Repositório RCAAP

O presente trabalho tem como objectivo estudar as acaracterísticas psicométricas do University Student Depression Inventory (USDI), tendo em vista a adaptaçãod este instrumento à população universitária portuguesa.

The main objective of this thesis was to develop a tool for Mental Workload (MWL) assessment, trying to answer to the growing interest of the community in evaluating the importance of this factor in task performance and human-machine interaction. The relevance of MWL in these areas has been the focus of an increasing number of scientific studies and publications despite the concept or definition of MWL in itself still fuzzy and the focus of many discussions. Furthermore, the objective assessment of this factor is still ill-defined, resourcing to many different methodologies and tools. Being a multivariable concept, Mental Workload should be assessed in a multimodal way. We used subjective methods including NASA task-load index; performance methods such as reaction time to make a decision; and objective methods including EEG device for brain related metrics, PPG for heart metrics and an eye-tracker for eye related metrics. In the case of the objective methods, the main issue for any sensor is the preprocessing stage, taking into account different type of noise and assuring a good quality data for reliable results. In order to contribute to the study of MWL and of its impact in task performance, we developed a tool based on the following assumptions: • Use of a non-medical-grade equipment in assessing mental workload induced by visual tasks; • Increase the MWL classification efficiency by correlation of different physiological measures, with different sensors; • Provide a common platform for sensor integration and signal processing, based on off-the-shelf, open source tools existing in the market. In this thesis, we showed how a multimodal system for MWL assessment can be made without resourcing to high cost / high complexity equipment, therefore enabling easier access for people interested in working in this field of study.

Automatic argument mining seeks to endow artificial devices with the ability to identify structured arguments from plain unstructured text in natural language. A major motivation for empowering artificial devices with such skill is the fundamental role of argumentation in human cognition, by allowing the transmission and acquisition of knowledge, a hallmark of human interactions. While inheriting the complexities of defining a structure for arguments that are faced by argumentation theory, in general, argument mining also addresses challenges that are specific to the automatic extraction of structured arguments, as for example, the need for a representation of natural language semantics for an argument to be handled computationally. Current natural language processing techniques have successfully captured, to some extent, the meaning of words and sentences with distributional semantic representations and different neural network architectures. The performance of these techniques has improved with ever larger volumes of data, particularly annotated data. However, annotating data is time-consuming and laborious, and several natural language processing tasks have an inherent scarcity of annotated data, including argument mining. In this dissertation, we explored several neural network solutions to argument mining inspired by distributional semantics and transfer learning to mitigate the scarcity of annotated data in argument mining and improve its performance. We make contributions regarding several argument mining sub-tasks, namely in the so-called argument component identification, clausal classification and relational classification. These solutions experiment and resort to cross-lingual transfer from the most resourced language, English, to a low-resourced language, Portuguese, as also diverse knowledge bases from different lexical semantic families and over forty confluent language processing tasks. Additionally, we assess the impact of data artifacts in annotated data through a reproduction study, followed by a study of Transformer-based architectures on a most demanding argument mining sub-task concerned with argument reasoning comprehension. Throughout these explorations into neural argument mining, we set new state-of-the-art results across several argument mining sub-tasks, providing empirical evidence that our proposed solutions outperform previous computational models with alternative approaches.

