Doctor of Philosophy / Department of Plant Pathology / Anna E. Whitfield / Tomato spotted wilt virus (TSWV) is an economically important plant virus dependent on insects (thrips) for transmission to plant hosts. Like many animal-infecting viruses, TSWV replicates in the cells of its insect vector. The virus is an emergent disease threatening food and fiber crops worldwide. The aim of this work was to develop novel control strategies against TSWV through a better understanding of the virus-vector interaction. Previously, the TSWV GN protein was shown to be the viral attachment protein, a molecule mediating attachment of virus particles to the midgut epithelial cells of vector thrips. The specific goals of my research were to further examine the utility of disrupting the virus-vector interaction for effective virus control by exploiting GN properties, and to track the route of TSWV in thrips using confocal microscopy. To achieve these goals, I expressed soluble and insoluble forms of GN fused to green fluorescent protein (GFP) transiently and transgenically and examined their cellular localization in planta. GN::GFP recombinant protein localized to Golgi stacks throughout the cells as indicated by a punctate pattern or co-localization to a Golgi marker. In contrast, the soluble form of GN, GN-S::GFP, localized to the ER and apparently also to the cytoplasm. Virus acquisition and transmission assays with GN-S::GFP transgenic tomato plants demonstrated that transmission of TSWV by F. occidentalis was reduced by 35 to 100%. These results indicated that transgenic expression of GN-S in tomato plants may have the potential to prevent secondary spread of the virus. Novel features of the morphology of principal (PSGs) and tubular salivary glands (TSGs) of the insect vector F. occidentalis and of their infection with TSWV were described. The virus colonized different cell types and regions within the PSGs with variable intensity and distribution; and accumulated at the lumen of individual cells. The TSGs of F. occidentalis are proposed as a route for TSWV infection into the PSGs. The transgenic plants and the new knowledge of the virus vector interaction are promising tools to control TSWV and a model approach for the control of other vector-borne viruses.

Os relaxantes neuromusculares não despolarizantes são fármacos indispensáveis nos procedimentos cirúrgicos que requerem entubação endotraqueal, visto que diminuem o tônus muscular. Quimicamente são divididos em derivados isoquinolinicos e derivados de esteróides, esses últimos com maior aplicação clinica e comercialização no Brasil. Assim sendo, é importante a validação de métodos analíticos para o controle de qualidade com alternativas confiáveis que garantam sua eficácia e segurança. Os brometos de vecurônio, de pancurônio e de rocurônio são fármacos relaxantes neuromusculares não despolarizantes derivados de esteróides. As propriedades farmacológicas destes fármacos são significativamente diferentes entre si, porém, as propriedades químicas são bastante similares. Na presente pesquisa foram desenvolvidos e validados métodos analíticos de separação (cromatografia líquida de alta eficiência e eletroforese capilar) para cada fármaco. Estes métodos foram aplicados a medicamentos comercializados no Brasil. Para todos os métodos cromatográficos foi utilizada uma coluna amino devido a seu caráter polar. Em função da baixa absorção dos três fármacos na região ultravioleta, os métodos eletroforéticos foram aplicados com detecções indiretas utilizando substâncias cromóforas. Comparando-se as técnicas utilizadas para determinação dos fármacos nos medicamentos isoladamente, não houve diferença significativa com nível de confiança de 95,0%. Nos testes de hipótese aplicados (F-Snedecor e t-Student), não foram observadas diferenças na precisão (variâncias) e no valor encontrado dos fármacos contidos nas amostras estudadas (comparação de duas médias). / The non-depolarizing neuromuscular relaxant drugs are essential in surgical procedures requiring endotracheal intubation, as they decrease muscle tonics. These drugs are chemically divided in isoquinolines derivatives and steroid derivatives, this latter group, with greater clinical application and commercialization in Brazil. Therefore, the study and validation of analytical methods are important for quality control with reliable alternatives to ensure their effectiveness and safety. The vecuronium, pancuronium and rocuronium bromides are steroid derivatives presenting non-depolarizing neuromuscular relaxant action. Pharmacological properties of these drugs differ significantly, however, its chemical properties are quite similar. In this research, analytical separation methods (high performance liquid chromatography and capillary electrophoresis), for each drug, were developed and fully validated. The methods were applied to pharmaceutical formulations commercialized in Brazil. For all chromatographic methods, an amino column was used, due to its polar characteristics. Due to the low absorption of the three drugs in the ultraviolet region, electrophoretic methods has been applied with indirect detection using chromophoric substances. When comparing both techniques used for quantitative determination of these drugs in pharmaceutical products, no significant difference was observed using a confidence level of 95.0%. By applying hypothesis tests (F-Snedecor and t-Student), it was not observed difference in precision (variance) and in the found amount of drugs contained in the selected samples (comparison of two means).

