Dengue type 2 outbreak in the south of the State of Bahia, Brazil: laboratorial and epidemiological studies

During March 1994 cases of a exanthematic acute disease were reported in the municipalities of Itagemirim, Eunápolis and Belmonte, state of Bahia. Dengue fever was confirmed by serology (MAC-ELISA) and by dengue virus type 2 isolation, genotype Jamaica. Signs and symptoms of classic dengue fever were observed with a high percentual of rash (73.8%) and pruritus (50.5%). Major haemorrhagic manifestations were unfrequent and only bleeding gum was reported. Dengue virus activity spreaded rapidly to important tourism counties like Porto Seguro, Ilhéus, Santa Cruz de Cabrália, Prado, Alcobaça and others, representing a risk for the spreading of dengue virus into the country and abroad.

Plasma levels of tumor necrosis factor-alpha in patients with visceral leishmaniasis (Kala-Azar). Association with activity of the disease and clinical remisson following antimonial therapy

Evaluation of TNF-alpha in patients with Kala-azar has drawn increasing interest due to its regulatory role on the immune system, in addition to its cachetizing activity. The objective of this study was to examine the association between plasma levels of TNF-alpha, measured by immunore-activity (ELISA) and bioactivity (cytotoxicity assay with L-929 cells), and clinical manifestations of visceral leishmaniasis. Plasma samples from 19 patients with Kala-azar were obtained before, during and at the end of antimonial therapy. TNF-alpha determinations was done by using the cytotoxicity assay (all patients) and the enzyme-linked immunoassay (ELISA - 14 patients). A discrepancy between results obtained by ELISA and cytotoxicity assay was observed. Levels of circulating TNF-alpha, assessed by ELISA, were higher in patients than in healthy controls, and declined significantly with improvement in clinical and laboratory parameters. Plasma levels before treatment were 124.7 ± 93.3 pg/ml (mean ± SD) and were higher than at the end of therapy 13.9 ± 25.1 pg/ml (mean ± SD) (p = 0.001). In contrast, plasma levels of TNF-alpha evaluated by cytotoxicity assay did not follow a predicted course during follow-up. Lysis, in this case, might be not totally attributed to TNF-alpha. The discrepancy might be attributed to the presence of factor(s) known to influence the release and activity of TNF-alpha.

Genetics of berry colour and anthocyanin content variation in grapevine (Vitis vinifera L.subsp. vinifera)

Dissertation presented in fulfillment of the requirements for the Degree of Doctor of Philosophy in Biology (Molecular Genetics) at the Instituto de Tecnologia Química e Biológica da Universidade Nova de Lisboa

Archaeometallurgical study of artefacts from Castro de Vila Nova de São Pedro (Azambuja, Portugal)

Dissertação para obtenção do Grau de Mestre em Conservação e Restauro

Study of materials, technique and conservation treatment of Sinfonia Heroica by Dordio Gomes

Dissertação apresentada na Faculdade de Ciências e Tecnologia da Universidade Nova de Lisboa para obtenção do grau de Mestre em Conservação e Restauro

Molecular modulation of brain development and function: lessons from “old” and “new”

Dissertation presented to obtain the Ph.D degree in Biology

The removal of xenobiotic compounds from wastewater through the use of biological processes and advanced oxidation technologies

Dissertação apresentada para obtenção do Grau de Doutor em Engenharia Química e Bioquímica pela Universidade Nova de Lisboa, Faculdade de Ciências e Tecnologia

Seroprevalence of Chagas disease in schoolchildren from two municipalities of Jequitinhonha Valley, Minas Gerais, Brazil; six years following the onset of epidemiological surveillance

Six years after the beginning of the epidemiological surveillance of Chagas disease in Berilo and José Gonçalves de Minas, Jequitinhonha Valley, MG, Brazil, a serological inquiry was performed to observe whether the transmission of this endemy was occurring in this area. A randomized sample of 1,412 children seven to 14 years old, was screened. Six asymptomatic children were found to be positive, leading to 0.4% of prevalence. Hemoculture confirmed infection in five out of the six positive cases. Additional epidemiological investigation revealed important antecedents, such as disease reports in relatives and predisposing ecological and housing conditions. Our results demonstrated similar seroprevalence (0.4%) in schoolchildren, ranging from seven to 14 years old, and that were observed six years ago (0.2%) for children 0-9 year-old. Thus, considering the constant presence of Panstrogylus megistus in the peridomicile these findings emphasize the need of continuous improved epidemiological surveillance of Chagas disease in this region.

