Equine herpesvirus-1 (EHV-1) can cause acute upper respiratory tract disease, abortion, neonatal death and neurological disease in horses. Rapid, accurate and timely diagnosis of EHV-1 infection in horses is important to curtail the spread of this pathogen. It has been reported that the neuropathogenic phenotype of EHV-1 can result from a single non-synonymous nucleotide substitution at position 2254 (A→G2254) in open reading frame 30 (ORF30). This was the basis for the development of an allelic discrimination, real-time PCR assay to distinguish between potential neuropathogenic and non-neuropathogenic EHV-1 strains. However, PCR analysis of a panel of EHV-1 abortion isolates revealed that other point mutations within ORF30 could produce false negative results with this previously described assay. Therefore, one of the objectives of this dissertation project was to develop a more sensitive and specific allelic discrimination real-time PCR assay for the detection of EHV-1. This was achieved by redesigning the primers and probes targeting ORF30. The new assay was ten times more sensitive than the original assay, with a lower detection limit of 10 infectious virus particles. While mutations within EHV-1’s genome can hinder diagnosis, they can also impact the virulence of the virus. Objective two, therefore, was to determine if sequential cell passage of T953 would induce sufficient attenuation of the EHV-1 genome to produce a low virulence phenotype. Two separate groups of 28 BALB/c mice were inoculated with either the parental strain or passage 135 (T953 P135) of EHV-1 strain T953. The animals were observed for fourteen days, euthanized and their tissues analyzed for the presence of EHV-1. At the conclusion of the fourteen day observation period, all of the mice infected with T953 P135 survived and regained their pre-inoculation body condition. Furthermore, there were significant differences in virus titer and viral DNA concentrations between T953 P135 and the parental strain, further confirming the attenuated phenotype of the virus. Taken together, data from this study clearly demonstrates that sequential cell culture passage of the neuropathogenic T953 strain of EHV-1 results in attenuation for young adult BALB/C mice.

This thesis addresses a number of problems in computer vision, image processing, and geometry processing, and presents novel solutions to these problems. The overarching theme of the techniques presented here is a multi-scale approach, leveraging mathematical tools to represent images and surfaces at different scales, and methods that can be adapted from one type of domain (eg., the plane) to another (eg., the sphere). The main problem addressed in this thesis is known as stereo reconstruction: reconstructing the geometry of a scene or object from two or more images of that scene. We develop novel algorithms to do this, which work for both planar and spherical images. By developing a novel way to formulate the notion of disparity for spherical images, we are able effectively adapt our algorithms from planar to spherical images. Our stereo reconstruction algorithm is based on a novel application of distance transforms to multi-scale matching. We use matching information aggregated over multiple scales, and enforce consistency between these scales using distance transforms. We then show how multiple spherical disparity maps can be efficiently and robustly fused using visibility and other geometric constraints. We then show how the reconstructed point clouds can be used to synthesize a realistic sequence of novel views, images from points of view not captured in the input images, in real-time. Along the way to this result, we address some related problems. For example, multi-scale features can be detected in spherical images by convolving those images with a filterbank, generating an overcomplete spherical wavelet representation of the image from which the multiscale features can be extracted. Convolution of spherical images is much more efficient in the spherical harmonic domain than in the spatial domain. Thus, we develop a GPU implementation for fast spherical harmonic transforms and frequency domain convolutions of spherical images. This tool can also be used to detect multi-scale features on geometric surfaces. When we have a point cloud of a surface of a particular class of object, whether generated by stereo reconstruction or by some other modality, we can use statistics and machine learning to more robustly estimate the surface. If we have at our disposal a database of surfaces of a particular type of object, such as the human face, we can compute statistics over this database to constrain the possible shape a new surface of this type can take. We show how a statistical spherical wavelet shape prior can be used to efficiently and robustly reconstruct a face shape from noisy point cloud data, including stereo data.

