further investigation of a functional link between simple sequences and meiotic recombination, including the putative role of non-B-DNA structures.

The honey possum Tarsipes rostratus, a marsupial endemic to South-Western Australia, feeds exclusively upon nectar and pollen. It is one of the smallest marsupials, with adult females (8-12g) significantly larger than adult males (6-9g). Honey possum males have the longest sperm (356µm) recorded for any mammaland the testes represent 4.2% body weight, amongst the largest recorded formammal species. These features suggest that sperm competition is an importantpart of the mating system. This study used a combination of field based studies,DNA analysis and histological examination of the female reproductive tract toinvestigate the life history, multiple paternity and reproduction of the honey possum innatural populations in the Fitzgerald River National Park (FRNP), on the south coastof Western Australia. This study drew upon earlier work on the honey possum in the FRNP in order to describe its life-history. The honey possum is short-lived (1-2 years), and attains sexual maturity whilst still growing. All four teats are occupied after birth, but the litter is reduced to 2 or 3 young during pouch life. The young have a relatively slow rate of growth. Breeding occurs continuously throughout the year, but is affected by the flowering phenologies of its foodplants. The greatest proportion of females with pouch-young occurs in winter; there are fewest pouch-young in autumn, a time of year when there is a dearth of flowers. Honey possums are essentially solitary animals, with no structured social unit, and male and female home ranges overlap. In captivity they are largely tolerant of one another, but larger females are behaviorally dominant to smaller females and to males. The densities and structure of the honey possum populations in the FRNP were analyzed from trapping data collected over 19 years. Population densities fluctuated significantly from season to season throughout the year, with changes in the flowering food resources available. There were also year-to-year differences in the intensity of those fluctuations, and these were significantly associated with rainfall in the previous year, and probably mediated through a lag effect in the flowering of the honey possum’s foodplants. The greatest densities of animals occurred over winter. In years following high rainfall, mean winter densities reached 88 individuals per hectare. The lowest densities occurred in spring, and in years following low rainfall mean spring densities fell to 8 individuals per hectare. Even at these lowest densities, there is still the potential for interaction between males and females. A succession from high to low, then back to high densities was seen during the three years of the present study (2000-2002) and this shadowed a similar succession of changes in rainfall. The proportion of females with pouch-young was significantly affected by the season, and by rainfall in the previous year. Years following low rainfall had a lower proportion of females in a condition to breed. The autumn dip in breeding that occurred in all years was exacerbated following dry years. Of those females that did breed in 2001, a time of low resources, there was no difference in the size of the litter compared to 2000 and 2002, times of higher resource availability. The sex-ratio of pouch young was at parity, but there was a slight bias towards males among both juveniles (56%) and adults (58%). This was probably due to the greater movements shown by males. Sex ratios were not affected by changes in rainfall and density. Male-biased dispersal was detected using genetic data and the movement patterns of males showed that they moved greater distances than females during their normal activity. Analysis of four microsatellite loci revealed extremely high levels of variation, with 28 to 50 alleles per locus and a mean expected heterozygosity of 0.95. These are amongst the highest seen in any microsatellite study of vertebrates. There was multiple paternity in 86% litters, using a minimum number of sires per litter method, and in 95% litters, using an estimated number of sires method based upon the relatedness of litter males. This indicates that multiple mating is frequent in female honey possums and is evidence for sperm competition. The estimated number of sires in a litter was often three or four. In 41% of cases, the number of sires was less than the number of young in the litter, indicating that some males were more successful