Estuaries are systems of ecological importance that also suffer from several anthropogenic pressures. Phytoplankton is considered an important water quality indicator due to its rapid response to variations in the environment. However, to adequately assess phytoplankton in estuaries it is important to understand how phytoplankton reacts to estuarine variability. Thus, the main aim of the present doctoral thesis is to improve the knowledge of the effects of temporal and spatial variations of the Tagus Estuary environment over the water quality metrics and to predict how it will respond in the future, under climate change scenarios. Thus, this study has the following objectives: i) understand the tidal effect over the estuarine physicochemical conditions and phytoplankton community; ii) assess the tidal effect over the water quality metrics; iii) understand the spatial and seasonal variability of physicochemical conditions and phytoplankton in the Tagus Estuary, as well as the effect of such variations over the Water Framework Directive (WFD) water quality metrics and iv) simulate the effects of increased outfall discharges and climate change over the Tagus Estuary physicochemical parameters and phytoplankton community and its effects on the WFD metrics. The results demonstrate that chlorophyll-a concentrations were higher during spring tides along with higher particulate matter concentrations. The low-neap tide was the most appropriate tidal condition to observe the worst-case scenario, mainly in the location more influenced by outfalls. In general, chlorophyll-a concentrations were higher during the Summer and Spring accompanied with higher suspended matter concentrations while nutrients concentrations were higher during the winter. Both, the simulations of sea level rise and reduction of river discharge led to a decrease of chlorophyll-a and nutrients concentrations while increasing salinity. In conclusion, the phytoplankton community of the Tagus Estuary is influenced by both temporal and spatial drivers, with a decrease in nutrients and chlorophyll-a concentrations expected in the future.

Extreme Value Theory and Statistics constitute the ideal toolbox to handle rare and extreme events, for which inference through classical Gaussian­based methodologies is unreliable. In this thesis, we start by reviewing the basis of Statistics of Extremes and statistical moments, then diving into open topics from a methodological standpoint, supported by simulation studies of the proposed contributions’ properties, together with illustrative analysis of data from several applied fields. Three new methods for appropriate threshold selection are suggested, developed under the Gen eralized Pareto and peaks­over­threshold parametric framework for univariate, independent and identically distributed data. The proposed Automatic L­moment Ratio Selection Method stands out as the superior technique, being automatic, objective and computationally effective while presenting overall good performance compared to state­of­the­art alternatives. Data sets of significant wave heights are considered, and results compared to previously existing studies of this data. Further, an analysis of very small data sets of measurements of Giant Squids is conducted with the aim of estimating the species’ maximum possible size – a novelty Extremes­driven take on this problem from the field of Zoology. This work moreover describes how the known semi­parametric mixed moment estimator for the Extreme Value Index may be used in the context of multivariate data, pooled across space and time. The setup considers the presence of extreme spatial dependence and a possibly non­monotonic trend in the frequency of extremes through both space and/or time (heteroscedasticity). The asymptotic properties of the estimator are derived under this more complex scenario, and its application is illustrated through the study of reanalysis daily precipitation series recorded for a set of locations in homogeneous regions of the United Kingdom. The estimators performance is compared with that of the common benchmark maximum likelihood estimator under the same conditions, and found to be generally favorable.

O Mieloma Múltiplo (MM) constitui a segunda patologia hemato-oncológica mais comum. É uma patologia complexa e heterogénea, caracterizada por recaídas recorrentes e resistência à terapêutica. A sua patogénese envolve a interação dos plasmócitos malignos com o seu microambiente na medula óssea, envolvendo mecanismos imunológicos. Clinicamente, evolui de um estado pré-maligno assintomático para mieloma ou outra doença maligna relacionada, cuja progressão é influenciada pela citogenética subjacente. A emergência de novas terapêuticas ao longo das últimas décadas transformou o panorama no MM, com uma melhoria significativa dos resultados clínicos e reportados pelo doente. O advento de terapêuticas dirigidas para mecanismos biopatológicos específicos modificou o paradigma terapêutico, com impacto nas taxas de resposta e sobrevivência. No que concerne ao Mieloma Múltiplo em Recaída ou Refratário, nos últimos anos têm sido desenvolvidas terapêuticas emergentes que podem constituir uma alternativa. O antigénio de superfície B-Cell Maturation Antigen (BCMA) é um alvo específico, cuja expressão é particularmente pronunciada nos plasmócitos malignos. A imunoterapia direcionada para o antigénio BCMA tem demonstrado resultados clínicos promissores em doentes refratários às terapêuticas convencionais, independentemente da heterogeneidade e risco genéticos. No entanto, foram descritos resultados desfavoráveis quanto à toxicidade, ausência de efetividade e mecanismos de resistência. A imunoterapia anti-BCMA inclui conjugados anticorpo-fármaco (ADC), anticorpos biespecíficos (BiTES) e terapias celulares autólogas (células CAR T). Os conjugados anticorpo-fármaco são compostos por anticorpos monoclonais ligados covalentemente a fármacos citotóxicos, envolvidos na lesão e morte celular. Os anticorpos biespecíficos contém um fragmento de cadeia única variável associado a dois anticorpos monoclonais que se ligam concomitantemente a BCMA e ao antigénio de superfície CD3 das células T, desencadeando a ligação cruzada de células T e células do MM, ativação das células T CD4+/CD8+ e secreção de citocinas citotóxicas, com subsequente morte celular. As células CAR T conectam-se a BCMA, com consequente ativação, secreção de citocinas citotóxicas e morte celular. As terapêuticas emergentes direcionadas para BCMA demonstraram resultados encorajadores no Mieloma Múltiplo em Recaída ou Refratário, em particular as terapias celulares. Todavia, permanecem alguns desafios a considerar, como a efetividade e segurança a longo prazo, a sequência no algoritmo terapêutico e a ocorrência de resistências à terapêutica.

