This research defines the morphological characteristics of the immune cells, coelomocytes, of the earthworm, Lumbricus terrestris. Such a definition of coelomocyte morphology can be correlated with immune function which will be of value in the development of a nonmammalian surrogate system to assess the immunotoxic potential of a terrestrial xenobiotics. Earthworms exposed to artificial soil dilutions of two complex xenobiotic mixtures, refuse derived fuel fly ash and a Superfund sire soil demonstrated an enhanced ability to produce hemolytic factors.

Urban forests perform essential ecological functions, and their performance is dependent on soil quality, which is often degraded by human activity. Because earthworms play a key role in soil health, augmenting earthworm populations in urban soils may improve tree performance. However, we know very little about earthworm ecology in highly urbanized soils. The objectives of our study were: (1) to assess earthworm demographics across a range of urban land uses and (2) to evaluate earthworm augmentation techniques for rehabilitating disturbed soils and improving tree growth. 1. We conducted an observational study across three landuse types to assess earthworm abundance and diversity as well as associated soil properties. Earthworm abundance and biomass in were affected by land use type, disturbance time frame, and seasonality. Earthworm abundance and biomass were affected by a suite of complex soil and temporal variables, and soil temperature and moisture seemed to be the most influential properties. 2. We conducted an earthworm inoculation experiment on a compacted cut-fill field soil with a very low existing earthworm population. In 2008, three soil treatments (control, compost, and compost + earthworm) were applied to 2 m2 plots into which two common urban tree species with contrasting soil tolerances were planted (Acer rubrum and Cornus florida). We measured soil physical and chemical properties along with earthworm survival and soil respiration. Earthworm inoculation did not succeed in improving bulk density or increasing soil organic matter, yet it was successful in increasing soil CEC, Fe, and Mn relative to compost only effects. / Master of Science

Submitted by Marcos Anselmo (marcos.anselmo@unipampa.edu.br) on 2016-05-02T13:45:06Z No. of bitstreams: 1 Natasha Pavin.pdf: 478428 bytes, checksum: 248a3efadd02d2695494d5e6ef077e2b (MD5) / Made available in DSpace on 2016-05-02T13:45:07Z (GMT). No. of bitstreams: 1 Natasha Pavin.pdf: 478428 bytes, checksum: 248a3efadd02d2695494d5e6ef077e2b (MD5) Previous issue date: 2016-04-02 / A ciclofosfamida (CP) é um agente antineoplásico e imunossupressor. É uma droga utilizada no tratamento de diversos tipos de tumores, usada para evitar a rejeição em transplante de órgãos e em doenças autoimunes, como artrite reumática e lúpus eritematoso. A CP é um pró-fármaco que sofre biotransformação no fígado pelo citocromo P450 para gerar metabólitos ativos como a mostarda fosfaramida e a acroleína. Seus metabólitos podem ocasionar um quadro de estresse oxidativo causando danos a diversos órgãos, entre eles o sistema reprodutor. A Tribulus terrestris (TT) faz parte da família das Zygophyllaceae, é uma erva rasteira perene com uma distribuição generalizada no Mediterrâneo, climas subtropicais e no deserto de todo o mundo. Na medicina popular tradicional, tem sido utilizada desde a antiguidade como um afrodisíaco, para energizar, vitalizar e melhorar a função sexual e desempenho físico em homens, bem como para tratar infecções urinárias, inflamações, edemas e outras doenças. Apesar de estudos experimentais e clínicos confirmarem parcialmente o fato de alguns efeitos da TT sobre a libido e a produção de esperma estarem envolvidos com o componente majoritário desta planta, a protodioscina, ainda há muito debate a respeito dos possíveis mecanismos de ação, bem como da sua aplicação terapêutica. Considerando o potencial antioxidante e esteroidogênico da TT, este estudo possui como objetivo investigar o potencial protetor do extrato seco da TT contra o dano testicular induzido pela ciclofosfamida em camundongos machos. Os camundongos Swiss machos receberam o extrato seco de TT como pré-tratamento por gavagem, durante 14 dias, na dose de 11 mg/Kg. No 14 ° dia, receberam uma única injeção intraperitoneal de ciclofosfamida na dose de 100 mg/Kg. Após 24 horas, esses animais foram anestesiados e eutanasiados, e o sangue, os testículos e os epidídimos foram retirados para posteriores análises bioquímicas, avaliação espermática e histopatológica. A presença e quantificação de protodioscina no vii extrato de TT foram avaliadas por HPLC (Cromatografia Líquida de Alta Eficiência), onde foi verificada uma concentração de 1,48% no extrato seco. A CP causou um desequilíbrio no sistema reprodutor destes animais, através do aumento das espécies reativas (ER), peroxidação