Les réactions d'hypersensibilité immédiate survenant au cours de l'anesthésie demeurent un sujet de préoccupation majeure pour les anesthésistes. Le médicament le plus fréquemment responsable d'une anaphylaxie sont les curares. Le but de ce travail est de préciser l'épidémiologie actuelle et d'identifier certains facteurs environnementaux susceptibles d'expliquer la fréquence élevée des réactions allergiques aux curares qui peuvent survenir en l'absence de toute exposition préalable. Dans la première partie de la thèse, une enquête nationale décrivant l'épidémiologie des réactions anaphylactiques peranesthésiques entre 2005 et 2007 en France. Elle a confirmé l'intérêt du dosage d'IgE spécifique anti-curare. La survenue de réactions d'hypersensibilité immédiate allergique à l'induction anesthésique lors d'une première exposition aux curares a conduit à évoquer l'existence d'une sensibilisation croisée avec des substances présentes dans l'environnement et possédant un ammonium quaternaire. Différentes hypothèses ont été proposées, notamment une exposition aux produits cosmétiques et/ou désinfectants, une exposition à des antigènes de type phosphorylcholine rencontrés dans les levures, ou certains parasites. Dans la deuxième partie de la thèse, deux populations professionnelles, des apprentis coiffeurs exposés à des agents chimiques contenant des ions ammoniums, et des boulanger-pâtissiers exposés à des allergènes de levures et de parasites, ont été comparées avec la population générale à la recherche de différences concernant la prévalence des anticorps IgE spécifique anti-curare. Au terme de ce travail, l'exposition à des produits cosmétiques chez les coiffeurs apparaît comme un facteur de risque de développer des anticorps IgE spécifique anti-curare, démontrant pour la première fois l'hypothèse du lien entre l'exposition répétée aux produits cosmétiques contenant l'ammonium quaternaire et la sensibilisation croisée aux curares / Immediate hypersensitivity reactions occurring during anesthesia remains a major concern for anesthesiologists. The drugs most frequently responsible for anaphylaxis are neuromuscular blocking agents (NMBAs). The purpose of this study is to determine the current epidemiology and to identify environmental factors that may explain the high frequency of allergic reactions to neuromuscular blocking agents that can occur without any prior exposure. In the first part of the thesis, a national survey describing the epidemiology of anaphylaxis during anesthesia between 2005 and 2007 in France was carried out. The value of testing specific IgE against NMBA was confirmed.The occurrence of immediate allergic hypersensitivity reaction when first exposure to an NMBA during induction led to evoke the existence of cross-sensitization with substances present in environment and having a quaternary ammonium compound. Different hypotheses have been proposed including exposure to cosmetics and / or disinfectants, exposure to antigens encountered in phosphorylcholine yeasts, or parasites, or exposure to a derivative opiate, pholcodine. In the second part of the thesis, two occupational populations, hairdressers apprentice exposed to chemical agents containing ammonium ions, baker and pastry maker exposed to allergens yeast and parasites were compared with the general population to search for differences regarding the prevalence of specific IgE antibody against NMBA. At the end of this work, hairdressers exposed to cosmetics agents appears to be a risk factor for developing specific IgE antibodies against NMBA and we demonstrated for the first time the hypothesis of the ross-sensitization between repeated exposures to cosmetics products containing quaternary ammonium and NMBAs

Este estudo discute uma climatologia de 59 anos (1949-2007) de bloqueios no Hemisfério Sul (SH) usando dados de altura geopotencial em 500-hPa das reanálises do National Center for Environmental Prediction / National Center for Atmospheric Research (NCEP-NCAR). A variabilidade espaço-temporal dos eventos de bloqueio e associações com o El Niño/Oscilação do Sul (ENOS) também foram examinadas. Adicionalmente, os bloqueios foram investigados em dois Modelos de Circulação Geral Acoplados Atmosfera-Oceano de clima (MCGAO) do Intergovernamental Painel for Climate Change (IPCC), o ECHAM5/MPI-OM e o MIROC 3.2. Dois cenários simulados foram analisados: O clima do século XX e o cenário de emissão A1B. Os episódios do ENOS foram identificados usando dois métodos. O primeiro foi o Índice Oceânico Mensal do Niño (ONI) do Climate Prediction Center (CPC-NCEP). O segundo método foi baseado em Funções Empíricas Ortogonais (EOF) e foi aplicado nos MCGAOs. Similarmente, também foi examinado a influencia combinada do ENOS e a Oscilação Antártica (AAO) na ocorrência e características dos bloqueios. O índice diário da AAO foi obtido pelo CPC-NCEP. Os índices convencionais de bloqueios detectam principalmente variações