Cheese is a type of food that has been used over time as a way of preserving milk. It is a very consumed food due to its flavor and versatility. For some years, popular cases of color defects in cheeses surfaces have been reported in the literature. Sporadic inconsistencies in cheese appearance may result in a downgrading of cheese and a consequential economic loss to producers. The dairy industry may be a good source of bacteria strains producing natural pigments. Pigments are compounds of importance to various industries, for example the food industry, where they can be used as additives, colour enhancers and antioxidants. As the current trend around the world is shifting towards the use of environmentally friendly products, the demand for natural dyes is increasing. Melanins are pigments produced by various microorganisms. Pseudomonas putida ESACB 191, isolated from goat cheese rind, has been described as a brown pigment producer. This strain produces a brown pigment in synthetic Müeller-Hinton broth. This brown compound was extracted, purified, analysed by FTIR and mass spectrometry, and identified as eumelanin. The maximum volumetric productivity obtained was 1.57 mg/L/h. The bioactivity of eumelanin was evaluated as the free radical scavenging capacity (antioxidant activity), EC50 of 74.0 ± 0.2 µg/mL, when determined using DPPH test and as an acetylcholinesterase inhibitor, with IC50 575 ± 4 µg/mL. This bacterial eumelanin showed no cytotoxicity towards A375, HeLa Kyoto, HepG2 or Caco2 cell lines. The effect of melanin on cholesterol absorption and drug interaction was evaluated in order to understand the interaction of melanin present in the cheese rind when ingested by consumers. However, it had no effect on cholesterol absorption across a simulated intestinal barrier formed by the Caco2 cell line or with the drug ezetimibe. Cynara cardunculus L. inflorescence infusion has been used for several centuries as coagulating agent to obtain curd in traditional cheese making, in some highly prized Portuguese cheeses. To promote the sustainable use of all C. cardunculus plants, C. cardunculus extract leaves decoction (CL), inflorescence decoction (CI), chlorogenic acid (CA) (a compound present in the plant leaves), and rosmarinic acid (RA) (a similar phenolic compound) aqueous solutions were tested for antimicrobial activity against bacteria that may appear on the cheese rind. The antimicrobial activity was evaluated using 15 bacterial strains utilizing two different methodologies: solid and liquid. The influence of these extracts and the phenolic compounds on melanin bioproduction by Pseudomonas putida ESACB 191 was also studied. CA and RA (1 mg/mL) showed antimicrobial activity. CL and CA reduced P. putida ESACB 191 growths in the liquid assay and melanin bioproduction by 6.20 Log CFU/mL and 50%, respectively. Cynarin, CA, and its derivates were identified as the main phenolic compounds (52%) of CL, which may justify its inhibitory action on bacterial growth and melanin bioproduction. The cheese rind is the natural active packaging of cheese and is subject to a wide range of external factors that compromise the appearance of the cheese, namely, color defects caused by spoilage microorganisms. To avoid the formation of brown rind due to microorganism spoilage eight films based on whey protein isolate (WPI) coatings were studied, of which IS3CA (WPI 5%+sorbitol 3%+citric acid 3%) was selected due to better properties. From the IS3CA film, novel films containing melanin M1 (74 µg/mL) and M2 (500 µg/mL) were developed and applied to cheese under proof-of-concept in industrial conditions. After 40 days of maturation, M2 presented the lowest microorganism count for all the microbial parameters analyzed. The cheese with M2 showed the lowest lightness, which indicates that it is the darkest cheese due to the melanin concentration. It was found that the mechanical and colorimetric properties are the ones that contribute the most to the distinction of the M2 film in cheese from the others. Using FTIR-ATR, it was possible to distinguish the rinds of M2 cheeses because they contained the highest concentrations of melanin. Thus, this study shows that the film with M2 showed the best mechanical, chemical, and antimicrobial properties for application in cheese. The novel melanin edible films also give the cheese a brown, uniform coating that contributes to improving the appearance of the cheese making it more attractive to the consumer.