lipídica (TBARS) e carbonilação de proteínas, bem como uma alteração nas enzimas antioxidantes: superóxido dismutase (SOD), catalase (CAT), glutationa peroxidase (GPx), glutationa S-transferase (GST) e glutationa redutase (GR). O pré tratamento com TT foi eficaz em proteger contra esses danos. Além disso, foi verificado uma redução na atividade da enzima 17β-hidroxiesteróide desidrogenase (17β-HSD), corroborando com a redução do nível de testosterona sérica verificado neste trabalho. Ao mesmo tempo, ao analisarmos a histopatologia dos testículos destes animais, verificamos uma moderada desorganização do epitélio espermatogenico, bem como, uma congestão dos vasos sanguíneos intersticiais. Estes achados corroboraram com os resultados encontrados na avaliação seminal dos animais tratados com CP, onde foi possível verificar uma diminuição da motilidade (41,07%), do vigor (23,52%), da integridade de membrana (43,44%) e da velocidade curvo-retilinea (25,43%), e o pré tratamento com TT foi eficaz em proteger contra esses danos. Em conclusão, este trabalho visou demonstrar pela primeira vez o potencial protetor do extrato seco de TT contra os danos ao sistema reprodutor masculino de camundongos expostos ao quimioterápico CP. À melhora dos parâmetros avaliados pode estar relacionada ao potencial antioxidante e esteroidogênico, bem como pela quantidade de protodioscina observada neste extrato. / Cyclophosphamide (CP) is an antineoplastic and immunosuppressive agent. It is a drug used to treat various tumours types, and also to prevent rejection in organ transplantation and autoimmune diseases such as rheumatoid arthritis and lupus erythematosus. The CP is a prodrug which is biotransformed in the liver by cytochrome P450 enzymes to generate active metabolites phosphoramide mustard and acrolein. Its metabolites can cause oxidative stress state, leading to damage in various organs, including the reproductive system. Tribulus terrestris (TT) is part of the family of Zygophyllaceae, it is a perennial creeping herb with a widespread distribution in the Mediterranean, subtropical climates and desert around the world. In traditional folk medicine, it has been used since ancient times as an aphrodisiac, to energize, vitalize and improve sexual function and physical performance in men and to treat urinary infections, inflammation, edema and other diseases. Although experimental and clinical studies partly confirm that some effects of TT on libido and sperm production are involved with the major component of this plant, protodioscin, there is still much debate about the possible mechanisms of action, as well as their therapeutic application. Considering the antioxidant and steroidogenic potential of TT, this study has the objective of investigating the protective potential of TT dry extract against testicular damage induced by cyclophosphamide in male mice. The male Swiss mice received the TT dry extract as pretreatment by gavage for 14 days at a dose of 11 mg kg-1. On day 14, animals received a single intraperitoneally injection of cyclophosphamide at a dose of 100 mg kg-1. After 24 hours, the animals were anesthetized and euthanized, and blood, testes and epididymis were removed for further biochemical analyzes, sperm and histological evaluation. The presence and quantification of protodioscin in TT extract were analyzed by HPLC (High Performance Liquid Chromatography), where a concentration of 1.48% of dry extract was verified. CP caused an imbalance in the reproductive system of these animals by increasing the reactive species (RS), ix lipid peroxidation (TBARS) and protein carbonylation, as well as change in antioxidant enzymes: superoxide dismutase (SOD), catalase (CAT), glutathione peroxidase (GPx), glutathione S-transferase (GST) and glutathione reductase (GR). The pre-treatment with TT was effective in protecting against this damage. Furthermore, a reduction in the activity of 17β-hydroxysteroid dehydrogenase (17β-HSD) was found, confirming the reduction of serum testosterone levels observed in this study. Additionally, when analyzing the histopathology of the animals’ testes, we perceived a moderate disruption of spermatogenesis epithelium, as well as a congestion of interstitial blood vessels. These findings corroborate the results found in seminal evaluation of the animals treated with CP, where we observed a decrease in motility (41.07%), vigor (23.52%), membrane integrity (43,44%) and curved-straight speed (25.43%), and the pre-treatment with TT was effective in protecting against this damage. In conclusion, this study aimed to demonstrate for the first time the protective potential of TT dry extract against damage to the male reproductive system of mice exposed to CP chemotherapy. Improvement of these parameters may be related to antioxidant and steroidogenic potential as well as the amount of protodioscin observed in this extract.