at siring offspring than others. Nevertheless, no more than two offspring in a litter were known to have been sired by the same male. Despite marked fluctuations in density from high in 2000, to low in 2001, then high again in 2002, the level of multiple paternity remained equally high in all years. Embryonic diapause and female reproduction was investigated in the honey possum. All adult females examined, both with and without pouch-young, were either close to oestrus, had ovulated or were carrying conceptuses. The honey possum has a postpartum oestrus and it was evident that this occurs approximately 2-4 days after birth. Cleavage and formation of the unilaminar blastocyst appears to occur rapidly over approximately 5 days. Embryonic diapause proceeded in a two phase manner similar to other small possum species. The unilaminar blastocyst expanded rapidly at first; and then, from about 18-20 days after birth, the diameter of the blastocyst remained constant at approximately 1.2-1.8mm. No growth or development beyond the unilaminar stage was observed during pouch-life. The first signs of reactivation occurred during lactation, after pouch exit, and expansion of the blastocyst only occurred in one post-lactational female. The development of the corpus luteum appeared different to patterns described for other marsupials, but its formation coincided with the formation of the unilaminar blastocyst. The diameter of the corpus luteum remained constant throughout diapause. The histology of the reproductive tract was generally similar to other marsupials. There were no sperm storage crypts in the female reproductive tract. The length of pouch-life in the honey possum was 55-65 days, and the interval between litters of the same size varied between 65 and 100 days. Embryonic diapause may reduce the time between production of successive litters in the honey possum, but lifetime reproductive potential is reasonably low. Females had up to four litters over the period that they were captured. Thus, each litter represents a substantial proportion (25%) of a female’s lifetime reproductive output. Reproductive amortization occurred, with 61% loss overall, due to overproduction of ova, loss of conceptuses and reduction of the litter during lactation. The behavioural dominance of females suggests that multiple mating is an active strategy, and this presumably allows the genetic quality of their offspring to be maximized. Males that succeed in sperm competition may be of better intrinsic quality. Overproduction of conceptuses by females presents the opportunity for them to select those fertilized by intrinsically viable males or genetically compatible males. Sexually active males are present all year round. Females were not synchronous in their sexual receptivity, and this would lead to a skewed operational sex ratio, with more reproductive males than oestrous females. Since adult males are significantly smaller than adult females and possess no ornaments or armaments, it is unlikely that males overtly fight for access to females. Rather, males appear to monitor the reproductive status of females through smell, and probably compete in their ability to locate oestrous females. The risk and intensity of sperm competition is high, sexual selection for a large investment in spermatogenesis is evident and competition after copulation is probably an important factor in the mating system. It is likely that males, as well as females mate multiply, and the mating system is promiscuous.

Diss. (sammanfattning) Linköping : Linköpings universitet, 2008. / Härtill 5 uppsatser.

Diss. (sammanfattning) Stockholm : Karol. inst., 2003. / Härtill 5 uppsatser.

Made available in DSpace on 2014-06-11T19:23:30Z (GMT). No. of bitstreams: 0 Previous issue date: 2010-06-17Bitstream added on 2014-06-13T19:50:20Z : No. of bitstreams: 1 mori_nt_me_botfca.pdf: 988090 bytes, checksum: 61ea693b25f67c44cb9358042b0c2f9c (MD5) / Coordenação de Aperfeiçoamento de Pessoal de Nível Superior (CAPES) / Handroanthus heptaphyllus (Vell.) Mattos, sinonímia Tabebuia heptaphylla ( Vell.) Toledo, popularmente conhecida por ipê-roxo, é uma espécie pertencente a família Bignoneaceae, muito apreciada pela sua beleza, madeira de excelente qualidade, além de algumas espécies dessa família possuírem substâncias as quais são usadas como produtos medicinais. A espécie é polinizada por abelhas, pássaros e outros visitantes que podem se alimentar das flores e dos frutos. Atualmente é utilizada em programas de reflorestamento de áreas degradadas, paisagismo e restauração. Ocorre em grande parte