Molecular regulation studies of plant embryogenesis have been conducted mostly in angiosperms. Nevertheless, it is estimated that angiosperm and gymnosperm lineages have diverged at least 300 million years ago, and their different embryo development features are well described. The microRNAs (miRNAs) are a class of small non-coding RNAs (sncRNAs) that regulate gene expression mostly post-transcriptionally by leading the cleavage or translational repression of their target transcripts, thereby acting as major regulators during plant development. A set of candidate miRNAs and corresponding mRNA transcripts have emerged as potential embryogenesis regulators based on a previous profiling of the Pinus pinaster sncRNA transcriptome in several embryogenic tissues. By focusing on the differential expression of some miRNAs throughout embryo development, a few P. pinaster miRNA-mRNA pairs were selected for further characterization. Such pairs included miR160-PpARF18, miR162-PpDCL1 and miR166-PpC3HDZ, potentially involved in auxin responses and miRNA biogenesis, among other processes. As a first characterization step, a transient expression system in Arabidopsis mesophyll protoplasts based on the activity of reporters which are either sensitive or resistant to miRNA cleavage, allowed the in vivo validation of the physical interaction between miR160 and the PpARF18 target site. Then, the miRNA-mRNA expression patterns across embryo development were studied using Picea abies somatic embryogenesis as a conifer model system, revealing expression trends similar to those previously reported for Arabidopsis. Finally, the role of miR160 under disturbed auxin homeostasis during P. abies embryo development was further explored. The expression profiles of the miR160:PabARF18 pair under such conditions suggest a central function of miR160 in the maintenance of adequate levels of PabARF18, possibly acting as a safeguard mechanism during embryogenesis. Overall, this work reports the first in vivo validation of a gymnosperm miRNA with its predicted target, supporting the conservation of the interaction between miR160 and ARF transcripts in conifers, and emphasising the relevant miRNA regulatory functions during embryogenesis. These results provide an important basis for further studies contributing to detail the molecular roadmap of embryo development in conifers.