Les Technosols ont des propriétés et une pédogenèse influencées par des matériaux technogéniques qui les constituent. Le modèle expérimental de Technosol construit a été choisi et résulte de l’utilisation délibérée de matériaux technogéniques au sein d’un profil. L’objectif scientifique est d’évaluer la contribution de la faune du sol aux processus impliqués dans le fonctionnement et l’évolution des Technosols. Des expérimentations en laboratoire (cosmes) et en conditions climatiques réelles (lysimètres, parcelles) ont été menées de l’échelle ultrastructurale à celle du pédon. Les résultats indiquent que (i) le Technosol construit est un support de vie de la faune, (ii) le modèle d’ingénieur de l’écosystème : Lumbricus terrestris, par la création de macroporosité et par ses structures biogéniques contribue à sa structuration, son agrégation et à la décomposition de sa matière organique, (iii) lorsque plusieurs groupes d’organismes assurant des fonctions différentes sont associés, des effets sont mesurés majoritairement sur la décomposition de la matière organique. La forte disponibilité des ressources du Technosol autorise la présence de la faune mais masquent en partie ses effets et inhibent les interactions faunistiques qui auront un effet sur les fonctions des sols. Un modèle d’évaluation de l’effet de Lumbricus terrestris sur la macroporosité est proposé. En termes de génie pédologique, un projet de modèle d’aide à la décision résulte des acquis scientifiques. L’inoculation de la faune contribuerait ainsi à initier des mécanismes d’évolution des sols et à les accélérer / Technosols are soils whose properties and pedogenesis are influenced by technical materials which take part in their constitution. The experimental model used here is a constructed Technosol which is deliberately composed of technical materials in a soil profile. The scientific objective is to evaluate the contribution of soil fauna to processes implied in soil functioning and evolution. Experiments in laboratory (cosms) and in climatic conditions (lysimeters, field) are conducted from ultrastructural scale to soil profile scale. Results indicate that (i) constructed Technosol can support soil fauna, (ii) the model of ecosystem engineer: Lumbricus terrestris, by macroporosity and by stables biogenic structures, contribute to the structuration, the aggregation and the decomposition of organic matter of the Technosol, (iii) when several groups of organisms implied in differents soil functions are used, effects are observed on organic matter decomposition. The huge availability of resources in the Technosol allow the presence of soil fauna but its effects and reduce soil fauna interactions which can have effects on soil functioning. A model whose objective is to evaluate effect of Lumbricus terrestris on macroporosity is proposed. In terms of soil engineering, a project of decision support model results of scientific experiences. Soil fauna inoculation in soil engineering could contribute to initiate and accelerate mechanisms of Technosol evolution

Le campagnol terrestre, Arvicola terrestris, occasionne en France, lors de ses pullulations cycliques interannuelles, d’importants dégâts aux prairies de montagne. Un groupe de travail constitué des équipes de recherche de l’Université de Franche-Comté (UFC), de l’INRA (Centre de Biologie et de Gestion des Populations) et d’organismes agricoles (Fédération Régionale de Défense contre les Organismes Nuisibles de Franche-Comté, FREDON), ont privilégié une approche « systémique » dans laquelle les interactions entre les campagnols, leur habitat (paysage, prédateurs) et les pratiques agricoles sont analysées de façon hiérarchisées (spatialement et temporellement). Un des objectifs est de mettre en évidence le plus grand nombre possible de facteurs de contrôle sur lesquels il est possible d’agir, et l'échelle à laquelle ces actions sont pertinentes. Ces études ont permis d’initier une stratégie, expérimentée avec succès, notamment en Franche-Comté et en Auvergne, et qui privilégie la lutte raisonnée. Il subsiste néanmoins des zones d’ombre relatives à la compréhension du cycle, notamment concernant les déterminants de la phase de déclin. Le rôle du cortège de pathogènes (parmi lesquels certains peuvent être transmis à l’homme) reste pour l’instant sujet de débat dans la littérature scientifique. La compréhension des facteurs clés déterminant cette phase devrait permettre aux éleveurs de mieux anticiper les impacts économiques et adopter les stratégies de contrôles des population les plus adéquates. Objectifs de la thèse (1) Tester les hypothèses des pathogènes et de la sénescence pour expliquer le déclin démographique. (2) Rechercher des indicateurs biologiques (diversité des pathogènes et/ou indicateurs immunitaires) qui permettent de prédire les phases de déclin et d’anticiper des mesures agricoles appropriées pour restaurer les prairies. (3) Evaluer le rôle de la transition entre la phase de forte densité et de déclin démographique pour l’émergence de pathogènes circulants par les populations de campagnols et responsables de maladies humaines. Méthodologie générale Des suivis de populations avec des prélèvements réguliers (mensuels) seront réalisés sur plusieurs populations (répliquats) dans la période qui encadre le déclin démographique. Des méthodes fondées sur le séquençage à haut débit (NGS : Next Generation Sequencing) pour l’épidémiologie permettent d’établir des catalogues complets des pathogènes (virus, bactéries, parasites) hébergées par les populations, et d’en mesurer les prévalences. / Context In France, during cyclic population surges, water voles, Arvicola terrestri, cause extensive damage to mountain grassland. A working group consisting of researchers from the University of Franche-Comté (UFC), INRA (Centre de Biologie et de Gestion des Populations) agricultural organizations (Fédération Régionale de Défense contre les Organismes Nuisibles de Franche-Comté, FREDON) are working on systems approach in which interactions between voles, their habitat (landscape, predators) and agricultural practices are analysed hierarchically (in