Electrochemical characterization of Dps, a DNA-protecting protein

Dissertação para obtenção do Grau de Mestre em Biotecnologia

Susceptibility of Cebus apella monkey (Primates: Cebidae) to experimental Leishmania (L.) infantum chagasi-infection

In Amazonian Brazil, the Cebus apella monkey (Primates: Cebidae) has been associated with the enzootic cycle of Leishmania (V.) shawi, a dermotropic parasite causing American cutaneous leishmaniasis (ACL). It has also been successfully used as animal model for studying cutaneous leishmaniasis. In this work, there has been investigated its susceptibility to experimental Leishmania (L.) infantum chagasi-infection, the etiologic agent of American visceral leishmaniasis (AVL). There were used ten C. apella specimens, eight adult and two young, four males and six females, all born and raised in captivity. Two experimental infection protocols were performed: i) six monkeys were inoculated, intra-dermal via (ID), into the base of the tail with 2 x 10(6) promastigotes forms from the stationary phase culture medium; ii) other four monkeys were inoculated with 3 x 10(7) amastigotes forms from the visceral infection of infected hamsters by two different via: a) two by intravenous via (IV) and, b) other two by intra-peritoneal via (IP). The parameters of infection evaluation included: a) clinical: physical exam of abdomen, weigh and body temperature; b) parasitological: needle aspiration of the bone-marrow for searching of amastigotes (Giemsa-stained smears) and promastigotes forms (culture medium); c) immunological: Indirect fluorescence antibody test (IFAT) and, Delayed-type hypersensitivity (DTH). In the six monkeys ID inoculated (promastigotes forms) all parameters of infection evaluation were negative during the 12 months period of follow-up. Among the four monkeys inoculated with amastigotes forms, two IV inoculated showed the parasite in the bone-marrow from the first toward to the sixth month p.i. and following that they cleared the infection, whereas the other two IP inoculated were totally negative. These four monkeys showed specific IgG-antibody response since the third month p.i. (IP: 1/80 and IV: 1/320 IgG) toward to the 12th month (IP: 1/160 and IV: 1/5120). The DTH-conversion occurred in only one IV inoculated monkey with a strong (30 mm) skin reaction. Considering these results, we do not encourage the use of C. apella monkey as animal model for studying the AVL.

In vitro and in vivo antileishmanial efficacy of a combination therapy of diminazene and artesunate against Leishmania donovani in BALB/c mice

The in vitro and in vivo activity of diminazene (Dim), artesunate (Art) and combination of Dim and Art (Dim-Art) against Leishmania donovani was compared to reference drug; amphotericin B. IC50 of Dim-Art was found to be 2.28 ± 0.24 µg/mL while those of Dim and Art were 9.16 ± 0.3 µg/mL and 4.64 ± 0.48 µg/mL respectively. The IC50 for Amphot B was 0.16 ± 0.32 µg/mL against stationary-phase promastigotes. In vivo evaluation in the L. donovani BALB/c mice model indicated that treatments with the combined drug therapy at doses of 12.5 mg/kg for 28 consecutive days significantly (p < 0.001) reduced parasite burden in the spleen as compared to the single drug treatments given at the same dosages. Although parasite burden was slightly lower (p < 0.05) in the Amphot B group than in the Dim-Art treatment group, the present study demonstrates the positive advantage and the potential use of the combined therapy of Dim-Art over the constituent drugs, Dim or Art when used alone. Further evaluation is recommended to determine the most efficacious combination ratio of the two compounds.

Apoptotic-like activity of staurosporine in axenic cultures of Trypanosoma evansi

Trypanosoma evansi is a blood protozoan parasite of the genus Trypanosoma which is responsible for surra (Trypanosomosis) in domestic and wild animals. This study addressed apoptotic-like features in Trypanosoma evansi in vitro. The mechanism of parasite death was investigated using staurosporine as an inducing agent. We evaluated its effects through several cytoplasmic features of apoptosis, including cell shrinkage, phosphatidylserine exposure, maintenance of plasma membrane integrity, and mitochondrial trans-membrane potential. For access to these features we have used the flow cytometry and fluorescence microscopy with cultures in the stationary phase and adjusted to a density of 10(6) cells/mL. The apoptotic effect of staurosporine in T. evansi was evaluated at 20 nM final concentration. There was an increase of phosphatidylserine exposure, whereas mitochondrial potential was decreased. Moreover, no evidence of cell permeability increasing with staurosporine was observed in this study, suggesting the absence of a necrotic process. Additional studies are needed to elucidate the possible pathways associated with this form of cell death in this hemoparasite.