Epidemiologic features of HIV infection in three municipalities of inner Rio de Janeiro State, Brazil

In Brazil relatively little attention is being paid to the study of the features of the spread of the AIDS epidemic towards small cities and rural areas. We report a descriptive study on the epidemiological features of HIV infection among 208 adult patients seen between July 1999 and May 2006 in the municipal HIV/AIDS Programs of three cities of inner Rio de Janeiro State: Saquarema, Santo Antonio de Pádua and Miracema. A portrait of a heterosexual epidemic emerged, with an overall male to female ratio of 1.1. More than 90% were residents of the studied cities, demonstrating a local demand for HIV-related assistance and the importance of municipal HIV/AIDS Programs. Past or current use of snorted cocaine was reported by a quarter of the patients. Older age and male gender were independent predictors of having a diagnosis of AIDS at presentation. The latter is in accordance with a more recent wave of epidemic spread towards female gender. A low frequency of male circumcision, an important determinant of heterosexual HIV transmission, was recorded. Almost 60% of the patients first presented in advanced stages of HIV infection, suggesting the existence of a large pool of undiagnosed cases in the community.

An assessment of the social and economic cohesion aspects of the development of the information society in Europe

Neste trabalho pretende consolidar-se a contribuição portuguesa para o estudo comunitário “Na Assessment of the Social and Economic Cohesion Aspects of the Development of the Information Society in Europe” elaborado por um consórcio europeu liderado pela Nexus Europe e em que intervém o ISEGI- Instituto Superior de Estatística e Gestão de Informação, da Universidade Nova de Lisboa, como parceiro nacional português.

The inhibitory activity of synthetic compounds and ions against transporters of multi-drug resistant bacteria

RESUMO: Sessenta e três derivados de hidantoína foram utilizados para avaliar possíveis efeitos de modulação na actividade das bombas de efluxo (BE) na Salmonella NCTC 13349 utilizando um método fluorimétrico semi-automático. Nenhum dos compostos apresentaram actividade anti-bacteriana até concentrações de 240 mg/L. Entre todos os compostos, SZ-7 demonstrou possuir propriedades de modulação de effluxo na presença de glucose. Para testar esta actividade, estirpes de Salmonella resistentes à ciprofloxacina, induzidas a elevados níveis de resistência com sobre-expressão de BE, foram expostas ao SZ-7. Este derivado afectou a susceptibilidade das estirpes à ciprofloxacina. Uma vez que os 63 compostos estudados apresentaram pouco efeito inibitório /cumulativo, apesar de serem conhecidos pelos seus efeitos moduladores de BE-dependentes de iões em eucariotas, foi questionado o papel dos iões na regulação de BE bacterianas, que poderão influenciar a eficácia de novos compostos. Para este estudo, utilizamos a Escherichia coli AG100 como modelo, devido ao extenso conhecimento no que respeita a estrutura e actividade das BE. Devido à importância de iões de cálcio (Ca2+) nos canais de transporte membranar e na actividade de ATPases, a sua actividade na modulação do efluxo foi investigada. De resultados anteriormente obtidos concluiu-se que a pH 5 o efluxo é independente de energia metabólica; contudo, a pH 8 é absolutamente dependente, sendo que o Ca2+ é indispensável para manter a actividade das ATPases bacterianas. A acumulação/effluxo de EtBr pela E. coli AG100 foi determinada na presença/ausência de Ca2+, clorpromazina (inibidor de ligação de Ca2+ a proteínas), e ácido etilenodiamino tetra-acético (quelante de Ca2+). Acumulação/effluxo aumentou a pH 8, contudo o Ca2+ reverte estes efeitos evidenciando a sua importância no funcionamento das BE bacterianas. Em resumo este trabalho colocou em evidência que muitos aspectos bioquímicos e bioenergéticos devem ser tomados em consideração no estudo da resistência bacteriana mediada por BE.------- ABSTRACT: Sixty-three hydantoin derivatives were evaluated for their modulating effects on efflux pump (EP) activity of Salmonella NCTC 13349 utilizing a semi-automatic fluorometric method. None of the compounds presented antibacterial activities at concentrations as high as 240 mg/L. Among all compounds, SZ-7 showed possible efflux modulating activity in the presence of glucose, indicative of a potential EP inhibitor. To verify its potential effects, ciprofloxacin-resistant Salmonella strains, induced to high level resistance with over-expressing EPs, were exposed to SZ-7. This derivative affected the susceptibility of the ciprofloxacin-resistant strains. Since the 63 compounds studied had very low inhibitory/accumulative effects, even though their known for being efficient in modulating ion-driven eukaryotic EPs, we questioned whether ions had a leading role in regulating bacterial EPs, influencing the effectiveness of new compounds. For this study we used Escherichia coli AG100 as a model, due to the extensive knowledge on its EPs structure and activity. Owing the importance of calcium ions (Ca2+) for membrane transport channels and activity of ATPases, the role of Ca2+ was investigated. From previous results we concluded that at pH 5 efflux is independent of metabolic energy; however, at pH 8 it is entirely dependent of metabolic energy and the Ca2+ ions are essential to maintain the activity of bacterial ATPases. Accumulation and efflux of ethidium bromide (EtBr) by E. coli AG100 was determined in the presence and absence of Ca2+, chlorpromazine (inhibitor of Ca2+-binding to proteins), and ethylenediaminetetraacetic acid (Ca2+ chelator). Accumulation of EtBr increased at pH 8; however Ca2+ reversed these effects providing information as to the importance of this ion in the regulation of bacterial EP systems. Overall this work puts in evidence that many biochemical and bioenergetic aspects related to the strains physiology need to be taken into consideration in bacterial drug resistance mediated by EPs.