do Brasil, desde o Estado da Bahia até o Rio Grande do Sul, Minas Gerais, Mato Grosso do Sul, Santa Catarina e São Paulo, compreendendo as latitudes de 13ºS (BA) a 30ºS (RS). O trabalho teve como objetivos estudar a variabilidade genética entre e dentro das subpopulações de H. heptaphyllus, por meio de marcadores microssatélites e conhecer sobre o seu sistema reprodutivo. Para tanto, foram colhidas sementes de 30 árvores, na região de Botucatu, S.P., sendo grande parte na Fazenda Experimental Lageado pertencente a UNESP - Campus de Botucatu. As sementes foram semeadas no viveiro da UNESP e as folhas das mudas produzidas foram coletadas para a extração de DNA e posteriormente analisadas em géis de poliacrilamida. No total, foram estudados oito locos microssatélites polimórficos, que apresentaram desde seis alelos por loco (loco TAU22) a 14 alelos (locos TAU12, TAU30 e TAU31). A média de heterozigosidade esperada ( e H ˆ ) para as seis subpopulações foi de 0,732, sendo que a heterozigosidade observada ( o H ˆ ) foi de 0,618. Os índices médios de fixação variaram de -0,082 (subpopulação 4) a 0,255 (subpopulação 3), com média de 0,152. Os resultados das subpopulações estudadas mostraram os índices de fixação em níveis aceitáveis, com média de 15,2%, no entanto... / Handroanthus heptaphyllus (Vell.) Mattos, sinonímia Tabebuia heptaphylla (Velloso) Toledo, known as ipê-roxo, belongs to the Family Bignoniaceae. It is a very important Brazilian forest tree species because of its beautiful flowers, excellent wood quality, and medicinal properties. Its flowers are usually visited by animals, like bees, birds, bats, etc, for feeding and for pollination purposes. The species has also been used in programs of reforestation of degraded areas, landscaping, and restoration. The ipê-roxo is widespread throughout Brazil, from Bahia State to Rio Grande do Sul, Minas Gerais, Mato Grosso do Sul, Santa Catarina, and São Paulo States, from 13ºS (BA) to 30ºS (RS) latitudes. The research has as objectives to study the genetic diversity within and between subpopulations of H. heptaphyllus by microsatellite molecular markers and to understand its mating system. We collected seeds of 30 trees, through the Botucatu region, Brazil, mostly from the Lageado Experimental Station, São Paulo State University (UNESP) – Botucatu. The seeds were sown in a nursery and the leaves were collected, to extract the DNA, and analyzed through polyacrilamide gels. In a total of eight polymorphic microsatellite loci were analyzed that varied from six alleles (TAU22 locus) to 14 alleles (TAU12, TAU30, and TAU31 loci). The expected heterozygosity mean ( e H ˆ ) for the six subpopulations was 0.7318, the observed heterozygosity mean ( o H ˆ ) was 0.6183, and the average of fixation index (f) between pairs of the six population varied from -0,082 (subpopulation 4) to 0.255 (subpopulation 3), with an average of 0.152. The results of the studied subpopulations have shown acceptable levels of fixation index, presenting an average of 15.2%, therefore, the subpopulation 4 has shown a higher amount of heterozygous than expected. The total genetic diversity ( IT fˆ ) for the six subpopulations... (Complete abstract click electronic access below)

Resumo: Este estudo investigou os efeitos do corte seletivo sobre a diversidade genética, sistema de reprodução e dispersão de pólen em uma população de Hymenaea courbaril L., situado na Floresta Nacional do Tapajós, Estado do Pará, utilizando nove locos microssatélites. A espécie é uma das mais importantes na Amazônia Brasileira devido o alto valor comercial de sua madeira. Antes do corte seletivo, todas as árvores adultas foram mapeadas, amostradas e genotipadas e 367 sementes foram coletadas de 20 árvores matrizes. Após o corte seletivo de 61% das arvores com DAP>81 cm, foram amostradas e genotipadas 250 sementes de 14 árvores reprodutivas. A análise da herança, ligação e desequilíbrio de ligação mostraram que os nove locos utilizados segregam conforme as leis mendelianas, não estão ligados e estão em equilíbrio gamético.As plantas não reprodutivas tinham maior número de alelos (122 alelos) do que as plantas reprodutivas (103) e as progênies (84). Destes alelos, 29 alelos eram exclusivos as plantas não reprodutivas, nove as plantas reprodutivas e 14 as progênies. O número médio de alelos por locos e a heterozigosidade esperada foram significativamente menores nas progênies antes ( k =16,7; Ho =0,710) e após a exploração madeireira ( k =9,33; Ho =0,555). O índice de fixação foi positivo e significativamente diferente de zero em todas as