This work is aimed at the study of the effect of 17α-ethinylestradiol (EE2) on the production and characteristics of ligninolytic enzymes (laccase, Mn-dependent peroxidase and lignin peroxidase) in I. lacteus, T. versicolor, P. chrysosporium and P. ostreatus cultures grown on two types of liquid media. Enzyme activity production in fungal cultures was affected by the composition of culture medium. In the case of P. chrysosporium, the addition of EE2 to the complex- medium cultures led to a MnP activity stimulation and simultaneously LiP production was partially repressed in these cultures. In the mineral MM medium, no effect of EE2 on enzyme production by P. chrysosporium was observed. In EE2 treated MM cultures of P. ostreatus lower MnP activities were found when compared to biotic controls. In the case of T. versicolor cultures, the addition of EE2 to the complex medium caused laccase and LiP stimulation in the cultures. In the MM medium, however, only laccase production was affected by EE2. I. lacteus MnP production was partially repressed by EE2 in MM medium. In contrast to that, significantly higher MnP activities were detected in complex- medium I. lacteus cultures after the treatment with EE2. Further EE2 degradation by the fungal cultures was studied. The highest degradation effeciency was...

The goal of this thesis was to create graphical demo in OpenGL, which would appropriately react on music, which is inseparable part of graphical demos. Work consists of two main parts, first is programming of real-time graphical engine, second is creating graphical demo which would run on graphical engine created in first part. First part focues on programming of OpenGL based application, which would render scene in real-time using techniques from computer graphics. Second part focuses mainly on graphical activity, which includes 3D modelling, texturing, creating animation and connection to music (reaction of environment and animations to changes in music)

Células tronco mesenquimais de tecido adiposo, são uma promissora ferramenta para aplicações clínicas em terapias celular e regenerativa, em vista da facilidade de sua extração e da maior quantidade de células por unidade de massa de tecido quando comparado a outras fontes clássicas de células mesenquimais como medula óssea. O protocolo clássico de extração e purificação dessas células, depende de sua adesão em plástico e xeno-materiais demandando muito tempo para ser utilizado por médicos para auxiliar pacientes em procedimentos de emergência. Estas células são capazes se diferenciar em diversos tipos celulares, o que as torna boas candidatas para terapia celular, embora sua capacidade de transdiferenciação para fenótipos neuronais seja ainda discutida. Neste trabalho demonstramos um novo processo para isolar essas células na base de epitopos específicos expressos (assinatura molecular de superfície) utilizando aptâmeros como ligantes de alta afinidade para estes sitios. Aptâmeros, moléculas de DNA simples fita identificadas a partir de uma biblioteca combinatória de sequencias de DNA simples-fita foram identificados por ciclos reiterativos de seleção in vitro (SELEX) utilizando células tronco do lipoaspirado como alvo. Dois aptâmeros isolados, denominados APT9 e APT11, foram capazes de identificar subpopulações (15,8 e 23,7% respectivamente) dentre as células tronco mesenquimais (classicamente CD29+/CD90+/CD45-) e separá-las usando nano-partículas magnéticas acopladas aos aptâmeros. Além disso, seguindo uma indução para diferenciação neuronal, as células tronco mesenquimais passam a apresentar morfologia neuronal e apresentam expressão e atividade de diversos receptores de neurotransmissores, avaliados por PCR real-time e imageamento de variações da concentração de cálcio intracelular ápos stimulação com vários agonistas de receptores metatrópicos e ionotrópicos. Ao longo da diferenciação, os níveis transcricionais de mRNA de receptores de cininas (B1 e B2), nicotínicos (alfa 7), muscarínicos (M1, M3 e M4), glutamatérgicos (AMPA2 e mGluR2), purinérgicos (P2Y1 e P2Y4) e GABAergicos (GABA-A, subunidade 3) e da óxido nítrico sintase neural aumentaram quando comparados aos níveis das células não diferenciadas, enquanto que os níveis de expressão de outros receptores incluindo purinérgicos P2X1, P3X4, P2X7 e P2Y6 e muscarínico M5 diminuíram. Os níveis de atividade das classes dos receptores estudados, por imageamento de variações da concentração de cálcio intrac, aumentaram para a maioria dos agonistas analisados durante a diferenciação neuronal com exceção para respostas induzidas por glutamato e NMDA. Células diferenciadas expressavam altos níveis de antígenos específicos de neurônios como β3-tubulina, NF-H, NeuN e MAP-2 indicando uma diferenciação em fenótipo neuronal bem sucedida. Desta maneira, esta tese, ao identificar aptâmeros, prove uma inovadora solução para médicos usarem as células tronco mesenquimais