space and time). One of the objectives is to highlight the largest possible number of control factors on which it is possible to act, and the scale at which these actions are relevant. These studies have helped initiate a strategy, successfully tested in Franche-Comté and in Auvergne, which promotes the integrated control of water vole populations. Nevertheless, there are still grey areas in the understanding of the cycle, particularly on the determinants of the decline phase. The role of pathogen communities (some species may even be transmitted to humans) so far remains the subject of debate in the scientific literature. The understanding of the key factors determining this phase should allow farmers to better anticipate economic impacts and to adopt optimal strategies for vole population control Objectives: (1) To test the pathogens and senescence hypotheses in order to explain the population decline. (2) To look for biological indicators (diversity of pathogens and / or immune indicators) that may predict the decline phase in order to anticipate appropriate measures to restore grasslands. (3) To assess the role of the transition between high population density phase and the decline phase for the emergence of pathogens in vole populations that may cause human diseases.General Methodology Population monitoring with regular (monthly) sampling will be made on several populations (replicates) in the period that brackets the vole population declines. Methods based on Next Generation Sequencing (NGS) makes it possible to establish extensive catalogues of pathogens (viruses, bacteria, other parasites) hosted by vole populations and to measure the prevalence.

A abelha Bombus terrestris é um eficiente polinizador, prestando importantes serviços ecossistêmicos na Europa e adjacências, onde é nativa. Suas colônias têm sido criadas em larga escala para polinização agrícola, as quais são comercializadas internacionalmente, inclusive em países fora de sua área de ocorrência nativa. Deliberada ou acidentalmente a espécie tem sido introduzida em ambientes alóctones, em muitos casos tornando-se invasora. Quando invasora, a espécie é um potencial vetor de doenças e um competidor com outras abelhas; vários impactos têm sido relatados em áreas invadidas ao redor do mundo. Na América do Sul, a espécie foi inicialmente introduzida em ambientes naturais no Chile, mas rapidamente a invasão se espalhou; atualmente é encontrada ocupando ambientes naturais na Argentina. A elevada capacidade invasiva da espécie e a alta velocidade de sua expansão, conforme tem sido relatada por pesquisadores, levantou a possibilidade de a espécie alcançar o Brasil por meio de corredores ambientais favoráveis que se conectam com áreas já invadidas, gerando preocupações sobre potenciais impactos aos sistemas naturais e agrícolas. Esta pesquisa empregou uma abordagem interdisciplinar, integrando uma variedade de métodos analíticos oriundos de diferentes áreas da ecologia e os mais avançados recursos de sistemas de informações geográficas para detectar globalmente as áreas susceptíveis à invasão por Bombus terrestris, considerando-se também as mudanças climáticas. Para o sul da América do Sul foram identificados os corredores de invasão que poderão permitir a espécie se espalhar e alcançar Brasil a partir de locais invadidos. Para o Brasil, foram identificados os municípios mais vulneráveis à entrada da espécie, e também aqueles que estão na rota de expansão da invasão Brasil adentro. Para os municípios brasileiros, por sua vez, foram verificadas as culturas agrícolas e as espécies de Bombus nativas que a invasora poderá interagir ao longo da rota de invasão potencial; com estas informações foram apontadas áreas prioritárias, subsidiando o planejamento de monitoramento e ações de controle do processo de invasão, mas também medidas preventivas e mitigadoras de impactos ambientais e econômicos após a invasão, caso venha a ocorrer. / The bee Bombus terrestris is an efficient pollinator, providing important ecosystem services in Europe and surrounding areas, where it is a native species. Their colonies have been reared in large-scale for agricultural pollination, which are internationally traded, including for countries outside its native range. The species has been deliberately or accidentally introduced into non-native environments, becoming invasive in many cases. When invasive, the species is a potential vector of diseases and competes with other bees for resources; many impacts have been reported in invaded countries around the world. In South America, the species was introduced in natural environments of Chile at first, but the invasion was quickly spread; nowadays it is found living in natural environments of Argentina. The high invasiveness of the species and the high speed of its invasive expansion, as has been reported by researchers, raised the possibility of the species reach Brazil through suitable environmental corridors that are connected with areas already invaded, raising the concerns about potential impacts to natural and agricultural systems. An interdisciplinary approach was used in this research, composing a variety of analytical methods from different areas of ecology and applying the most advanced resources of geographic information systems to detect areas susceptible to invasion by Bombus terrestris at global scale, considering also climate change. For the southern South America, the corridors of invasion that could allow the spreading of the species and that potentially reach Brazil were identified. For Brazil, the most susceptible municipalities at the entrance of the species have been identified, and also those that are on the route of expansion inside the country. For Brazilian municipalities, crops and native species of Bombus that the invasive species can interact with along the potential invasion route were verified; with this information, priority areas were identified, supporting the planning of monitoring and control actions of the invasion process, but also preventive and mitigating measures of environmental and economic impacts after the invasion, if it eventually occurs.