amostras ( F ad=0,111 , F juv=0,160 e F prog=0,045 ), sugerindo endogamia. A estimativa da taxa de cruzamento multilocos populacional ( tm ) foi significativamente menor do que a unidade ( tm =0,962) e a diferença entre a taxa de cruzamento multilocos e unilocos ( tm−t s =0,066) sugere que ocorreram cruzamentos entre parentes. A estimativa do número efetivo de árvores doadoras de pólen antes da exploração ( Nep =3,79) foi significativamente menor que após a exploração de ( Nep =7,2). A taxa de imigração ... (Resumo completo, clicar acesso eletrônico abaixo) / Abstract: This study evaluated the effects of selective logging on the genetic diversity, reproductive system and pollen dispersal of a Hymenaea courbaril L. population by using nine microsatellite loci. Because of the characteristics of the wood, this species is one of the most logged in the Amazon resulting in significant reductions in its natural population. The study is located in the Tapajós National Forest, Para State, Brazil, in a 546 ha stand that is part of a governmental initiative for public forest concessions. To determine the effects of logging, both pre- and post-logging populations were analysed. Before logging, all adult trees of H. courbaril were mapped, sampled and genotyped; additionally progeny was analysed based on 367 seeds collected from 20 matrices. After logging took place (61% of the population above 81cm diameter at breast height), 250 seeds from 14 trees were collected and genotyped. Initially, tests for Mendelian inheritance, linkage and gametic disequilibrium were carried out which confirmed no linkage, deviation or disequilibrium for the used loci. The results for genetic diversity showed that the non-reproductive population presented a higher number of alleles (122) than the reproductive population (103) and progenies (84). From all alleles, 29 were exclusive to non-reproductive trees, nine to reproductive and 14 to progeny. The average number of alleles and expected heterozygosity were significantly lower for progenies in both pre- and post-logging situations ( k =16,7; Ho =0,710; k =9,33; Ho =0,555, respectively). The fixation index was positive and significantly different from zero for all sampled populations which suggests endogamy ( F ad=0,111 , F juv=0, 160 e F prog=0,045 ). The multilocus crossing rate was statistically significant ( tm =0,962) and the difference between multilocus and single-locus outcrossing rate was also significant ... (Complete abstract click electronic access below) / Orientador: Alexandre Magno Sebbenn / Coorientador: André Eduardo Biscaia de Lacerda / Banca: Miguel Luiz Menezes Freitas / Banca: Milton Kanashiro / Mestre

Le corégulateur transcriptionnel RIP140 est un facteur ubiquitaire majeur impliqué dans la régulation de nombreux processus physiopathologiques, qui possède la capacité d’être un coactivateur ou un corépresseur des voies de signalisation selon son recrutement sur les gènes cible. Des résultats du laboratoire ont montré que RIP140 est un facteur de bon pronostic de la tumorigenèse intestinale sporadique. Ce travail s’intéresse à l’implication de ce facteur de transcription dans les cancers colorectaux familiaux et, plus particulièrement, en lien avec le syndrome de Lynch (LS). Le syndrome de Lynch est une prédisposition héréditaire aux cancers, majoritairement colorectaux, caractérisés par un défaut du système de réparation des mésappariements de l’ADN (Mismatch Repair, MMR), dû à une première mutation germinale d’un des gènes de ce système. La perte de fonctionnalité MMR est responsable du phénotype d’instabilité microsatellitaire (MSI). Cependant, il existe des formes familiales de cancers colorectaux, avec MSI, où il n’est pas retrouvé d’atteinte germinale ou somatique de l’un des gènes du système MMR. Ce sont les syndromes apparentés au syndrome de Lynch (Lynch Like Syndrome, LLS) dont la prise en charge est identique à celle du LS. L’utilisation de modèles murins et de lignées cellulaires colorectales, présentant des modulations d’expression de RIP140 ont permis de mettre en évidence l’effet positif de ce corégulateur sur la régulation transcriptionnelle de l’expression des gènes du système MMR, MSH2 et MSH6. La validité fonctionnelle de cette régulation a été explorée par des analyses d’instabilité microsatellitaire et de sensibilité à différentes molécules cytotoxiques. Des cohortes de tumeurs ont permis de confirmer la corrélation d’expression entre RIP140 et les gènes MSH2 et MSH6 chez les patients. En outre, la régulation de l’expression par RIP140 d’une polymérase translésionnelle