dentro de uma sala de cirurgia, através de um método que é capaz de purificar essas células em um tempo clínico viável, com pureza e sem contato com contaminantes. Além disso, nós mostramos aqui que com um protocolo como o proposto para diferenciação neuronal, nós poderíamos induzir essas células para se diferenciar em neurônios, através da ativação de fatores de transcrição específicos, levando às células tronco mesenquimais a serem possivelmente utilizadas em terapias celulares de reparo neuronal. / Adipose mesenchymal stem cells are promising tools for clinical applications in cellular and regeneration therapies, in view of easiness of extraction and higher amount of isolated stem cells per mass of tissue when compared to other classical mesenchymal stem cell sources including bone marrow. The classical protocol to extract and purify these cells, depending on plastic adherence and xeno-materials, is too time consuming to be used by physicians to help patients at emergency procedures. These cells are able to differentiate into various cell types, making them good candidates for cell therapy, however their capability for transdifferentiation into neural phenotypes is yet discussed. Here we show a novel process to isolate these cells using their surface molecular signature and aptamers, ssDNA molecules identified through the SELEX technique, denominated APT9 and APT11 that are able to identify subpopulations (15,8 and 23,7% respectively) within the mesenchymal stem cells (classically CD29+/CD90+/CD45-) and separate them using magnetic nano-particles attached to the aptamers. Moreover, following induction to neural differentiation, mesenchymal cells presents neuronal morphology and present expression and activity of several neurotransmitter receptors, as evaluated by real-time PCR and calcium imaging. During this process, mRNA transcription levels of bradykinin (B1 and B2), cholinergic (alpha 7), muscarinic (M1, M3 and M4), glutamatergic (AMPA2 and mGlu2), purinergic (P2Y1 and P2Y4) and GABAergic (GABA-A, subunit 3) receptors and neuronal nitric oxide synthase were augmented when compared to levels of undifferentiated cells, while the expression levels of other receptors including purinergic P2X1, P2X4, P2X7 and P2Y6 and muscarinic M5 receptors were down-regulated. Activity levels of the studied receptor classes, as studied by calcium imaging, increased for most of the agonists analyzed during the neuronal differentiation with the exception for glutamate- and NMDA-induced receptor responses. Differentiated cells expressed high levels of neuron-specific antigens such as β3-tubulin, NF-H, NeuN and MAP-2, indicating a successful differentiation into neuronal phenotypes. This thesis, by identifying aptamers, provides a novel solution for physicians to use mesenchymal stem cells inside a surgery room, by using a method that are able to purify the cells in a clinical viable time, with purity and no contact with contaminats. Furthermore, we show here that with a protocol as provided for neuronal differentiation, we could induce these cells to differentiate into neurons, by activating specific transcription factors,making mesenchymal stem cells to possibly be used in neuronal repair cell therapies.

Este trabalho apresenta um estudo sobre sistemas operacionais de tempo real (RTOS) utilizados na implementação da infraestrutura de controle digital para sistemas mecatrônicos, mas serve também como referência para outros sistemas que possuam restrições de tempo. Além de ter um caráter experimental, onde foram medidos e analisados dados como o pior tempo de resposta dos sistemas e a latência para tratamento de interrupções, este trabalho de pesquisa ainda contempla a implementação e uso de RTOS em situações práticas, bem como contempla a construção de uma plataforma geral de pesquisa que servirá de base para futuros trabalhos no laboratório de mecatrônica. Os sistemas analisados neste trabalho foram o VxWorks, QNX, Linux, RTAI, Windows XP, Windows CE e \'mü\'C/OS-II. Outro produto gerado durante este trabalho foi um Live CD para auxiliar na implementação e ensino de conceitos e sistemas de tempo real. / This work presents a study about real time operating systems (RTOS) that are utilized as infrastructure to create digital control systems for mechatronics systems, and also for systems that have critical time constraints. Parameters like worst case response time and interrupt latency were measured for each operating system. This research project also covers the implementation and use of RTOS in practical situations. A general research platform for robotics and real time research was also developed and will be used for future works in the Mechatronics Laboratory. The tested systems were VxWorks, QNX, Linux, RTAI, Windows XP, Windows CE and \'mü\'C/OS-II. Another product released during this work was a Live CD to aid the implementation and teaching of real time systems and concepts.