Studies on BolA and ribonuclease R: two important factors in the control of bacterial gene expression

Dissertation presented to obtain the Ph.D degree in Biology

On-line system for faults detection in induction motors based on PCA

Dissertation to obtain the degree of Master in Electrical and Computer Engineering

Tumors of the Brachial Plexus in a Tertiary Referral Center: a Case Series and Literature Review

Introduction: Brachial plexus (BP) tumors are very rare tumors, with less than 800 cases been described in the literature worldwide since 1970. These tumors often present as local or radicular pain, with scant or no neurological deficits. These symptoms are shared by many other more common rheumatologic diseases, thus making their diagnosis difficult in most cases. Additionally, these tumors often present as lumps and are therefore biopsied, which carries a significant risk of iatrogenic nerve injury. Material and Methods: In this paper the authors describe their experience with the management of 5 patients with BP tumors followed up for at least 2 years. There were 4 males and 1 female. Median follow-up time was 41 ± 21 months. Average age at diagnosis was 40,0 ± 19,9 years. The most common complaints at presentation were pain and sensibility changes. All patients had a positive Tinel sign when the lesion was percussed. In all patients surgery was undertaken and the tumors removed. In 4 patients nerve integrity was maintained. In one patient with excruciating pain a segment of the nerve had to be excised and the nerve defect was bridged with sural nerve grafts. Results: Pathology examination of the resected specimens revealed a Schwannoma in 4 cases and a neurofibroma in the patient submitted to segmental nerve resection. Two years postoperatively, no recurrences were observed. All patients revealed clinical improvement. The patient submitted to nerve resection had improvement in pain, but presented diminished strength and sensibility in the involved nerve territory. Conclusion: Surgical excision of BP tumors is not a risk free procedure. Most authors suggest surgery if the lesion is symptomatic or progressing in size. If the tumor is stationary and not associated with neurological dysfunction a conservative approach should be taken.

Customer Lifetime Value: a framework for application in the insurance industry - Building a Business Process to Generate and Maintain an Automatic Estimation Agent

Research Project submited as partial fulfilment for the Master Degree in Statistics and Information Management

Estudo da expressão génica da Legionella pneumophila estirpe paris após co-cultura em Acanthamoeba castellanii

RESUMO: A Legionella é um bacilo Gram-negativo que replica dentro de protozoários como Acanthamoeba castellanii (A. castellanii) e no interior de macrófagos alveolares humanos, podendo resultar numa pneumonia grave. A Legionella em meio líquido tem um ciclo de vida bifásico, apresentando traços replicativos na fase exponencial e expressando factores transmissíveis na fase estacionária. Estudos recentes demonstraram que a Legionella precisa de assegurar um tempo preciso no seu ciclo de vida para efectuar com êxito a infecção das células hospedeiras. Muitos modelos de estudo foram desenvolvidos a fim de aumentar o conhecimento sobre o ciclo de vida intracelular e identificar os genes necessários para a modulação da célula hospedeira. Embora o conhecimento sobre a interacção bactéria-hospedeiro ainda seja limitado, parece que esta interacção gera um conjunto de características de virulência permitindo que a bactéria infecte células fagocíticas humanas e cause doença. O objectivo do presente projecto de investigação foi investigar e seleccionar genes críticos para a infecciosidade da Legionella pneumophila estirpe Paris (Lp Paris), desenhar e optimizar uma técnica de PCR em tempo real para o estudo da expressão génica e comparar o perfil de expressão da Lp Paris antes e depois da co-cultura em A. castellanii. Os resultados mostraram que oito dos 12 genes em estudo alteraram a sua expressão relativa após co-cultura em A. castellanii quando os ensaios foram realizados com culturas de Lp Paris na fase estacionária precoce (cinco foram induzidos e três reprimidos) Quando os ensaios foram realizados com culturas de Lp Paris na fase estacionária tardia 11 genes apresentaram repressão na sua expressão relativa. Analisando os resultados, concluímos que o perfil de expressão de Lp Paris foi modificado pela interacção com A. castellanii, no entanto essa mudança foi dependente da fase do seu ciclo de vida.-------ABSTRACT: Legionella is a pathogenic Gram-negative bacterium that replicates not only within aquatic protozoa like Acanthamoeba castellanii (A. castellanii), but also within human alveolar macrophages, which can result in a severe pneumonia. Legionella has a biphasic life cycle in broth, where exponential phase cultures display replicative traits and stationary bacteria express transmissive factors. Recent studies demonstrated that for successful infection of host cells, Legionella needs to ensure a precise timing of its life cycle. Many models of study were developed in order to learn about the intracellular life cycle and to identify the genes necessary for the host cell modulation. Although knowledge about the bacteria-host interaction is still limited, it appears that this interaction generate a pool of virulence traits, allowing the bacterium to infect human phagocytic cells and cause disease. The purpose of the present study was to investigate and select de critical genes for the infectivity of Legionella pneumophila strain Paris (Lp Paris), design and optimize a real time PCR technique for gene expression study and compare the expression profile of Lp Paris before and after co- culture of A. castellanii. The results show that eight of 12 genes in study changed its relative expression after coculture in A. castellanii when we performed the intracellular assays with early stationary phase Lp Paris cultures (five were induced and tree were repressed). When we performed the intracellular assays with late stationary phase Lp Paris cultures 11 genes showed a repressed relative expression. Analysing the results, we conclude that the expression profile of Lp Paris was modified by interaction with A. castellanii but this change was dependent of the timing of its life cycle.