particulière, la polymérase Polκ, a été étudiée. Cette polymérase assure la réplication des séquences microsatellitaires du génome. Nous avons démontré que RIP140 stimule l’expression du gène POLK dans nos modèles cellulaires et que son expression est corrélée à celle de RIP140 au sein des tumeurs colorectales humaines. Enfin, par séquençage de différentes lignées cellulaires, nous avons mis en évidence une mutation de RIP140 qui entraîne un décalage du cadre de lecture et génère une protéine tronquée avec perte de deux domaines répresseurs de la protéine. Un séquençage à très haut débit nous a permis de rechercher cette mutation parmi des échantillons de tumeurs colorectales avec MSI. Cette mutation est retrouvée dans 19% des tumeurs, notamment LLS (16,2%), où elle est associée à une moins bonne survie globale. Elle affecte les propriétés antiprolifératives et transrépressives de RIP140 ainsi que les régulations positives des gènes MSH2, MSH6 et POLK. Le développement d’un outil anticorps spécifique de cette mutation serait extrêmement utile pour suivre l’expression de la forme mutée au sein des tumeurs et des premiers essais ont été réalisés en ce sens. En conclusion de ce travail, RIP140 contrôle l’expression de gènes majeurs impliqués dans le maintien de l’intégrité du génome et une mutation de ce corégulateur transcriptionnel pourrait être responsable de l’instabilité microsatellitaire de certaines tumeurs où des altérations des gènes MMR ne sont pas retrouvées. Des études cliniques sur des cohortes plus conséquentes seront nécessaires pour valider son intérêt en tant que marqueur utilisable dans la prise en charge des patients. / The transcriptional coregulator RIP140 is an ubiquitous cofactor playing a major role in the regulation of many physiopathological processes. It can either act as a coactivator or as a corepressor of signaling pathways depending on its recruitment on target genes. It has been shown that RIP140 is a good prognostic marker in sporadic intestinal tumorigenesis. This work focuses on its role in familial colorectal cancers and particularly in relation to the Lynch syndrome (LS). Lynch syndrome is a hereditary cancer predisposition, mostly colorectal, characterized by a defect in the Mismatch Repair (MMR) system, due to a first germline mutation of one gene of this system. Loss of MMR function induces a microsatellite instability (MSI) phenotype. However, there are some MSI familial colorectal cancers, where neither germinal nor somatic alteration of one MMR gene is found. They are referred to as Lynch like Syndrome (LLS) and their overall management is identical to that of LS. Murine models and colorectal cell lines, harboring modulations of RIP140 expression, allowed us to demonstrate the positive transcriptional regulation of the MMR genes, MSH2 and MSH6 by RIP140. Functional validation of this regulation was explored by microsatellite instability and sensitivity to various cytotoxic drugs analyses. A positive correlation has been confirmed between RIP140 and MSH2 and MSH6 gene expression in a cohort of 396 patients. Moreover, the transcriptional regulation by RIP140 of a specialized translesional DNA polymerase, the Polκ polymerase, has been investigated. Polκ ensures microsatellite sequences replication. We have demonstrated that RIP140 positively stimulates the expression of the POLK gene in our cell models and which appears correlated with that of RIP140 in human colorectal tumors. Finally, by sequencing different cell lines, we found a frameshift mutation of RIP140, generating a truncated protein with loss of the last two repression domains. High-throughput sequencing allowed us to look for this mutation in patient MSI colorectal tumor samples. This mutation was found in 19% of these tumors, especially LLS (16,2%), where it has been associated with lower overall survival. This mutation affects the antiproliferative and transrepressive properties of RIP140, as well as the positive regulation of the MSH2, MSH6 and POLK gene. Development of a specific antibody for this mutation would be extremely useful in following the expression of this mutated form within tumors and first tests have been already carried out. In conclusion, RIP140 controls expression of major genes involved in genome integrity maintenance and a mutation of this transcriptional coregulator could be responsible for microsatellite instability of some tumors where alterations of MMR genes are not found. Clinical studies on larger cohorts will be necessary to validate its interest as a marker usable in patient management.