longitudinais. Este trabalho propõe um índice de bloqueio que detecta, além de variações longitudinais também as variações latitudinais dos bloqueios. Cinco setores relevantes de bloqueios foram examinados em detalhes: Indico Sudoeste (SB1), Pacífico Sudoeste (SB2), Pacífico Central (SB3), Pacífico Sudeste (SB4) e Atlântico Sudoeste (SB5). Além disso, foram investigados duas grandes regiões do Pacífico Sul: Pacífico Oeste e Pacífico Leste. Foi encontrado que a escala média típica dos eventos de bloqueio varia entre 1,5 e 2,5 dias. Além disso, a duração dos eventos depende da latitude, com eventos de maior duração observados em latitudes mais altas. Diferenças longitudinais estatisticamente significativas na freqüência do escoamento bloqueado foram observadas entre as fases Quente e Neutra do ENOS desde o outono a primavera. Episódios intensos da fase Quente do ENOS (isto é, moderados a fortes) tendem a modificar o local preferencial de bloqueio, mas não a freqüência. Por outro lado, os episódios fracos da fase Quente do ENOS estiveram associados relativamente com alta freqüência. Os Eventos de bloqueio durante o ENOS+ duram, em média, mais um dia relativamente aos episódios Neutros. Em contraste, a fase Fria do ENOS (ENOS-) caracterizou-se pela redução dos eventos de bloqueio sobre o setor do Pacífico Central, exceto durante os meses do verão austral. Entretanto, nenhuma diferença estatisticamente significativa foi detectada na duração dos eventos. Composições de anomalias de vento em 200-hPa indicam que o enfraquecimento (fortalecimento) do jato polar em torno de 60ºS durante a AAO negativa (positiva) em ambas as fases do ENOS tem uma importância significativa no aumento (redução) dos eventos de bloqueio. Um significativo aumento estatístico dos eventos sobre o setor do Pacífico Sudeste foi observado durante a AAO negativa em ambas as fases do ENOS. Ainda, um aumento (redução) dos eventos foi observado sobre a região do Pacífico Oeste na fase negativa (positiva) da AAO durante o ENOS-. Em contraste, durante o ENOS+ não houve diferenças estatisticamente significativas na distribuição longitudinal dos eventos separado de acordo com as fases opostas da AAO, embora haja um aumento (redução) dos eventos da região do Pacífico Oeste para o Pacífico Leste durante a fase negativa (positiva) da AAO. Os MCGAOs simularam corretamente a amplitude do ciclo anual observado. Também, ambos os MCGAOs simularam melhor a duração e o local preferencial do que freqüência. Nenhum MCGAO simulou adequadamente a freqüência durante a fase Neutra do ENOS. O ECHAM5/MPI-OM (rodada 2) mostra um erro sistemático que levam a uma superestimativa na freqüência de eventos sobre o Pacífico Leste durante as fases Neutra e Fria do ENOS. As diferenças entre as duas versões do MIROC 3.2 indicam que a alta resolução nos modelos melhora o desempenho em simular a freqüência de bloqueios. / This study discusses 59-yr climatology (1949-2007) of Southern Hemisphere (SH) blockings using daily 500-hPa geopotential height data from National Center for Environmental Prediction / National Center for Atmospheric Research (NCEP-NCAR reanalysis. The spatiotemporal variability of blocking events and associations with El Nino/Southern Oscillation (ENSO) are examined. Additionally, blockings were examined in two Intergovernmental Panel for Climate Change (IPCC) Coupled General Circulation Models (CGCM), ECHAM5/MPI-OM and MIROC 3.2. Two sets of simulations were examined: the climate of the 20th century and the A1B emission scenario. ENSO episodes were identified using two methods. The first method was the Monthly Oceanic Niño Index (ONI) from the Climate Prediction Center (CPC-NCEP). The second method was based on Empirical Orthogonal Function (EOF) and was applied to identify ENSO episodes in the CGCMs. Similarly, the combined influence of ENSO and the Antarctic Oscillation (AAO) on the occurrence and characteristics of blockings was also examined. The daily AAO index was obtained from CPC/NCEP. Most conventional blocking indices detect longitudinal variations of blockings. In this study we propose a new blocking index that detects longitudinal and latitudinal variations of blockings. The following relevant sectors of blocking occurrence were identified and examined in detail: Southeast Indian (SB1), Southwest Pacific (SB2), Central Pacific (SB3), Southeast Pacific (SB4) and Southwest Atlantic (SB5) oceans. In addition, we investigated two large regions of South Pacific: West Pacific and East Pacific. We found that the typical timescale of a blocking event is about ~1.5 2.5 days. Nonetheless, the duration of events depends on the latitude, with larger durations observed at higher latitudes. Statistically significant differences in the longitude of blockings are observed between Warm (ENSO+) and Neutral ENSO phases from the Austral fall to spring. Moderate to strong Warm ENSO episodes modulate the preferred locations of blockings but do not