No summary/description provided

O tema desta dissertação é a relação entre biopolítica e ciências da vida em Foucault, cujo corpus tem estimulado uma considerável produção de comentário, auxiliando, assim, a pensar o presente (pós-)pandémico. O tema permite, nesse sentido, ver algumas dificuldades importantes que afectam Foucault e o actual debate sobre aquela relação: 1) A relação entre um governo e uma ciência que se focam na vida acontece por via ideológica, isto é, enganadora, o que contraria, pois, a tese foucaultiana sobre a “produtividade” da relação entre o poder e o saber; 2) O contributo das ciências da vida para uma biopolítica de promoção da vida parece se realizar apenas pela morte. Por exemplo, expondo os mais vulneráveis à morte para melhorar a saúde da população, ou colocando a população à mercê de vírus potentes criados em laboratório. Pergunta-se, então: existe uma relação produtiva, ou seja, nem ideológica nem mortífera, entre biopolítica e ciências da vida em Foucault? A nosso ver, o modo como Foucault lê os cientistas da vida que lhe são contemporâneos permite explicar o seu abandono dos temas da ideologia e da morte para pensar as ciências da vida, isto é, quando se dedica à biopolítica da “segurança” no curso dado no Collège de France em 1977- 1978, Sécurité, Territoire, Population. A nossa hipótese é que uma relação produtiva entre biopolítica e ciências da vida passa pela gestão da relação entre regularidade-irregularidade da vida. Quer dizer, uma biopolítica que estimula, com o auxílio das ciências da vida, o inesperado da vida, de modo a se tornar um poder dinâmico, que aprende, pois, com o novo, abrindo-se ao futuro para o melhor capturar. Finaliza-se com um estudo de caso para pensar uma biopolítica da comunicação de ciência à luz do tema da relação entre regularidade-irregularidade.

Virulence is defined as the degree of disease severity and lethality produced by a pathogen and is conditioned by virulence factors, which vary depending on the pathogen and determine the range of hosts it can infect. As a result, virulence is a complex phenotype, dependent on multiple molecular determinants. Omic techniques are thus a good way to investigate it. This thesis discusses two virulence-related phenotypes in two pathogens: surfactant biosynthesis in Pseudomonas aeruginosa and host specificity in Mycobacterium tuberculosis. P. aeruginosa is an opportunistic gram-negative pathogen that infects immunocompromised patients. Its collective behaviors are crucial during infection. They require the secretion of biosurfactants, secondary metabolites that enable the bacteria to spread through moist surfaces, which is highly regulated by environmental cues. Conversely, M. tuberculosis is a mammal obligate pathogen whose first infection targets are the macrophages. There are several M. tuberculosis lineages, which present host association. Identifying the factors that affect the inclination towards a host can show host-specific virulence patterns. Here, through a multi-omic approach we gained insight about these two phenotypes. We observed that surfactant production is inconsistent across P. aeruginosa’s phylogeny and added an additional layer of regulation of this phenotype, dependent on oxidative stress. Regarding M. tuberculosis, through metabolic modeling we predicted the lineage-specific genomic differences to produce host-correlated convergent alterations in metabolic pathways important for its virulence. Furthermore, we pinpointed candidate genes for their involvement in host specificity, such as Phospholipase C ones. Finally, by using an in vitro infection model we expanded the current knowledge about the adaptation of animal-associated lineages to their hosts and found experimental evidence supporting the involvement of the phospholipase C plcC gene in the adaptation to humans. Metabolism was central in the exploration of these phenotypes. Because it is at the end of the biological information chain, it is closer to phenotype than any other omic dataset. Metabolomics gave a comprehensive picture of the researched biological systems, allowing for the formulation of hypotheses that could later be empirically tested. The bioinformatic pipelines developed in this thesis can be applied to the study of other complex phenotypes in other organisms. The knowledge gained about these virulence related phenotypes might serve as a basis for developing new therapies against these pathogens.

The degree of erythrocyte deformability (ED) changes as a consequence of alterations on either its membrane properties, or shape or internal composition may provide information about erythrocyte functional disturbances. Several metabolic, cerebral and cardiovascular diseases are characterized by decreased ED that may compromise the blood flow in microcirculation. “In vitro” effects of compounds on ED under different shear stresses may be studied using an ektacytometer apparatus which give us the erythrocyte elongation index (EEI) as a degree of the ED ability. When phosphorylation level of erythrocyte band 3 protein was modified by specific PTK and PTP inhibitors, no ED changes were observed. Cytoskeleton protein phosphorlation levels modifications obtained with PKC inhibitor or activator decreased and increased EEI respectively. When the thiol status of the erythrocyte were preserved with dithiothreitol (DTT), no alterations on EEI were verified. ED values increased for all shear stresses when in presence of the acetylcholine at variance when submitted to the Spermine NONOate that only at high shear stresses an EEI increase was observed. Better understanding of those molecules on erythrocyte signal transduction mechanisms may bring potential targets for drug intervention on ED-dependent blood rheology.