Este trabalho propõe um novo método de proteção de geradores síncronos baseado em inteligência artificial, mais especificamente lógica fuzzy, com o propósito de melhorar a proteção destes componentes vitais aos sistema elétricos de potência. Um sistema elétrico de potência completo composto por geradores, linhas de transmissão e cargas foi simulado através do Real Time Digital Simulator para fornecer dados para realização de testes e validação do algoritmo de proteção inteligente. Adicionalmente, foram desenvolvidos dois esquemas experimentais que proporcionaram a obtenção de um conjunto extensivo de ensaios de faltas internas em dois geradores síncronos reais, com o objetivo de fornecer dados para comprovar a eficácia da proteção proposta. Um sistema integrado de software e hardware cujo objetivo é servir de plataforma para desenvolver e executar em tempo real algoritmos de proteção aplicados foi desenvolvido. Esse sistema integrado foi utilizado nos testes embarcados em tempo-real do algoritmo de proteção desenvolvido. Apresenta-se também um arranjo composto por funções de proteção tradicionais mais utilizadas em campo, as quais serviram como base de comparação de desempenho para o esquema de proteção inteligente desenvolvido. Os resultados apresentados mostram que o método de proteção inteligente proposto é mais sensível para detectar instantaneamente faltas fase-terra em geradores síncronos com aterramento de alta-impedância, além de ser capaz de identificar faltas entre-espiras e entre-caminhos / This work presents the development of a synchronous generator protective technique based on artificial intelligence, specifically fuzzy logic, in order to improve the protection of these vital components of the electric power system. A complete electric power system composed of generators, transmission lines and loads was simulated using the Real Time Digital Simulator to provide data for testing and validating the intelligent protection algorithm. Additionally, an extensive set of internal fault experiments conducted on two actual synchronous generators provided oscillograms to demostrate the proposed protection effectiveness. An integrated hardware and software system whose purpose is to serve as a platform for developing and executing real-time protection algorithms is presented. This integrated system was used for real-time embedded testing of the protection algorithm developed. An arrangement composed of traditional protection functions most used in the field is also presented, this arrangement served as a comparison basis for the intelligent protection scheme performance. The results show that the intelligent protection is more sensitive to detect instantly ground faults in synchronous generators with high-impedance grounding, it is also shown that the proposed scheme is able to identify inter-turns and inter-circuits faults

Este trabalho propõe um novo método de proteção de geradores síncronos baseado em inteligência artificial, mais especificamente lógica fuzzy, com o propósito de melhorar a proteção destes componentes vitais aos sistema elétricos de potência. Um sistema elétrico de potência completo composto por geradores, linhas de transmissão e cargas foi simulado através do Real Time Digital Simulator para fornecer dados para realização de testes e validação do algoritmo de proteção inteligente. Adicionalmente, foram desenvolvidos dois esquemas experimentais que proporcionaram a obtenção de um conjunto extensivo de ensaios de faltas internas em dois geradores síncronos reais, com o objetivo de fornecer dados para comprovar a eficácia da proteção proposta. Um sistema integrado de software e hardware cujo objetivo é servir de plataforma para desenvolver e executar em tempo real algoritmos de proteção aplicados foi desenvolvido. Esse sistema integrado foi utilizado nos testes embarcados em tempo-real do algoritmo de proteção desenvolvido. Apresenta-se também um arranjo composto por funções de proteção tradicionais mais utilizadas em campo, as quais serviram como base de comparação de desempenho para o esquema de proteção inteligente desenvolvido. Os resultados apresentados mostram que o método de proteção inteligente proposto é mais sensível para detectar instantaneamente faltas fase-terra em geradores síncronos com aterramento de alta-impedância, além de ser capaz de identificar faltas entre-espiras e entre-caminhos / This work presents the development of a synchronous generator protective