play a significant role for variations in their frequency. On the other hand, weak ENOS+ episodes were associated with relatively high frequency of blockings. Blocking events during ENSO+ last on average one more day compared to events that occur during Neutral episodes. In contrast, Cold (ENOS-) ENSO episodes are characterized by a decrease of blockings over the Central Pacific sector, except during the Austral summer months. However, no statistically significant differences are detected in the duration. Composites of 200-hPa zonal wind anomalies indicate that the weakening (strengthening) of the polar jet around 60oS during negative (positive) AAO phases in both ENSO phases plays a major role for the relative increase (decrease) of blocking events. A statistically significant increase of events over Southeast Pacific is observed during negative AAO phases in both ENSO phases. Moreover, an increase (decrease) of events is observed over West Pacific region when negative (positive) AAO phases occur during ENSO-. In contrast, during ENSO+ there is no statistically significant difference in the longitudinal distribution of events separated according to opposite AAO phases, although there is an increase (decrease) in the events from West Pacific region to East Pacific during negative (positive) AAO phase. The CGCMs investigated in this study correctly simulated the amplitude of observed annual cycle of geopotential height in the extratropics. Also, both CGCMs show a better performance in simulating the duration and preferred locations of blockings than their frequency. None of these CGCMs simulated well the frequency during Neutral ENSO phase. The ECHAM5/MPI-OM (run2) shows systematic biases in some regions. For instance, this model overestimates the frequency of blockings over East Pacific region during Cold and Neutral ENSO phases. The differences between the two versions of MIROC 3.2 indicate that the increase in resolution improves the performance of the model in simulating the frequency of blockings.

La caractérisation des réseaux trophiques est nécessaire pour comprendre le fonctionnement des écosystèmes et les mécanismes impliqués dans leur stabilité. Il est parfois difficile de déterminer les régimes alimentaires notamment pour des espèces discrètes et difficiles à observer comme les grands carnivores. Cependant, ces espèces jouent un rôle clé dans les écosystèmes dont elles influencent le fonctionnement et la biodiversité. Ainsi, connaitre le régime alimentaire des grands prédateurs avec précision est essentiel pour établir des stratégies de conservation. Diverses méthodes basées sur le monitoring, l'analyse d'échantillons invasifs ou non ont été utilisées pour étudier les régimes alimentaires. Elles sont généralement biaisées ou peu résolutives. Les méthodes basées sur l'identification des fragments d'ADN dans les fèces ont le potentiel de fournir une meilleure information, notamment dans le cadre d'une approche métabarcoding. Il s'agit de caractériser simultanément l'ensemble des espèces dont l'ADN est présent dans un échantillon environnemental, en utilisant les Nouvelles Techniques de Séquençage. Dans ce cas, les amorces universelles nécessaires pour amplifier toutes les proies potentielles amplifient également l'ADN du prédateur s'il y a proximité taxonomique (par exemple mammifères). Ainsi les produits PCR obtenus à partir des fèces sont essentiellement composés d'ADN du prédateur et ne reflètent pas l'ensemble du régime alimentaire. L'utilisation d'un oligonucléotide de blocage limitant spécifiquement l'amplification de l'ADN du prédateur peut résoudre ce problème. Nous avons développé une méthode de ce type basée sur l'utilisation d'amorces universelles pour les vertébrés (amplifiant la région 12SV5) et d'oligonucléotides de blocage. Bien que non quantitative, cette méthode s'est montrée robuste, adaptée à l'étude de prédateurs à très large spectre de proies, et très résolutive pour identifier les proies au niveau du genre et de l'espèce. Nous l'avons appliquée à l'étude du régime alimentaire du chat léopard (Prionailurus bengalensis) qui s'est avéré très diversifié (mammifères, oiseaux, amphibiens et poissons) dans les deux populations du Pakistan étudiées. Avec la même approche, nous avons démontré la réalité du conflit entre l'homme et le léopard commun (Panthera pardus) dont le régime est presque exclusivement composé d'animaux domestiques. Enfin, nous avons pu proposer des actions de conservations pertinentes après avoir montré que le régime de la très menacée panthère des neiges (Panthera uncia) est principalement composé d'ongulés sauvages. / Information on food webs is central to understand ecosystem functioning. It also provides information of ecosystem stability by evaluating the resource availability and use. Obtaining information on the diet can be critical especially when dealing with elusive carnivores, which are difficult to observe. However, these