DIS3L2, member of the highly conserved RNase II family, is a 3’-5’ exoribonuclease that degrades different types of RNAs in an exosome-independent manner, including mRNAs and several types of non-coding RNAs. DIS3L2-mediated decay operates in tight cooperation with the terminal uridylyl transferases 4 and 7 (TUT4/7), which uridylate specific mRNAs that are subsequently recognized and degraded by DIS3L2. Recently, it was discovered that these three factors are involved in the nonsense-mediated mRNA decay (NMD) pathway. NMD is a surveillance pathway that selectively degrades mRNAs harboring premature translation-termination codons (PTCs) located more than 50-55 nucleotides upstream of an exon-exon junction, thus protecting the cell from the expression of truncated proteins. Furthermore, NMD controls the expression levels of normal and fully functional mRNAs, becoming an important mechanism of gene expression regulation in the cell. However, the mechanism by which DIS3L2 and uridylation take place in the NMD pathway remains poorly characterized. On the other hand, there is some evidence that dysregulated activity of DIS3L2 may contribute to cancer development. This ribonuclease has been implicated in cancer-related cellular processes such as cell proliferation and apoptosis, and mutations in DIS3L2 gene lead to deregulation of cell cycle genes inducing chromosomal instability. In this PhD project, we aimed to characterize the role of DIS3L2 in cancer and to dissect the mechanism by which DIS3L2 and uridylation are involved in the NMD pathway. By analyzing publicly available RNA datasets from The Cancer Genome Atlas (TCGA), we found higher DIS3L2 mRNA levels in colorectal cancer (CRC) tissues versus normal colonic samples as well as worse prognosis for CRC patients with high DIS3L2 expression. Moreover, our RNA deep-sequencing analysis showed that knockdown (KD) of DIS3L2 results in a significant transcriptomic disturbance in SW480 CRC cells. In addition, gene ontology (GO) analysis of significant upregulated genes exhibited enrichment in mRNAs encoding proteins involved in cell cycle regulation and cancer-related pathways, which guided us to investigate which specific hallmarks of cancer are differentially regulated by DIS3L2. Four CRC cell lines (HCT116, SW480, Caco-2 and HT-29) with distinct mutational background and oncogenicity were used to test the role of DIS3L2 in this cancer type. Our results indicate that lack of DIS3L2 impairs cell viability of highly oncogenic SW480 and HCT116 CRC cells, but has little or no impact in the more differentiated Caco-2 and HT 29 cells. Interestingly, the mTOR signaling pathway, crucial for cell survival and growth, is downregulated upon DIS3L2 depletion, whereas AZGP1, an mTOR pathway inhibitor, is upregulated. Furthermore, we demonstrate that DIS3L2 KD leads to the disruption of metastasis-associated properties, such as cell migration and invasion, only in highly oncogenic CRC cells. Our bioinformatic analysis corroborate the predictive value of specific NMD-eliciting genomic features characterized in previous studies, such as a high GC content in the 3’UTR, the presence of 3’UTR introns or a higher incidence of uORFs. Interestingly, the 3’UTRs of NMD-targeted mRNAs degraded by DIS3L2 tend to have a high GC content and to be devoid of 3’UTR introns when compared to 3’UTRs of transcripts showing resistance to NMD and DIS3L2-mediated degradation. Notably, gene expression assessment in a system of constructs containing different genomic regions of the NMD/DIS3L2 target GADD45A unveiled that its 3’UTR enhances susceptibility to NMD and contributes for DIS3L2 target recognition. Furthermore, we discovered that interruption of the mRNA closed-loop configuration does not sensitize NMD targets to degradation by DIS3L2. Curiously, we found that DIS3L2-mediated degradation seems to be more efficient in circularized NMD targets. Throughout this work, several results have shown that lack of uridylation along with DIS3L2 mitigates the impact observed upon single depletion of this ribonuclease. In an attempt to identify additional factors contributing to this phenomenon, we demonstrate that neither of the canonical cytoplasmic exoribonucleases DIS3, DIS3L1 nor XRN1 is responsible for such compensatory mechanism. Altogether, the work here presented unveils a novel oncogenic role for DIS3L2 in sustaining CRC cell proliferation and provides evidence that this ribonuclease is required to support the viability and invasive behavior of dedifferentiated CRC cells. Moreover, this work shed light on the mechanism by which DIS3L2 is involved in NMD, highlighting the biological relevance of the 3’UTR on DIS3L2 target recognition. Taking into account that the NMD pathway regulates important biological processes in the pathophysiology of cancer, understanding the mechanism by which DIS3L2 regulates specific NMD targets may provide vital information for the development of new therapeutic strategies.