Tapirus terrestris (Linnaeus, 1758), o maior frugívoro do Brasil, desempenha um papel importante na dinâmica dos ambientes e é muito suscetível à perda de habitats. Devido à Floresta Estacional Semidecidual ser o ecossistema mais fragmentado e ameaçado do Domínio Florestal Atlântico do Brasil e o Pontal do Paranapanema ser igualmente constituído apenas por remanescentes florestais, as populações de antas (T. terrestris) nessa região, sentindo os efeitos dessas mudanças ambientais, podem estar reduzidas a níveis insustentáveis ao longo do tempo, e conseqüentemente, todo ecossistema pode ser prejudicado. O conhecimento da ecologia alimentar é um dos fatores mais importantes para realização de ações que visem a conservação das espécies e as interações entre plantas e animais são fundamentais para manutenção da dinâmica florestal, assim, os objetivos desse estudo foram conhecer a composição de frutos consumidos pela anta e verificar se houve variação sazonal em sua dieta e sua ação potencial como dispersor de sementes. Diante da paisagem fragmentada da região onde o estudo foi conduzido e da possibilidade dessa alteração afetar a dieta de espécies, objetivou-se verificar se a fragmentação de habitats influencia o consumo de frutos pelas antas. Para isso, foram analisadas 170 amostras fecais e dois conteúdos estomacais, coletados entre maio de 2003 e maio de 2005, no Parque Estadual Morro do Diabo (PEMD), nos fragmentos florestais da Estação Ecológica Mico-Leão-Preto e em outros remanescentes florestais da região do Pontal do Paranapanema, Estado de São Paulo. Depois de coletadas, as fezes foram lavadas, secas em estufa, as sementes foram retiradas das fezes e pesadas separadamente das partes vegetativas. Posteriormente, os frutos foram identificados por pesquisador especialista e foi montado experimento de germinação para verificar a viabilidade das sementes. Sua dieta foi composta por 65,5% de fibras e 34,5% de frutos e sementes. Comparando a composição sazonal da dieta, não houve diferença entre a massa consumida de fibras e frutos (seca: t=0.15, gl=114, p=0,88; chuvosa: t=1,431, gl=56, p=0,16). Com relação à fragmentação florestal, a massa de fibras e frutos consumidos no PEMD não apresentou diferença (t=1.54, gl=129, p=0,13); já nos fragmentos foi composta por maior massa de fibras do que sementes (t=-5.69, gl=41, p<0.001). Foi identificado o consumo de 58 tipos de frutos, pertencentes a 23 famílias vegetais. Dentre estes, 22 itens e oito famílias são registros inéditos. Os frutos mais representativos foram Syagrus romanzoffiana (Cham.) Glassm, Psychotria spp., Bromelia balansae (Mez, 1891), Ilex spp. e Annona cacans (Warm.). Syagrus romanzoffiana, Psychotria spp. e Bromelia balansae foram os frutos mais representativos, apresentando importância considerável nas duas estações, sendo freqüentemente consumidos no PEMD e nos fragmentos florestais. Foi estimada uma riqueza maior de frutos consumidos na estação seca e no PEMD, enquanto a estação chuvosa e os fragmentos apresentaram menores valores (seca=72,77, chuvosa=41,77; PEMD=79,8, fragmentos=33). Não houve predação de sementes durante a mastigação e para certas espécies o processo digestório não inviabilizou as sementes. Estes resultados sugerem que a espécie pode atuar como potencial dispersora ou predadora de sementes. O consumo de maior riqueza de frutos na época seca acompanhou o período com maior número de espécies frutificando na região e conseqüentemente, mais recursos alimentares disponíveis durante essa estação. Como nos remanescentes fragmentados há diminuição das interações ecológicas, levando a redução na abundância de frutos, riqueza e diversidade de espécies, menores massa de sementes e riqueza de frutos nos fragmentos pode ser uma conseqüência do isolamento de habitats. Assim, apesar de T. terrestris consumir uma grande variedade de frutos no Pontal do Paranapanema, sugerindo uma dieta rica e diversa, ao longo do tempo, a população da região pode ser afetada pela fragmentação florestal. / The lowland tapir Tapirus terrestris (Linnaeus, 1758), the largest mammalian frugivore in Brazil, plays an important role in environmental dynamics and is particularly susceptible to habitat loss. The Atlantic Plateau Forest is the most threatened ecosystem of the Brazilian Atlantic Forest Domain. In the Pontal do Paranapanema region (western São Paulo State), this system is entirely composed of remnant forest fragments. Here, the tapir population may decrease to an unsustainable size in long-term and, consequently, the whole ecosystem may become compromised. Food habits are some of the most important data needed for species conservation and to understand the plant-animal interactions necessary to maintain forest dynamics. Thus, the objectives of this study were to investigate in tapirs: fruit species composition in the diet, seasonal variation of consumed items and the animals? potential as seed