large carnivores are keystone species that influence the ecosystem through trophic cascades and maintain biodiversity. Thus, precise knowledge of their diet is a prerequisite for designing conservation strategies of these endangered species. Direct and indirect monitoring as well as invasive and non-invasive approaches that have been used to study the diet are either biased or have a low resolution. The DNA-based analysis of feces is an alternative method that may provide better information. It can be implemented through a metabarcoding approach, which is the simultaneous identification of multiple species from a single environmental sample containing degraded DNA by using Next Generation Sequencing. In this case, the use of universal primers for vertebrates amplifying all potential prey also amplifies the predator DNA when it belongs to a close taxon (e.g. mammals). Thus, the PCR products obtained from feces extracts will mainly consist of predator sequences and may not represent the full diet. The use of oligonucleotides specifically blocking the amplification of the predator DNA may overcome this problem. We have developed such a method based on the concomitant use of a universal primer pair (12SV5, amplifying all vertebrates) and blocking oligonucleotides for identifying the prey DNA fragments from predators feces. Even if the method developed is not quantitative, it is robust and adequate for studying predator with a very large dietary range and has a better resolution than traditional methods for identifying prey at the genus or species level. This methodology has been applied to characterize the highly eclectic diet (mammals, birds, amphibians and fishes) of two Northern-Pakistani populations of leopard cat (Prionailurus bengalensis). With the same approach, we demonstrated the importance of the Human-leopard conflict in Pakistan, due to the almost exclusive consumption of domestic animals by the common leopard (Panthera pardus). We could also highlight relevant conservation issues for the highly endangered and cryptic snow leopard (Panthera uncia), based on the fact that it mainly fed on wild ungulates.

Les effets non-linéaires dépendant essentiellement de l’intensité du champ électrique de l’onde et du guide, sont indispensables à la génération des régimes impulsionnels dans les lasers à fibre dopée. L’effet Kerr, qui se manifeste quel que soit l’énergie de propagation et de pompage, va engendrer le phénomène de l’auto modulation de phase (SPM) qui se traduira par un élargissement spectral. La SPM peut se voir aussi dans l’interaction entre les deux ondes qui se propagent le long des axes lents et rapides du guide (XPM). Dans un guide uniforme passif, cette auto modulation de phase en se conjuguant avec la dispersion chromatique du guide peut conduire à l’impulsion soliton, mais ce cadre doit être largement dépassé pour aborder la mise en forme d’impulsions dans un système dissipatif, tel qu’une cavité laser. Ce cadre élargi est celui du soliton dissipatif. Dans ce travail, nous avons passé en revue la fibre optique dopée en tant que milieu actif. Nous avons présenté le principe physique du blocage de modes, en introduisant l’absorbant saturable virtuel reposant sur l’évolution non linéaire de la polarisation (ENLP). Dans la partie expérimentale, on s’est penché plus particulièrement sur de nouvelles dynamiques à haute cadence (multi-GHz) d’un laser à fibre dopée, à modes bloqués. Avant de présenter et positionner notre travail, on s’est intéressé dans un premier temps à ce qui a été publié sur la génération des trains d’impulsions ultracourtes à haute cadence. Nous avons étudié chaque configuration, allant de la génération de régime harmonique à haute cadence par le blocage de modes habituel jusqu’aux techniques utilisant l’instabilité de modulation avec un filtrage interférométrique (µcavité, F.P, SFBG). En évaluant les opportunités de réalisation de chacune, nous avons été attirés par la configuration de Mao, publiée dans Sci. Reports, toute fibrée, qui sera le premier volet de notre contribution expérimentale. Effectivement, dans un premier temps nous avons repris les travaux de Mao et al. où le MZI est pris en série dans la cavité fondamentale. Dans le souci d’apporter plus de stabilité, nous avons proposé une autre configuration où le MZI est pris dans une boucle de recirculation qui permettra plus de filtrage des modes cavité en phase. Cette stabilité du régime impulsionnel, à haute cadence, a été observée en temps réel. / Non-linear effects, which depend essentially on the intensity of the electric field of the wave and the guide, are essential for the generation of pulse regimes in doped fiber lasers. The Kerr effect, which occurs regardless of the propagation and pumping energy, will generate the phenomenon of phase auto modulation (SPM), which will result in a spectral expansion. The SPM can also be seen in the interaction between the two waves that propagate