Grapevine (Vitis vinifera L.) is a fruit crop with high economic value due to its final products like wine. However, every year, the vineyards are affected by the attack of different pathogens, leading to a great loss of production. One of the most common grapevine diseases is downy mildew caused by the obligate biotrophic oomycete Plasmopara viticola ((Berk. and Curt.) Berl. & de Toni). Downy mildew is a cryptogamic disease that attacks all the green parts of the plant, leaves, tendrils, branches and bunches, whose development is favoured by mild temperatures and high humidity. In this infection process, the apoplast is the subcellular compartment where there is the greatest contact between the pathogen's infection structures, as well as the molecules secreted by it, and the plant's molecules. A comprehensive knowledge on the grapevine strategies to overcome pathogens, as well as the evolution of pathogen infection mechanisms is paramount for the development of new sustainable disease control strategies and avoid the massive use of pesticides in each grapevine growing season. The apoplast is the first hub of plant-pathogen communication where pathogen effectors are recognized by plant defensive proteins and cell receptors, thus activating signal transduction pathways. As a result of this first contact, the host triggers a defence response that involves the modulation of extra and intracellular proteins, including proteases. On the side of the pathogens, they secrete a myriad of molecules, called effectors, in the plant apoplast that have the function of overcoming the plant's barriers and inducing the disease. Within proteases, subtilases have been the focus of several studies and seems to be involved in plant defence, resistance, and immunity mechanisms. In grapevine, some of them present a constitutive expression, without the plant being under the influence of any stress, quite high in some resistant cultivars, compared to a susceptible cultivar. After inoculation with P. viticola, an expression increase of several subtilases was observed in grapevine leaves. A specific subtilase, VviSBT4.19, showed interesting results that indicate a possible involvement in grapevine resistance to this oomycete. Considering all the above, the aims of this work were: 1) The development of a methodology for extracting the apoplast from grapevine leaves, focusing on two grapevine cultivars with different degrees of tolerance to downy mildew; 2) The study of the apoplast proteome during grapevine-downy mildew interaction, identifying both plant and pathogen proteins; 3) The analysis of VviSBT4.19 gene expression in grapevine leaves infected with other pathogens, namely the fungi Erysiphe necator and Botrytis cinerea, and through the identification of interacting proteins with the VviSBT4.19 during 'Regent'-P. viticola interaction. It is expected that this work will contribute in an active and relevant way to the deep knowledge of the interaction between the grapevine and the downy mildew for the development or improvement of sustainable alternatives for a more environmentally friendly viticulture.