Submitted by Ana Maria Fiscina Sampaio (fiscina@bahia.fiocruz.br) on 2012-07-30T18:02:27Z No. of bitstreams: 1 Marcelo Magalhaes Silva Estudo de alterações genéticas....pdf: 2168493 bytes, checksum: 48c50f165408a4314aa5ad8de3c72ca0 (MD5) / Made available in DSpace on 2012-07-30T18:02:27Z (GMT). No. of bitstreams: 1 Marcelo Magalhaes Silva Estudo de alterações genéticas....pdf: 2168493 bytes, checksum: 48c50f165408a4314aa5ad8de3c72ca0 (MD5) Previous issue date: 2012 / Fundação Oswaldo Cruz. Centro de Pesquisas Gonçalo Moniz. Salvador, Bahia, Brasil / A leucemia/linfoma de células T do adulto (ATL) é uma severa doença linfoproliferativa de células T CD4+ associada ao HTLV-1. Por apresentar diferentes manifestações clínicas, essa neoplasia pode ser classificada em cinco formas: aguda, crônica, smoldering, linfomatosa e tumoral primária de pele. Embora alguns trabalhos venham estudando o processo oncogênico mediado pelo HTLV-1, diversos fatores responsáveis pelo desenvolvimento da ATL ainda permanecem desconhecidos. Este estudo teve como objetivo a investigação de alterações genéticas em células ATL (mutações pontuais em genes supressores de tumor e alterações microssatélites) e sua associação com a evolução clínica da doença e sobrevida dos pacientes. A presença de mutações pontuais nos supressores de tumor TP53, p15INK4B e p16INK4A foram avaliadas em 31 pacientes com diferentes formas da ATL (16 agudos, dez crônicos e cinco smoldering) por análise de seqüenciamento de DNA. Cinco pacientes (16%) apresentaram mutações pontuais no gene TP53, sendo que quatro dentre os mesmos foram classificados com a forma aguda. A presença de mutações nos genes avaliados foi associada com pior prognóstico em pacientes com a forma aguda. Em um dos pacientes incluídos neste trabalho (forma aguda) foi verificada a presença de alteração no éxon 2 do gene p16INK4A. Mutações pontuais não foram detectadas no gene p15INK4B em nenhum dos pacientes incluídos. Os marcadores D10S190, D10S191, D11S1391 e D18S21 foram utilizados para a análise de alterações microssatélites por metodologia semi-automatizada. Dentre os 25 pacientes ATL avaliados (seis agudos, oito crônicos, dez smoldering e um linfomatoso), sete apresentaram alterações microssatélites. Três desses pacientes apresentaram instabilidade (MSI), três pacientes apresentaram perda de heterozigosidade (LOH) e em um paciente foi verificado ambas as alterações. Na Bahia, mutações pontuais em TP53 foram detectadas principalmente na forma aguda da ATL e parece estar associada com pior prognóstico. Além disso, de acordo com nossos conhecimentos, este é o primeiro estudo a descrever tanto MSI com LOH em pacientes portando formas crônica e smoldering da ATL. / Adult T-cell leukemia/lymphoma (ATL) is a severe CD4+ lymphoproliferative disease associated to human T-cell lymphotropic virus type 1 (HTLV-1). ATL has different clinical manifestations and is classified in five clinical forms: acute, chronic, smoldering, lymphoma and primary cutaneous tumoral. Although the mechanisms of oncogenesis of the HTLV-1 have been investigated, the factors related to ATL development are still unknown. The goal of this study was to investigate genetic alterations in ATL cells (point mutations in tumor suppressor genes and microsatellite alterations) and their association with the clinical evolution of the disease and survival. The presence of point mutations in TP53, p15INK4B e p16INK4A were evaluated in 31 ATL patients (16 acute, ten chronic and five smoldering) by direct sequencing. Five of them (16%) had TP53 point mutations, four of them with the acute form of ATL. The presence of point mutations in this gene was associated to poor prognosis in acute patients. Only one case (acute form) has an alteration in the exon 2 of the p16INK4A gene. No point mutations of the p15INK4B were found in the patients included. The markers D10S190, D10S191, D11S1391 and D18S21 were considered for the microsatellite analysis using a semiautomated technique. From the twenty-five ATL cases included (six acute, eight chronic, ten smoldering and one lymphoma), seven showed microsatellite alteration. Among them, three patients had microsatellite instability (MSI), three had loss of heterozygosity and one patient presented both alterations. In Bahia, point mutations in TP53 were detected mainly in acute form of ATL and were associated to poor prognosis. The presence of MSI and LOH in the smoldering and chronic forms of ATL were demonstrated for the first time in this study.