along the slow and fast axes of the guide (XPM). In a passive uniform guide, this self-phase modulation in combination with the chromatic dispersion of the guide can lead to the soliton pulse, but this framework must be largely overcome to address pulse shaping in a dissipative system, such as a laser cavity. This extended framework is that of the dissipative soliton. In this work, we reviewed doped optical fiber as an active medium. We presented the physical principle of mode blocking, introducing the virtual saturable absorber based on the non-linear evolution of polarization (ENLP). In the experimental part, we focused more particularly on new high rate dynamics (multi-GHz) of a doped fiber laser with blocked modes. Before presenting and positioning our work, we first focused on what was published on the generation of ultra-short high speed pulse trains. We studied each configuration, ranging from the generation of high-rate harmonic regime by blocking the usual modes to techniques using modulation instability with interferometric filtering (µcavity, F.P, SFBG). In assessing the opportunities for each, we were attracted by the configuration of Mao, published in Sci. Reports, all fiber, which will be the first part of our experimental contribution. Indeed, initially we resumed the work of Mao et al. where the MZI is taken in series in the fundamental cavity. In order to provide more stability, we have proposed another configuration where the MZI is caught in a recirculation loop that will allow more filtering of the cavity modes in phase. This stability of the impulse regime, at high repetition rate, was observed in real time.

Os relaxantes neuromusculares não despolarizantes são fármacos indispensáveis nos procedimentos cirúrgicos que requerem entubação endotraqueal, visto que diminuem o tônus muscular. Quimicamente são divididos em derivados isoquinolinicos e derivados de esteróides, esses últimos com maior aplicação clinica e comercialização no Brasil. Assim sendo, é importante a validação de métodos analíticos para o controle de qualidade com alternativas confiáveis que garantam sua eficácia e segurança. Os brometos de vecurônio, de pancurônio e de rocurônio são fármacos relaxantes neuromusculares não despolarizantes derivados de esteróides. As propriedades farmacológicas destes fármacos são significativamente diferentes entre si, porém, as propriedades químicas são bastante similares. Na presente pesquisa foram desenvolvidos e validados métodos analíticos de separação (cromatografia líquida de alta eficiência e eletroforese capilar) para cada fármaco. Estes métodos foram aplicados a medicamentos comercializados no Brasil. Para todos os métodos cromatográficos foi utilizada uma coluna amino devido a seu caráter polar. Em função da baixa absorção dos três fármacos na região ultravioleta, os métodos eletroforéticos foram aplicados com detecções indiretas utilizando substâncias cromóforas. Comparando-se as técnicas utilizadas para determinação dos fármacos nos medicamentos isoladamente, não houve diferença significativa com nível de confiança de 95,0%. Nos testes de hipótese aplicados (F-Snedecor e t-Student), não foram observadas diferenças na precisão (variâncias) e no valor encontrado dos fármacos contidos nas amostras estudadas (comparação de duas médias). / The non-depolarizing neuromuscular relaxant drugs are essential in surgical procedures requiring endotracheal intubation, as they decrease muscle tonics. These drugs are chemically divided in isoquinolines derivatives and steroid derivatives, this latter group, with greater clinical application and commercialization in Brazil. Therefore, the study and validation of analytical methods are important for quality control with reliable alternatives to ensure their effectiveness and safety. The vecuronium, pancuronium and rocuronium bromides are steroid derivatives presenting non-depolarizing neuromuscular relaxant action. Pharmacological properties of these drugs differ significantly, however, its chemical properties are quite similar. In this research, analytical separation methods (high performance liquid chromatography and capillary electrophoresis), for each drug, were developed and fully validated. The methods were applied to pharmaceutical formulations commercialized in Brazil. For all chromatographic methods, an amino column was used, due to its polar characteristics. Due to the low absorption of the three drugs in the ultraviolet region, electrophoretic methods has been applied with indirect detection using chromophoric substances. When comparing both techniques used for quantitative determination of these drugs in pharmaceutical products, no significant difference was observed using a confidence level of 95.0%. By applying hypothesis tests (F-Snedecor and t-Student), it was not observed difference in precision (variance) and in the found amount of drugs contained in the selected samples (comparison of two means).