dispersers. Due to the fragmented landscape in the study region and the possibility that such alteration affects the species? diet, a subsequent objective was to verify if forest fragmentation influences fruit consumption. We analyzed 170 fecal and two stomach-content samples obtained from May 2003 to May 2005. These were collected from Morro do Diabo State Park (MDSP, 36000 ha), forest fragments from the Mico-Leão-Preto Ecological Station and other fragments in the Pontal do Paranapanema region. The tapir diet was composed of 65.5% fibers and leaves and 34.5% fruits and seeds. Tapirs consumed the same amount of fibers and fruits during the wet and dry seasons (t=1.431, p=0.16; t=0.15, p=0.88, respectively). Seed and fiber weight were not significantly different from fruits weight in MDSP (t=1.54, d.f.=129, p=0.13), although in forest patches a lower amount of fruits was consumed (t=-5.69, d.f.=41, p<0.001). Fifty-eight different items from 23 Families of plants were identified ? among these, 22 fruits and eight Families were recorded for the first time in the species? diet. Syagrus romanzoffiana (Cham.) Glassm, Psychotria spp. and Bromelia balansae (Mez, 1891) were the most consumed fruits, important during both seasons and in both MDSP and other forest fragments. The variables associated with the higher richness of fruits consumed were in the dry season and in MDSP (relative to rainy season and other forest fragments): dry=72.77; rainy=41.77; MDSP=79.8, other fragments=33). Observations of seeds found in feces indicated that mastication rarely contributes to seed damage. Germination experiments were undertaken with both whole and damaged seeds originated from feces, several of these remained viable. Suggesting that tapirs can act as seed dispersers. Tapirs consumed the highest richness of fruits during the dry season, likely associated with higher fruit production in the habitat during these months. Our data further suggests that forest fragmentation may be associated with the lower seed mass and species richness found in the tapir diet. Despite the diversity of fruit consumed by tapirs in the Pontal do Paranapanema, the long-term population stability in this region can be affected by habitat fragmentation.

A biomassa vegetal pode ser uma importante fonte de obtenção de diversos produtos a partir da desestruturação de suas frações por um vasto grupo de enzimas. No entanto, a geração de compostos de alto valor agregado a partir da biomassa lignocelulósica requer o desenvolvimento de novos sistemas enzimáticos. Pensando nisso, a prospecção e a caracterização de novas enzimas que estão presentes no secretoma de fungos degradadores da biomassa lignocelulósica tem sido fonte de pesquisa por pesquisadores do mundo todo. O objetivo deste trabalho foi prospectar, clonar e expressar de maneira heteróloga o gene codificante de uma enzima putativa do fungo termofílico T. terrestris em cepas do fungo filamentoso A. nidulans A773 e promover sua caracterização bioquímica e biofísica. O gene da enzima foi amplificado, clonado e inserido no vetor pEXPYR antes de ser inserido no sistema de expressão do A. nidulans. Os transformantes obtidos foram induzidos em meio mínimo de cultivo contendo 3% (m/v) de maltose e 1% (m/v) de glicose em meio estacionário para a produção, seguido da purificação da enzima. Estudos bioquímicos foram realizados para determinar o pH e a temperatura ótima de reação, bem como, a especificidade aos substratos e a determinação dos parâmetros cinéticos. A termoestabilidade da enzima também foi avaliada por estudo de dicroísmo circular (DC). Além disso, foi avaliado o efeito colaborativo entre uma xilanase GH10 e a enzima em estudo na hidrólise do bagaço de cana-de-açúcar pré-tratado. A enzima obtida por expressão heteróloga foi caracterizada como uma xilo-oligossacarídeo oxidase (XylO). Por meio da análise filogenética das sequências de aminoácidos entre a enzima expressa e outras enzimas oxidativas, a XylO foi classificada como pertencente a família das flavoproteínas e subfamília das BBE. A enzima TtXylO demonstrou ter especificidade em oligossacarídeos de C5 apresentando boa atividade enzimática em substratos complexos de xilana. A enzima possui pH ótimo de 5,5 e temperatura ótima de 25 ºC. As análises de DC indicaram temperatura de desnaturação de 62,7 ºC, caracterizando esta enzima como termofílica. Contudo, novos estudos ainda são necessários para avaliar os produtos gerados a partir da oxidação dos diferentes xilo-oligossacarídeos pela XylO e seu potencial uso na indústria. / Plant biomass is an important source for generation of several products obtained from enzymatic cleavage of its fractions by a large group of enzymes. However, the generation of high value compounds from lignocellulosic biomass requires the development of new enzymatic systems. Considering that, prospection and characterization of enzymes present in the biomass-degrading