technique based on artificial intelligence, specifically fuzzy logic, in order to improve the protection of these vital components of the electric power system. A complete electric power system composed of generators, transmission lines and loads was simulated using the Real Time Digital Simulator to provide data for testing and validating the intelligent protection algorithm. Additionally, an extensive set of internal fault experiments conducted on two actual synchronous generators provided oscillograms to demostrate the proposed protection effectiveness. An integrated hardware and software system whose purpose is to serve as a platform for developing and executing real-time protection algorithms is presented. This integrated system was used for real-time embedded testing of the protection algorithm developed. An arrangement composed of traditional protection functions most used in the field is also presented, this arrangement served as a comparison basis for the intelligent protection scheme performance. The results show that the intelligent protection is more sensitive to detect instantly ground faults in synchronous generators with high-impedance grounding, it is also shown that the proposed scheme is able to identify inter-turns and inter-circuits faults

Este estudo teve como objetivo detectar a presença do vírus da raiva em diferentes órgãos de morcegos do gênero Artibeus empregando as técnicas moleculares como RT-PCR, hnRT-PCR e Real Time RT-PCR. De aproximadamente 4000 espécimes de morcegos recebidas no Instituto Pasteur para o diagnóstico da raiva, foram selecionados 30 morcegos do gênero Artibeus, com resultados positivos para raiva pelas técnicas tradicionais de IFD e inoculação em células N2A utilizando suspensões feitas a partir do SNC. Para as técnicas moleculares, foram retirados glândulas salivares, bexigas urinárias, rins, pulmões e conteúdos fecais e ainda foram lavadas as calotas cranianas dos espécimes. Os órgãos e conteúdos fecais foram diluídos a 1:10 (P/V) e as bexigas urinárias a 1:20 (P/V). As suspensões foram inoculadas em células N2A para o isolamento viral. Foi realizada a extração do RNA total usando o TRIzol®, foram realizadas a transcrição reversa seguida da PCR e hnRT-PCR com utilização de primers específicos para o gene codificante da proteína N. A partir do produto da transcrição reversa foi realizada a técnica de Real Time RT-PCR, utilizando primers e sonda específicos para variante antigênica 3. Das 30 suspensões de lavado cerebral, 28 (93,33%) resultaram positivos, na inoculação em cultura de células, seguido de glândulas salivares (36,67%), bexigas (16,67%) e conteúdos fecais (3,33%). Os resultados encontrados da sensibilidade nas técnicas de RT-PCR, hnRT-PCR e Real Time RT-PCR foram 56,25%, 82,57% e 82,19% quando avaliadas as 180 amostras analisadas. A comparação das técnicas de hnRT-PCR e Real Time RT-PCR feita pelo teste exato de Fisher quanto a proporção de positivos detectados mostrou que para o lavado cerebral, órgãos e conteúdos fecais a proporção foi igual (P>0,05). Em relação à positividade os resultados encontrados nas técnicas de hnRT-PCR e Real Time RT-PCR foram 100% em lavado cerebral; 90% e 93,33% em glândulas salivares; 83,33% e 90% em bexigas; 80% e 93,33% em rins; 76,67% e 50% em pulmões e 43,33% em ambas as técnicas em conteúdos fecais. Esses resultados sugerem que tanto as técnicas de hnRT-PCR como Real Time RT-PCR podem ser utilizadas como métodos complementares para o diagnóstico da raiva e são sensíveis o bastante para o uso em estudos de patogênese. A técnica de Real Time RT-PCR realizada neste estudo se mostrou eficiente em detectar o RABV em diferentes órgãos e tecidos extraneurais com a vantagem de ser uma técnica mais rápida e sensível. / This study was aimed to detect the presence of rabies virus in different organs of the genus Artibeus bats using molecular techniques such as RT-PCR, hnRT-PCR, and the Real Time RT-PCR. From about 4,000 specimens of bats received for rabies diagnosis at the Pasteur Institute, 30 bats of the genus Artibeus were then selected. The selected bats presented positive results by the traditional DFA and N2A-cells inoculation test using brain tissue suspensions. Samples of salivary glands, urinary bladders, kidneys, lungs, and fecal contents and washings of the skulls were collected for the molecular techniques testing. The organs and the fecal contents were diluted at 1:10 (w/v) and the urinary bladder, at 1:20 (w/v) and these suspensions were inoculated into N2A cells for viral isolation. The extraction of the total RNA was performed by using TRIzol® and followed by the reverse transcription and the PCR and the hnRT-PCR were performed by using specific primers for the gene encoding the protein N. The product obtained by the reverse transcription technique was submitted to the Real Time RT-PCR technique, using primers and probe specific for antigenic variant 3 of the rabies virus. Of the 30 suspensions of the brain washings, 28 (93.33%) were positive in N2A cell culture inoculation, followed by the suspensions of the salivary glands (36.67%), bladders (16.67%) and fecal contents (3.33%). For the 180 samples evaluated, the results of sensitivity found for the RT-PCR, hnRT-PCR and Real Time RT-PCR techniques were 56.25%, 82.57%, and 82.19%, respectively. A comparison of hnRT-PCR and Real Time RT-PCR techniques performed by Fisher\'s exact test showed that the proportion of positives detected by the brain washings, organs and of the fecal content was non-significant (P> 0.05). Regarding the results found in hnRT-PCR and Real Time RT-PCR techniques, 100% positives were in brain washing, 90% and 93.33% in salivary glands, 83.33% and 90% in bladders, 80% and 93.33% in kidneys, 76.67% and 50% in lungs and 43.33% for both techniques on fecal contents. These results suggest that both hnRT-PCR and Real-Time PCR techniques can be used as complementary methods for the diagnosis of rabies and are sensitive enough for use in pathogenesis studies. The Real Time RT-PCR technique performed in this study proved to be faster and more sensitive and effective in detecting RABV in different organs and extra neural tissues of bats.

Este trabalho propõe investigar, desenvolver e validar uma metodologia de projeto para sistemas de diagnóstico para detecção de falhas e anomalias em aplicações de acionamento de motores elétricos, comumente utilizados na indústria de manufatura. A metodologia proposta é baseada na coleta e interpretação de dados de processo de redes PROFINET, perfil PROFIdrive, e ferramentas de aprendizagem de máquinas. Técnicas de extração e redução de atributos são aplicadas nos dados de processo coletados. Estes atributos são utilizados em algoritmos para reconhecimento de padrões, os algoritmos investigados são o k-Nearest Neighbor, Redes Neurais Artificiais, Support Vector Machines, e adicionalmente uma adaptação da metodologia é feita utilizando um algoritmo para detecção de novidades. A avaliação da metodologia considerou quatro cenários para estudos de caso, para falhas comuns em aplicações de máquinas rotativas. Os resultados alcançados demonstram a eficácia da metodologia, que foi capaz de detectar as falhas e anomalias investigadas de maneira satisfatória, similares a trabalhos correlatos, com o diferencial de não exigirem sensores adicionais dedicados na coleta de dados. Desta maneira, o trabalho contribui para área de redes de comunicação industrial, mais especificamente o protocolo PROFINET, diagnósticos de anomalias em máquinas acionadas por motores elétricos, e ferramentas de aprendizagem de máquinas. / This work proposes to investigate, develop and validate a methodology to design diagnostic systems to detect faults and anomalies in motion control applications, commonly used in manufacturing industry. The proposed methodology is based on collection and interpretation of process data from PROFINET networks, PROFIdrive profile, and machine learning tools. Feature extraction and selection techniques are applied to the collected process data. These features are used in algorithms for pattern recognition problems. Investigated algorithms are k-Nearest Neighbor, Artificial Neural Networks, Support Vector Machines and in addition, an adaptation of the methodology is held for novelty detection. Four scenarios were considered as case of studies for methodology evaluation, based on common faults in rotating machine applications. The results proved the methodology effectiveness for diagnostic system design, which were able to detect satisfactorily the investigated faults and anomalies, similar to related work, with the differential of not requiring additional dedicated sensors for data collection. In this way, the work contributes to the area of industrial communication networks, more specifically in PROFINET protocol, diagnostic systems for fault detection in motion control applications, and machine learning tools.