Introdução e objetivos: a monitorização neuromuscular objetiva é prática médica baseada em evidências e deve ser empregada rotineiramente quando do uso de fármacos bloqueadores neuromusculares. Entretanto, pesquisas relacionadas a esta monitorização em pacientes pediátricos não estão vastamente documentadas como nos adultos. Em pesquisa clínica, o monitor neuromuscular deve apresentar resposta estável (menor que 5% de variação na altura de T1) por um período de dois a cinco minutos antes da administração do bloqueador neuromuscular. O tempo necessário para se alcançar esta estabilidade na resposta pode variar, mas pode ser encurtado por meio da aplicação de um estímulo tetânico por cinco segundos. Aventouse a hipótese de que a aplicação de estímulo tetânico antes da calibração poderia levar a diferenças nos parâmetros de início de ação e nos tempos de recuperação. O objetivo primário deste estudo foi comparar o tempo de início de ação e os tempos de recuperação após dose única de rocurônio 0,6 mg/kg seguido de recuperação espontânea, entre dois grupos de pacientes com sequências diferentes para a calibração (com e sem o uso de estímulo tetânico). Os objetivos secundários foram a avaliação da altura inicial e final de T1, tempo para se obter estabilidade da altura de T1 e os seguintes ajustes do monitor neuromuscular: corrente elétrica e sensibilidade. Método: consentimento informado dos responsáveis, foram incluídos no estudo 50 pacientes, estado físico 1 ou 2, de dois a 11 anos, agendados para cirurgias abdominais e/ou perineais com tempo cirúrgico estimado superior a 60 minutos. Os pacientes (25 por grupo) foram submetidos a anestesia intravenosa e alocados randomicamente para receber estímulo tetânico (grupo T) ou não (grupo C), antes da calibração do monitor. Após a calibração do monitor, a modalidade sequência de quatro estímulos foi iniciada e mantida em intervalo de 15 segundos. Resultados: não houve diferença significativa no início de ação (C: 57,5±16,9 versus T: 58,3±31,2 s; p=0,917). O tempo normalizado para as relações da sequência de quatro estímulos 0,7, 0,8 e 0,9 diferiram significativamente entre os grupos (C: 40,1 ± 7,9 versus T: 34,8±10 min; p=0,047, C: 43,8±9,4 versus T: 37,4±11 min; p=0,045 e C: 49,9±12,2 versus T: 41,7±13,1 min; p=0,026, respectivamente). O tempo necessário para a estabilização da altura de T1 não mostrou diferença estatística entre os grupos (C: 195±203 versus T: 116±81,6 s; p=0,093). Os valores de altura inicial de T1 mostraram diferença significativa entre os grupos (C: 98 versus T: 82,7%; p < 0,001). Os valores de altura final de T1 também mostraram diferença significante entre os grupos (C: 95,3 versus T: 69,3%; p < 0,001). Conclusões: o estímulo tetânico encurtou o tempo normalizado das relações da sequência de quatro estímulos 0,7, 0,8 e 0,9. As alturas inicial e final de T1 foram menores no grupo tétano. Não houve diferença estatística entre os grupos relativo ao tempo necessário para estabilização da altura de T1. Os ajustes do monitor (corrente elétrica e sensibilidade) não apresentaram diferenças entre os gruposEffect of tetanic stimulation, prior to calibration, on the onset of action and recovery times of neuromuscular blockade in pediatric patients / Background and objective: objective neuromuscular monitoring is evidence-based medical practice and should be routinely used when using neuromuscular blocking drugs. However, research related to this monitoring in pediatric patients is not widely documented as in adults. In clinical research, the neuromuscular monitor should have a stable response (less than 5% change in T1 height) for a period of two to five minutes before administration of the neuromuscular block agent. The time required to achieve this stability in response may vary, but may be shortened by the application of a tetanic stimulus for 5 seconds. It was hypothesized that the application of tetanic stimulus prior to calibration could lead to differences in the parameters of onset of action and recovery times. The primary outcome of this study was to compare time to onset and recovery times after single dose rocuronium 0.6 mg.kg-1 followed by spontaneous recovery between two groups of patients with different sequences for calibration (with and without use of tetanic stimulus). The secondary outcomes were the evaluation of the initial and final T1 height, time to obtain stability of T1 height and the following neuromuscular monitor settings: electric current and sensitivity. Methods: after approval by the Institutional Ethics Committee and obtaining the informed consent of those responsible for the patient, were included in the study 50 patients, physical status 1 or 2, from 2 to 11 years, scheduled for abdominal and/or perineal surgeries with estimated surgical time greater than 60 minutes. Patients (25 per group) underwent intravenous anesthesia and were allocated randomly to receive tetanus stimulation (group T) or not (group C) prior to calibration of the monitor. After calibration of the monitor, train-of-four mode was initiated and maintained at interval of 15 seconds. Results: there was no significant difference in onset of action (C: 57,5±16,9 versus T: 58,3±31,2 s; p=0,917). The train-of-four normalized times 0.7, 0.8 and 0.9 differed significantly between the groups (C: 40,1±7,9 versus T: 34,8±10 min; p=0,047, C: 43,8±9,4 versus T: 37,4±11 min; p=0,045 and C: 49,9±12,2 versus T: 41,7±13,1 min; p=0,026, respectively). The time required to stabilize the T1 height did not show statistical difference between the groups (C: 195±203 versus T: 116±81,6 s; p=0,093), The initial values of T1 height showed a significant difference between the groups (C: 98 versus T: 82,7%; p < 0,001). The final T1 height values also showed a significant difference between the groups (C: 95,3 versus T: 69,3%; p < 0,001). Conclusions: the tetanic stimulus shortened the normalized time of the fourstimulus sequence ratios 0.7, 0.8, and 0.9. The initial and final T1 heights were lower in the tetanus group. There was no statistical difference between the groups regarding the time required to stabilize the T1 height. The monitor settings (electric current and sensitivity) did not show differences between groups. Trial registration: Clinicaltrials.gov identifier: NCT0249867

<p>This thesis deals with the development of efficient and reliable algorithms and library software for factorizing matrices and solving matrix equations on high-performance computer systems. The architectures of today's computers consist of multiple processors, each with multiple functional units. The memory systems are hierarchical with several levels, each having different speed and size. The practical peak performance of a system is reached only by considering all of these characteristics. One portable method for achieving good system utilization is to express a linear algebra problem in terms of level 3 BLAS (Basic Linear Algebra Subprogram) transformations. The most important operation is GEMM (GEneral Matrix Multiply), which typically defines the practical peak performance of a computer system. There are efficient GEMM implementations available for almost any platform, thus an algorithm using this operation is highly portable.</p><p>The dissertation focuses on how recursion can be applied to solve linear algebra problems. Recursive linear algebra algorithms have the potential to automatically match the size of subproblems to the different memory hierarchies, leading to much better utilization of the memory system. Furthermore, recursive algorithms expose level 3 BLAS operations, and reveal task parallelism. The first paper handles the Cholesky factorization for matrices stored in packed format. Our algorithm uses a recursive packed matrix data layout that enables the use of high-performance matrix--matrix multiplication, in contrast to the standard packed format. The resulting library routine requires half the memory of full storage, yet the performance is better than for full storage routines.</p><p>Paper two and tree introduce recursive blocked algorithms for solving triangular Sylvester-type matrix equations. For these problems, recursion together with superscalar kernels produce new algorithms that give 10-fold speedups compared to existing routines in the SLICOT and LAPACK libraries. We show that our recursive algorithms also have a significant impact on the execution time of solving unreduced problems and when used in condition estimation. By recursively splitting several problem dimensions simultaneously, parallel algorithms for shared memory systems are obtained. The fourth paper introduces a library---RECSY---consisting of a set of routines implemented in Fortran 90 using the ideas presented in paper two and three. Using performance monitoring tools, the last paper evaluates the possible gain in using different matrix blocking layouts and the impact of superscalar kernels in the RECSY library. </p>

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