fungi secretome has been a source of study by researchers around the world. The aim of this work was to prospect, clone and heterologously express a putative enzyme encoding gene from the thermophilic fungus T. terrestris in A. nidulans A773 strains and to promote its biochemical and biophysical characterization. The gene was amplified, cloned and inserted into the pEXPYR vector before being inserted into A. nidulans expression system. The transformants were induced by culture in minimal médium containing 3% (w/v) maltose and 1% (w/v) glucose by stationary culture for the production, followed by enzyme purification. Biochemical analyses were performed to determine optimum pH and temperature as well as the substrate specificities and kinetic parameters. The enzyme thermostability was also evaluated by circular dichroism (CD). In addition, the collaborative effect between the enzyme and a GH10 on hydrolys of pre-treated sugarcane bagasse was evaluated. The enzyme obtained by heterologous expression was characterized as a xylooligosaccharide oxidase (XylO). Phylogenetic analysis between amino acid sequences of expressed enzyme and other oxidative enzymes classified XylO as belonging to flavoproteins family and subfamily of BBE. TtXylO has been shown to have specificity on C5 oligosaccharides exhibiting good enzymatic activity on complex xylan substrates. The enzyme has an optimum pH of 5.5 and optimum temperature of 25 ºC. DC analyses showed melting temperature of 62.7 ºC, characterizing this enzyme as thermophilic. In general, further studies are still needed to evaluate the products generated from oxidation of xylooligosaccharides by XylO and their potential use in the industry.

A biomassa vegetal pode ser uma importante fonte de obtenção de diversos produtos a partir da desestruturação de suas frações por um vasto grupo de enzimas. No entanto, a geração de compostos de alto valor agregado a partir da biomassa lignocelulósica requer o desenvolvimento de novos sistemas enzimáticos. Pensando nisso, a prospecção e a caracterização de novas enzimas que estão presentes no secretoma de fungos degradadores da biomassa lignocelulósica tem sido fonte de pesquisa por pesquisadores do mundo todo. O objetivo deste trabalho foi prospectar, clonar e expressar de maneira heteróloga o gene codificante de uma enzima putativa do fungo termofílico T. terrestris em cepas do fungo filamentoso A. nidulans A773 e promover sua caracterização bioquímica e biofísica. O gene da enzima foi amplificado, clonado e inserido no vetor pEXPYR antes de ser inserido no sistema de expressão do A. nidulans. Os transformantes obtidos foram induzidos em meio mínimo de cultivo contendo 3% (m/v) de maltose e 1% (m/v) de glicose em meio estacionário para a produção, seguido da purificação da enzima. Estudos bioquímicos foram realizados para determinar o pH e a temperatura ótima de reação, bem como, a especificidade aos substratos e a determinação dos parâmetros cinéticos. A termoestabilidade da enzima também foi avaliada por estudo de dicroísmo circular (DC). Além disso, foi avaliado o efeito colaborativo entre uma xilanase GH10 e a enzima em estudo na hidrólise do bagaço de cana-de-açúcar pré-tratado. A enzima obtida por expressão heteróloga foi caracterizada como uma xilo-oligossacarídeo oxidase (XylO). Por meio da análise filogenética das sequências de aminoácidos entre a enzima expressa e outras enzimas oxidativas, a XylO foi classificada como pertencente a família das flavoproteínas e subfamília das BBE. A enzima TtXylO demonstrou ter especificidade em oligossacarídeos de C5 apresentando boa atividade enzimática em substratos complexos de xilana. A enzima possui pH ótimo de 5,5 e temperatura ótima de 25 ºC. As análises de DC indicaram temperatura de desnaturação de 62,7 ºC, caracterizando esta enzima como termofílica. Contudo, novos estudos ainda são necessários para avaliar os produtos gerados a partir da oxidação dos diferentes xilo-oligossacarídeos pela XylO e seu potencial uso na indústria. / Plant biomass is an important source for generation of several products obtained from enzymatic cleavage of its fractions by a large group of enzymes. However, the generation of high value compounds from lignocellulosic biomass requires the development of new enzymatic systems. Considering that, prospection and characterization of enzymes present in the biomass-degrading fungi secretome has been a source of study by researchers around the world. The aim of this work was to prospect, clone and heterologously express a putative enzyme encoding gene from the thermophilic fungus T. terrestris in A. nidulans A773 strains and to promote its biochemical and biophysical characterization. The gene was amplified, cloned and inserted into the pEXPYR vector before being inserted into A. nidulans expression system. The transformants were induced by culture in minimal médium containing 3% (w/v) maltose and 