Ankylosing spondylitis: genomic and functional characterization of candidate genes and their repercussion in clinical practice

RESUMO: Introdução: A espondilite anquilosante (EA) é uma doença inflamatória crónica caracterizada pela inflamação das articulações sacroilíacas e da coluna. A anquilose progressiva motiva uma deterioração gradual da função física e da qualidade de vida. O diagnóstico e o tratamento precoces podem contribuir para um melhor prognóstico. Neste contexto, a identificação de biomarcadores, assume-se como sendo muito útil para a prática clínica e representa hoje um grande desafio para a comunidade científica. Objetivos: Este estudo teve como objetivos: 1 - caracterizar a EA em Portugal; 2 - investigar possíveis associações entre genes, MHC e não-MHC, com a suscetibilidade e as características fenotípicas da EA; 3 - identificar genes candidatos associados a EA através da tecnologia de microarray. Material e Métodos: Foram recrutados doentes com EA, de acordo com os critérios modificados de Nova Iorque, nas consultas de Reumatologia dos diferentes hospitais participantes. Colecionaram-se dados demográficos, clínicos e radiológicos e colhidas amostras de sangue periférico. Selecionaram-se de forma aleatória, doentes HLA-B27 positivos, os quais foram tipados em termos de HLA classe I e II por PCR-rSSOP. Os haplótipos HLA estendidos foram estimados pelo algoritmo Expectation Maximization com recurso ao software Arlequin v3.11. As variantes alélicas dos genes IL23R, ERAP1 e ANKH foram estudadas através de ensaios de discriminação alélica TaqMan. A análise de associação foi realizada utilizando testes da Cochrane-Armitage e de regressão linear, tal como implementado pelo PLINK, para variáveis qualitativas e quantitativas, respetivamente. O estudo de expressão génica foi realizado por Illumina HT-12 Whole-Genome Expression BeadChips. Os genes candidatos foram validados usando qPCR-based TaqMan Low Density Arrays (TLDAs). Resultados: Foram incluídos 369 doentes (62,3% do sexo masculino, com idade média de 45,4 ± 13,2 anos, duração média da doença de 11,4 ± 10,5 anos). No momento da avaliação, 49,9% tinham doença axial, 2,4% periférica, 40,9% mista e 7,1% entesopática. A uveíte anterior aguda (33,6%) foi a manifestação extra-articular mais comum. Foram positivos para o HLA-B27, 80,3% dos doentes. Os haplótipo A*02/B*27/Cw*02/DRB1*01/DQB1*05 parece conferir suscetibilidade para a EA, e o A*02/B*27/Cw*01/DRB1*08/DQB1*04 parece conferir proteção em termos de atividade, repercussão funcional e radiológica da doença. Três variantes (2 para IL23R e 1 para ERAP1) mostraram significativa associação com a doença, confirmando a associação destes genes com a EA na população Portuguesa. O mesmo não se verificou com as variantes estudadas do ANKH. Não se verificou associação entre as variantes génicas não-MHC e as manifestações clínicas da EA. Foi identificado um perfil de expressão génica para a EA, tendo sido validados catorze genes - alguns têm um papel bem documentado em termos de inflamação, outros no metabolismo da cartilagem e do osso. Conclusões: Foi estabelecido um perfil demográfico e clínico dos doentes com EA em Portugal. A identificação de variantes génicas e de um perfil de expressão contribuem para uma melhor compreensão da sua fisiopatologia e podem ser úteis para estabelecer modelos com relevância em termos de diagnóstico, prognóstico e orientação terapêutica dos doentes. -----------ABSTRACT: Background: Ankylosing Spondylitis (AS) is a chronic inflammatory disorder characterized by inflammation in the spine and sacroiliac joints leading to progressive joint ankylosis and in progressive deterioration of physical function and quality of life. An early diagnosis and early therapy may contribute to a better prognosis. The identification of biomarkers would be helpful and represents a great challenge for the scientific community. Objectives: The present study had the following aims: 1- to characterize the pattern of AS in Portuguese patients; 2- to investigate MHC and non-MHC gene associations with susceptibility and phenotypic features of AS and; 3- to identify candidate genes associated with AS by means of whole-genome microarray. Material and Methods: AS was defined in accordance to the modified New York criteria and AS cases were recruited from hospital outcares patient clinics. Demographic and clinical data were recorded and blood samples collected. A random group of HLA-B27 positive patients and controls were selected and typed for HLA class I and II by PCR-rSSOP. The extended HLA haplotypes were estimated by Expectation Maximization Algorithm using Arlequin v3.11 software. Genotyping of IL23R, ERAP1 and ANKH allelic variants was carried out with TaqMan allelic discrimination assays. Association analysis was performed using the Cochrane-Armitage and linear regression tests as implemented in PLINK, for dichotomous and quantitative variables, respectively. Gene expression profile was carried out using Illumina HT-12 Whole-Genome Expression BeadChips and candidate genes were validated using qPCR-based TaqMan Low Density Arrays (TLDAs). Results: A total of 369 patients (62.3% male; mean age 45.4±13.2 years; mean disease duration 11.4±10.5 years), were included. Regarding clinical disease pattern, at the time of assessment, 49.9% had axial disease, 2.4% peripheral disease, 40.9% mixed disease and 7.1% isolated enthesopathic disease. Acute anterior uveitis (33.6%) was the most common extra-articular manifestation. 80.3% of AS patients were HLA-B27 positive. The haplotype A*02/B*27/Cw*02/DRB1*01/DQB1*05 seems to confer susceptibility to AS, whereas A*02/B*27/Cw*01/DRB1*08/DQB1*04 seems to provide protection in terms of disease activity, functional and radiological repercussion. Three markers (two for IL23R and one for ERAP1) showed significant single-locus disease associations. Association of these genes with AS in the Portuguese population was confirmed, whereas ANKH markers studied did not show an association with AS. No association was seen between non-MHC genes and clinical manifestations of AS. A gene expression signature for AS was established; among the fourteen validated genes, a number of them have a well-documented inflammatory role or in modulation of cartilage and bone metabolism. Conclusions: A demographic and clinical profile of patients with AS in Portugal was established. Identification of genetic variants of target genes as well as gene expression signatures could provide a better understanding of AS pathophysiology and could be useful to establish models with relevance in terms of susceptibility, prognosis, and potential therapeutic guidance.