Rodovias podem fragmentar populações por dois mecanismos, mortalidade e evitamento. Como espécies que evitam rodovias são raramente atropeladas por veículos e, então, não são detectadas em monitoramentos de fauna atropelada, outras abordagens são necessárias para identificar se eles estão sendo afetados. Ctenomys minutus (tuco-tuco) é um roedor subterrâneo que habita campos arenosos nas margens de rodovias e são raramente registrados em monitoramentos de fauna atropelada. Buscamos identificar se as rodovias são uma barreira para o fluxo gênico de tuco-tuco baseado em nove loci de microssatélite. Coletamos amostras de tecido epitelial de indivíduos de quatro populações: duas com a presença (Weber e Amaral) e, como controle, duas com a ausência de rodovia (Maribo I e Maribo II). Mensuramos diversidade genética, diferenciação genética (estatística F) e acessamos estrutura genética (agrupamento bayesiano). Não observamos redução na variabilidade genética e encontramos um baixo nível de isolamento entre Weber e Amaral e um isolamento ainda menor entre Maribo I e Maribo II. O método bayesiano separou os indivíduos em dois grupos, onde Maribo I e Maribo II são um grupo consistente e Weber e Amaral possuem fracas diferenciações. Os resultados nos indicam que um efeito de barreira entre as populações separadas pela rodovia está em processo e que é necessário mais tempo para observarmos de forma mais clara o isolamento. São necessários mais estudos genéticos e comportamentais para certificar este padrão. Sob aspectos práticos, seria adequado monitorar as populações afetadas e, eventualmente, aplicar alguma medida de mitigação na estrada pra proporcionar conectividade. Por fim, a abordagem genética se mostrou muito interessante para avaliar este impacto. / Roads can fragment populations by two mechanisms, mortality and avoidance behavior. Since species that avoid roads are rarely killed by vehicles and thus cannot be detected in roadkill surveys, other approaches are necessary to identify whether they are affected. Ctenomys minutus (tuco-tuco) is a subterranean rodent who inhabits sand fields including at the margins of roads, however is rarely recorded on roadkill surveys. We aimed to identify if roads are a barrier to the gene flow of tuco-tuco based on nine microsatellite loci. We collected tissue samples from individuals of four populations: a pair with the presence (Weber and Amaral) and, as control, a pair with absence of a road (Maribo I and Maribo II). We measured the genetic diversity, the genetic differentiation (F-statistics), and assessed the genetic structure (Bayesian clustering). We observed no reduction in genetic variability and a low isolation level in pairwise comparison of Weber and Amaral, which was even lower between Maribo I and Maribo II. The Bayesian method separated individuals into 2 clusters, where Maribo I and Maribo II are one consistent cluster and Weber and Amaral present weak differentiations. The results indicate that a barrier effect between populations separated by roads is in process. More genetic and behavioral studies are needed to confirm this pattern. Under practical aspects, it would be appropriate to monitor the affected populations and possibly apply some mitigation measure on the road to provide connectivity. Finally, genetic approach proved very interesting to evaluate this impact.