16,685 research outputs found

    Pollution-induced community tolerance in freshwater biofilms – from molecular mechanisms to loss of community functions

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    Exposure to herbicides poses a threat to aquatic biofilms by affecting their community structure, physiology and function. These changes render biofilms to become more tolerant, but on the downside community tolerance has ecologic costs. A concept that addresses induced community tolerance to a pollutant (PICT) was introduced by Blanck and Wängberg (1988). The basic principle of the concept is that microbial communities undergo pollution-induced succession when exposed to a pollutant over a long period of time, which changes communities structurally and functionally and enhancing tolerance to the pollutant exposure. However, the mechanisms of tolerance and the ecologic consequences were hardly studied up to date. This thesis addresses the structural and functional changes in biofilm communities and applies modern molecular methods to unravel molecular tolerance mechanisms. Two different freshwater biofilm communities were cultivated for a period of five weeks, with one of the communities being contaminated with 4 μg L-1 diuron. Subsequently, the communities were characterized for structural and functional differences, especially focusing on their crucial role of photosynthesis. The community structure of the autotrophs was assessed using HPLC-based pigment analysis and their functional alterations were investigated using Imaging-PAM fluorometry to study photosynthesis and community oxygen profiling to determine net primary production. Then, the molecular fingerprints of the communities were measured with meta-transcriptomics (RNA-Seq) and GC-based community metabolomics approaches and analyzed with respect to changes in their molecular functions. The communities were acute exposed to diuron for one hour in a dose-response design, to reveal a potential PICT and uncover related adaptation to diuron exposure. The combination of apical and molecular methods in a dose-response design enabled the linkage of functional effects of diuron exposure and underlying molecular mechanisms based on a sensitivity analysis. Chronic exposure to diuron impaired freshwater biofilms in their biomass accrual. The contaminated communities particularly lost autotrophic biomass, reflected by the decrease in specific chlorophyll a content. This loss was associated with a change in the molecular fingerprint of the communities, which substantiates structural and physiological changes. The decline in autotrophic biomass could be due to a primary loss of sensitive autotrophic organisms caused by the selection of better adapted species in the course of chronic exposure. Related to this hypothesis, an increase in diuron tolerance has been detected in the contaminated communities and molecular mechanisms facilitating tolerance have been found. It was shown that genes of the photosystem, reductive-pentose phosphate cycle and arginine metabolism were differentially expressed among the communities and that an increased amount of potential antioxidant degradation products was found in the contaminated communities. This led to the hypothesis that contaminated communities may have adapted to oxidative stress, making them less sensitive to diuron exposure. Moreover, the photosynthetic light harvesting complex was altered and the photoprotective xanthophyll cycle was increased in the contaminated communities. Despite these adaptation strategies, the loss of autotrophic biomass has been shown to impair primary production. This impairment persisted even under repeated short-term exposure, so that the tolerance mechanisms cannot safeguard primary production as a key function in aquatic systems.:1. The effect of chemicals on organisms and their functions .............................. 1 1.1 Welcome to the anthropocene .......................................................................... 1 1.2 From cellular stress responses to ecosystem resilience ................................... 3 1.2.1 The individual pursuit for homeostasis ....................................................... 3 1.2.2 Stability from diversity ................................................................................. 5 1.3 Community ecotoxicology - a step forward in monitoring the effects of chemical pollution? ................................................................................................................. 6 1.4 Functional ecotoxicological assessment of microbial communities ................... 9 1.5 Molecular tools – the key to a mechanistic understanding of stressor effects from a functional perspective in microbial communities? ...................................... 12 2. Aims and Hypothesis ......................................................................................... 14 2.1 Research question .......................................................................................... 14 2.2 Hypothesis and outline .................................................................................... 15 2.3 Experimental approach & concept .................................................................. 16 2.3.1 Aquatic freshwater biofilms as model community ..................................... 16 2.3.2 Diuron as model herbicide ........................................................................ 17 2.3.3 Experimental design ................................................................................. 18 3. Structural and physiological changes in microbial communities after chronic exposure - PICT and altered functional capacity ................................................. 21 3.1 Introduction ..................................................................................................... 21 3.2 Methods .......................................................................................................... 