Municipalities of higher vulnerability to Sylvatic Yellow Fever occurrence in the São Paulo State, Brazil

Until 1999 the endemic cases of Sylvatic Yellow Fever were located in the states of northern, midwestern and pre-Amazon regions. Since then, the disease progressively expanded its territory of occurrence, cases being registered beyond the traditional boundaries of endemism. The São Paulo State is considered to be part of this context, since after decades without registration of autochthonous cases of the disease, it reported, in 2000 and 2008-2009, epizootic occurrence in non-human primates and 30 cases in humans. Facts like these, added to the increase in incidences of serious adverse effects resulting from the Yellow Fever vaccination, have highlighted the importance of defining priority municipalities for vaccination against the disease in the state. Two groups of municipalities, some affected and some non-affected by YF, were compared for environmental variables related to the eco-epidemiology of the disease according to literature. The Multiple Correspondence Analysis (MCA) was used to pinpoint the factor able to differentiate the two groups of municipalities and define the levels of risk. The southeast region of the São Paulo State was considered to be the area with a higher number of municipalities classified as high risk and should be considered a priority for the application of prevention measures against Yellow Fever.

SEASONAL DISTRIBUTION OF MALARIA VECTORS (DIPTERA: CULICIDAE) IN RURAL LOCALITIES OF PORTO VELHO, RONDÔNIA, BRAZILIAN AMAZON

We conducted a survey of the malaria vectors in an area where a power line had been constructed, between the municipalities of Porto Velho and Rio Branco, in the states of Rondônia and Acre, respectively. The present paper relates to the results of the survey of Anopheles fauna conducted in the state of Rondônia. Mosquito field collections were performed in six villages along the federal highway BR 364 in the municipality of Porto Velho, namely Porto Velho, Jaci Paraná, Mutum Paraná, Vila Abunã, Vista Alegre do Abunã, and Extrema. Mosquito captures were performed at three distinct sites in each locality during the months of February, July, and October 2011 using a protected human-landing catch method; outdoor and indoor captures were conducted simultaneously at each site for six hours. In the six sampled areas, we captured 2,185 mosquitoes belonging to seven Anopheles species. Of these specimens, 95.1% consisted of Anopheles darlingi, 1.8% An. triannulatus l.s., 1.7% An. deaneorum, 0.8% An. konderi l.s., 0.4 An. braziliensis, 0.1% An. albitarsis l.s., and 0.1% An. benarrochi. An. darlingi was the only species found in all localities; the remaining species occurred in sites with specific characteristics.

Regulation of PLK4 levels and activity to ensure centriole number control

Dissertation presented to obtain a Ph.D degree in Cellular Biology