Studies on the wine spoilage yeast Brettanomyces bruxellensis: 4-ethylphenol production and improved detection

Dissertação para obtenção do Grau de Mestre em Biotecnologia

Thermo-hydro-mechanical analysis of CO2 injection into deep aquifers

One of the biggest challenges for humanity is global warming and consequently, climate changes. Even though there has been increasing public awareness and investments from numerous countries concerning renewable energies, fossil fuels are and will continue to be in the near future, the main source of energy. Carbon capture and storage (CCS) is believed to be a serious measure to mitigate CO2 concentration. CCS briefly consists of capturing CO2 from the atmosphere or stationary emission sources and transporting and storing it via mineral carbonation, in oceans or geological media. The latter is referred to as carbon capture and geological storage (CCGS) and is considered to be the most promising of all solutions. Generally it consists of a storage (e.g. depleted oil reservoirs and deep saline aquifers) and sealing (commonly termed caprock in the oil industry) formations. The present study concerns the injection of CO2 into deep aquifers and regardless injection conditions, temperature gradients between carbon dioxide and the storage formation are likely to occur. Should the CO2 temperature be lower than the storage formation, a contractive behaviour of the reservoir and caprock is expected. The latter can result in the opening of new paths or re-opening of fractures, favouring leakage and compromising the CCGS project. During CO2 injection, coupled thermo-hydro-mechanical phenomena occur, which due to their complexity, hamper the assessment of each relative influence. For this purpose, several analyses were carried out in order to evaluate their influences but focusing on the thermal contractive behaviour. It was finally concluded that depending on mechanical and thermal properties of the pair aquifer-seal, the sealing caprock can undergo significant decreases in effective stress.

Online consumer behavior: The tablet shopper – A new kind of customer?

Tablet computers are on the rise and are increasingly superseding stationary computers in terms of modern online shopping. This paper therefore aims on understanding how tablet and website characteristics might impact online consumer behavior. The collected data resulted from focus groups and in-depth interviews conducted with a technology affine audience. It has shown that tablets indeed may have shifted shopping behavior from utilitarian towards rather recreational attributes. Discussions also revealed high customer expectations towards visited shopping websites and prevailing deficits. Thus, harnessing the concept of experiential flow is crucial. Compelling experiences are required to increase the duration of stay and consequently the likelihood of increased purchases.

Optimal design and operation of compact simulated moving bed processes for enantioseparations

Simulated moving bed (SMB) chromatography is attracting more and more attention since it is a powerful technique for complex separation tasks. Nowadays, more than 60% of preparative SMB units are installed in the pharmaceutical and in the food in- dustry [SDI, Preparative and Process Liquid Chromatography: The Future of Process Separations, International Strategic Directions, Los Angeles, USA, 2002. http://www. strategicdirections.com]. Chromatography is the method of choice in these ¯elds, be- cause often pharmaceuticals and ¯ne-chemicals have physico-chemical properties which di®er little from those of the by-products, and they may be thermally instable. In these cases, standard separation techniques as distillation and extraction are not applicable. The noteworthiness of preparative chromatography, particulary SMB process, as a sep- aration and puri¯cation process in the above mentioned industries has been increasing, due to its °exibility, energy e±ciency and higher product purity performance. Consequently, a new SMB paradigm is requested by the large number of potential small- scale applications of the SMB technology, which exploits the °exibility and versatility of the technology. In this new SMB paradigm, a number of possibilities for improving SMB performance through variation of parameters during a switching interval, are pushing the trend toward the use of units with smaller number of columns because less stationary phase is used and the setup is more economical. This is especially important for the phar- maceutical industry, where SMBs are seen as multipurpose units that can be applied to di®erent separations in all stages of the drug-development cycle. In order to reduce the experimental e®ort and accordingly the coast associated with the development of separation processes, simulation models are intensively used. One impor- tant aspect in this context refers to the determination of the adsorption isotherms in SMB chromatography, where separations are usually carried out under strongly nonlinear conditions in order to achieve higher productivities. The accurate determination of the competitive adsorption equilibrium of the enantiomeric species is thus of fundamental importance to allow computer-assisted optimization or process scale-up. Two major SMB operating problems are apparent at production scale: the assessment of product quality and the maintenance of long-term stable and controlled operation. Constraints regarding product purity, dictated by pharmaceutical and food regulatory organizations, have drastically increased the demand for product quality control. The strict imposed regulations are increasing the need for developing optically pure drugs.(...)

A ação combinada da carbonatação e dos cloretos: avaliação do coeficiente de difusão de cloretos em betões carbonatados

Dissertação de mestrado Internacional em Sustentabilidade do Ambiente Construído

Real-time monitoring of progression towards renal failure in primary care patients

First published online: December 16, 2014.