23 3.2.1 Biofilm cultivation ...................................................................................... 23 3.2.2 Dry weight and autotrophic index ............................................................. 23 3.2.4 Pigment analysis of periphyton ................................................................. 23 3.2.4.1 In-vivo pigment analysis for community characterization ....................... 24 3.2.4.2 In-vivo pigment analysis based on Imaging-PAM fluorometry ............... 24 3.2.4.3 In-vivo pigment fluorescence for tolerance detection ............................. 26 3.2.4.4 Ex-vivo pigment analysis by high-pressure liquid-chromatography ....... 27 3.2.5 Community oxygen metabolism measurements ....................................... 28 3.3 Results and discussion ................................................................................... 29 3.3.1 Comparison of the structural community parameters ............................... 29 3.3.2 Photosynthetic activity and primary production of the communities after selection phase ................................................................................................. 33 3.3.3 Acquisition of photosynthetic tolerance .................................................... 34 3.3.4 Primary production at exposure conditions ............................................... 36 3.3.5 Tolerance detection in primary production ................................................ 37 3.4 Summary and Conclusion ........................................................................... 40 4. Community gene expression analysis by meta-transcriptomics ................... 41 4.1 Introduction to meta-transcriptomics ............................................................... 41 4.2. Methods ......................................................................................................... 43 4.2.1 Sampling and RNA extraction................................................................... 43 4.2.2 RNA sequencing analysis ......................................................................... 44 4.2.3 Data assembly and processing................................................................. 45 4.2.4 Prioritization of contigs and annotation ..................................................... 47 4.2.5 Sensitivity analysis of biological processes .............................................. 48 4.3 Results and discussion ................................................................................... 48 4.3.1 Characterization of the meta-transcriptomic fingerprints .......................... 49 4.3.2 Insights into community stress response mechanisms using trend analysis (DRomic’s) ......................................................................................................... 51 4.3.3 Response pattern in the isoform PS genes .............................................. 63 4.5 Summary and conclusion ................................................................................ 65 5. Community metabolome analysis ..................................................................... 66 5.1 Introduction to community metabolomics ........................................................ 66 5.2 Methods .......................................................................................................... 68 5.2.1 Sampling, metabolite extraction and derivatisation................................... 68 5.2.2 GC-TOF-MS analysis ............................................................................... 69 5.2.3 Data processing and statistical analysis ................................................... 69 5.3 Results and discussion ................................................................................... 70 5.3.1 Characterization of the metabolic fingerprints .......................................... 70 5.3.2 Difference in the metabolic fingerprints .................................................... 71 5.3.3 Differential metabolic responses of the communities to short-term exposure of diuron ............................................................................................................ 73 5.4 Summary and conclusion ................................................................................ 78 6. Synthesis ............................................................................................................. 79 6.1 Approaches and challenges for linking molecular data to functional measurements ...................................................................................................... 79 6.2 Methods .......................................................................................................... 83 6.2.1 Summary on the data ............................................................................... 83 6.2.2 Aggregation of molecular data to index values (TELI and MELI) .............. 83 6.2.3 Functional annotation of contigs and metabolites using KEGG ................ 83 6.3 Results and discussion ................................................................................... 85 6.3.1 Results of aggregation techniques ........................................................... 85 6.3.2 Sensitivity analysis of the different molecular approaches and endpoints 86 6.3.3 Mechanistic view of the molecular stress responses based on KEGG functions ............................................................................................................ 89 6.4 Consolidation of the results – holistic interpretation and discussion ............... 93 6.4.1 Adaptation to chronic diuron exposure - from molecular changes to community effects.............................................................................................. 93 6.4.2 Assessment of the ecological costs of Pollution-induced community tolerance based on primary production ............................................................. 94 6.5 Outlook ............................................................................................................ 9