No summary/description provided

Anthropogenic activities are increasingly shaping ecosystems worldwide, threatening the functioning of global ecological processes that maintain life on Earth. Endemic-rich areas, such as oceanic islands, hold unique biotas and socio-ecological systems, which are particularly susceptible to habitat degradation and biological invasions. Terrestrial molluscs comprise a highly diverse taxonomic group which support critical ecosystem functions but have suffered some of the highest extinction rates. The oceanic islands of the Gulf of Guinea host a high proportion of single-island endemic terrestrial mollusc species that remain little studied. This thesis explores the links between terrestrial molluscs, their habitats, and the people living in the Gulf of Guinea oceanic islands, using a combination of techniques from social and ecological sciences. The islands hold 96 species of terrestrial mollusc, 65% of which are endemic. On the largest island, São Tomé, land-use intensification decreased native species richness and abundance, while it promoted introduced species, such as the invasive Archachatina marginata. This species was an important source of food and income, especially for vulnerable people in rural areas, where it was also widely preferred and recognized by children. Wildlife conservation preferences were mainly justified by species attractiveness or profitability, and male, poorer children from rural areas were more likely to correctly identify animal species. The endemic A. bicarinata is the only terrestrial mollusc from the islands that has been listed as threatened, and most of its genetic diversity was found on Príncipe Island. This thesis showed that habitat loss and introduced species are important threats to this malacofauna, and that maintaining the integrity of forest ecosystems is critical to preventing invasions, but also that management for conservation will have to consider the relevance of introduced species for people. Urgent in and ex situ efforts are needed to protect the valuable forests where most endemic molluscs persist.

O presente artigo apresenta e discute os dados arqueológicos referentes aos mais antigos vestígios de uso de argila e de cerâmica no atual concelho de Mafra, partindo da informação compulsada nos últimos 25 anos, através de ações desenvolvidas pelas signatárias, no âmbito das intervenções promovidas pela Câmara Municipal de Mafra. Discute-se e apresentam-se os dados disponíveis para os sítios de Cova da Baleia (2007), São Julião (1999, 2000, 2007) e Lizandro (2018) e referem-se outros achados superficiais, numa leitura cruzada com o panorama disponível para o Ocidente Peninsular.

Recentemente foram identificados monumentos funerários romanos em pedra que vêm enriquecer o património do concelho de Mafra. Pertencem a uma tipologia centrada no século I d. C. denominada cupa, palavra que designa uma cobertura tumular em forma semicilíndrica. Estes monumentos ainda mantêm em muitos casos os seus epitáfios, inscrições epigráficas que nos fornecem informações sobre as pessoas que neles foram sepultadas, fontes inestimáveis para a reconstituição do passado romano deste extremo ocidental do Império. No presente estudo abordamos o conjunto dos monumentos em forma de cupa até agora descobertos no que é, hoje em dia, território mafrense.

No summary/description provided

An effective vaccine that will provide sterile protection against Plasmodium, the etiologic agent of malaria, is crucial for disease eradication. Efforts in the last five decades culminated in a WHOendorsed malaria vaccine. Albeit significantly reducing severe disease cases, this vaccine consistently fails at conferring sterile immunity to vaccinees. The high protection attained following experimental immunization with attenuated parasites halting their development soon after hepatocyte infection raises the possibility that early events during pre-erythrocytic infection are important for protective immunity against Plasmodium. Prior to the productive infection of an hepatocyte, Plasmodium parasites journey across biological barriers by traversing several host cells first in the skin, and subsequently in the liver sinosoids and parenchyma. By employing the cell traversal-deficient parasite lines Pbspect1- and Pbplp1- , we show that cell traversal is crucial for the establishment of sterile protection upon radiation-attenuated sporozoitesbased immunization of C57BL/6J mice. Furthermore, we show that loss of protection in the absence of cell traversal is independent of the timing of intrahepatic development arrest of parasites, as CPS-based immunizations with Pbspect1- parasites equaly failed to confer full protection upon infectious challenge. Analysis of the immune mechanisms relevant for sterile protection revealed that, while humoral resposes were not impacted by the lack of cell traversal, the establishment of CD8+ T cell responses following vaccination was impaired in Pbspect1- RAS-immunized mice. Together, our data suggest that cell traversal by Plasmodium parasites is a critical determinant for the establishment of sterile protection following vaccination by eliciting and/or maintaining cellular immunity aimed at eliminating Plasmodium intrahepatic forms. These findings challenge the current understanding that malaria vaccination largely depends on parasite development, and raise the possibility that enhancing cell traversal motility of immunizing parasites may constitute a valid strategy to boost malaria vaccine candidates efficacy and take us one step closer to a malaria-free world.