1% (w/v) glucose by stationary culture for the production, followed by enzyme purification. Biochemical analyses were performed to determine optimum pH and temperature as well as the substrate specificities and kinetic parameters. The enzyme thermostability was also evaluated by circular dichroism (CD). In addition, the collaborative effect between the enzyme and a GH10 on hydrolys of pre-treated sugarcane bagasse was evaluated. The enzyme obtained by heterologous expression was characterized as a xylooligosaccharide oxidase (XylO). Phylogenetic analysis between amino acid sequences of expressed enzyme and other oxidative enzymes classified XylO as belonging to flavoproteins family and subfamily of BBE. TtXylO has been shown to have specificity on C5 oligosaccharides exhibiting good enzymatic activity on complex xylan substrates. The enzyme has an optimum pH of 5.5 and optimum temperature of 25 ºC. DC analyses showed melting temperature of 62.7 ºC, characterizing this enzyme as thermophilic. In general, further studies are still needed to evaluate the products generated from oxidation of xylooligosaccharides by XylO and their potential use in the industry.

Les applications thérapeutiques de la neurostimulation ultrasonore représentent un terrain de recherche très prometteur, auquel il fait défaut un modèle valide décrivant les biomécanismes sous-jacents. Le premier objectif de ce travail de thèse était de proposer un modèle nerveux propice à une étude mécanistique du phénomène de neurostimulation ultrasonore. L’objectif suivant était de prouver l’intérêt d’exploiter ce modèle pour recueillir des informations concernant les interactions biophysiques ayant lieu entre les ultrasons (US) focalisés et le système nerveux. La majorité des études réalisées a porté sur le système nerveux du ver de terre commun, Lumbricus terrestris. Elles ont consisté d’une part à comparer entre elles les caractéristiques temporelles des réponses nerveuses associées à différentes modalités de stimulation, et d’autre part à évaluer l’influence de chacun des paramètres acoustiques du stimulus ultrasonore sur le taux de succès de neurostimulation (NSR). Dans les deux cas, la méthodologie suivie reposait sur l’administration de différents stimuli aléatoirement alternés. Complémentairement, le rôle joué par la cavitation acoustique a été étudié. La faisabilité de la stimulation du système nerveux du ver de terre, au moyen d’US et dans des conditions in vivo, a été prouvée. Les aires sensorielles et la dynamique de réponses associées aux trois modalités de stimulation ont été caractérisées. Il a été conclu que, dans ce modèle nerveux invertébré, pendant le phénomène de neurostimulation ultrasonore, les structures nerveuses interagissant fonctionnellement avec les US sont les nerfs afférents segmentaux. Les résultats des études paramétriques ont indiqué que le NSR augmente avec l’intensité acoustique, la durée de pulse et la fréquence de répétition des pulses. Il a été proposé que la structure nerveuse visée est sensible à la « force de radiation moyenne » transportée par le stimulus US, indépendamment des paramètres menant à cette valeur / Ultrasound neurostimulation applied to therapy is a promising field of research but still lacks of a validated model explaining the biomechanisms underlying the phenomenon. The first objective of this PhD thesis was to propose a nervous model suited for a mechanistic study of the phenomenon of ultrasound neurostimulation. In a second time, it was intended to practically prove the interest of this model by using it to gain knowledge regarding the biophysical interactions between focused ultrasound and the nervous system. Studies were performed on the nervous system of the anesthetized earthworm, Lumbricus terrestris. They consisted in either comparing the timings of the nervous responses associated with different modalities of stimulation, or evaluating the influence of each acoustic parameter on the neurostimulation success rate (NSR). In both cases, the methodology followed was to administer randomly mixed sequences of different stimuli. The feasibility of the in vivo activation of the anesthetized earthworm’s nervous system was proven. The sensory fields and response dynamics associated with the three modalities of stimulation were characterized. The parametric studies indicated that the NSR increases with pulse amplitude, pulse duration, pulse repetition frequency, but is more weakly influenced by the harmonic content and number of pulses. By applying a causal approach to interpret the results, we concluded that, in this nervous model, during the phenomenon of ultrasound neurostimulation, the structures functionally responding to the ultrasound stimulus are the segmental afferent nerves. We hypothesize that the main interaction with the axonal regions is mediated by ultrasound radiation force, without excluding the involvement of other biomechanisms