    Anuário científico da Escola Superior de Tecnologia da Saúde de Lisboa - 2021

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    É com grande prazer que apresentamos a mais recente edição (a 11.ª) do Anuário Científico da Escola Superior de Tecnologia da Saúde de Lisboa. Como instituição de ensino superior, temos o compromisso de promover e incentivar a pesquisa científica em todas as áreas do conhecimento que contemplam a nossa missão. Esta publicação tem como objetivo divulgar toda a produção científica desenvolvida pelos Professores, Investigadores, Estudantes e Pessoal não Docente da ESTeSL durante 2021. Este Anuário é, assim, o reflexo do trabalho árduo e dedicado da nossa comunidade, que se empenhou na produção de conteúdo científico de elevada qualidade e partilhada com a Sociedade na forma de livros, capítulos de livros, artigos publicados em revistas nacionais e internacionais, resumos de comunicações orais e pósteres, bem como resultado dos trabalhos de 1º e 2º ciclo. Com isto, o conteúdo desta publicação abrange uma ampla variedade de tópicos, desde temas mais fundamentais até estudos de aplicação prática em contextos específicos de Saúde, refletindo desta forma a pluralidade e diversidade de áreas que definem, e tornam única, a ESTeSL. Acreditamos que a investigação e pesquisa científica é um eixo fundamental para o desenvolvimento da sociedade e é por isso que incentivamos os nossos estudantes a envolverem-se em atividades de pesquisa e prática baseada na evidência desde o início dos seus estudos na ESTeSL. Esta publicação é um exemplo do sucesso desses esforços, sendo a maior de sempre, o que faz com que estejamos muito orgulhosos em partilhar os resultados e descobertas dos nossos investigadores com a comunidade científica e o público em geral. Esperamos que este Anuário inspire e motive outros estudantes, profissionais de saúde, professores e outros colaboradores a continuarem a explorar novas ideias e contribuir para o avanço da ciência e da tecnologia no corpo de conhecimento próprio das áreas que compõe a ESTeSL. Agradecemos a todos os envolvidos na produção deste anuário e desejamos uma leitura inspiradora e agradável.info:eu-repo/semantics/publishedVersio

    Machine Learning Research Trends in Africa: A 30 Years Overview with Bibliometric Analysis Review

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    In this paper, a critical bibliometric analysis study is conducted, coupled with an extensive literature survey on recent developments and associated applications in machine learning research with a perspective on Africa. The presented bibliometric analysis study consists of 2761 machine learning-related documents, of which 98% were articles with at least 482 citations published in 903 journals during the past 30 years. Furthermore, the collated documents were retrieved from the Science Citation Index EXPANDED, comprising research publications from 54 African countries between 1993 and 2021. The bibliometric study shows the visualization of the current landscape and future trends in machine learning research and its application to facilitate future collaborative research and knowledge exchange among authors from different research institutions scattered across the African continent

    Therapeutic effect of mesenchymal stem cell derived extracellular vesicles on 3D model of oligocortical spheroids

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    Extracellular vesicles (EVs) are released by nearly every cell type and are an important structure in inter-cellular communication. Abnormal EV signaling is found in many conditions including ischemia, Alzheimer’s Disease (AD) and Down Syndrome (DS). However, EVs from stem cells from healthy animals have recently emerged as a possible therapeutic intervention to address a variety of neurological conditions. Mesenchymal stromal cell-derived (MSCs) extracellular vesicles from the bone marrow of young healthy monkeys contain microRNAs and proteins and previous studies have shown that MSC-EV treatment mitigates inflammation and oxidative stress, promotes myelination, and improves functional recovery in a rhesus monkey model of cortical injury. EVs have also been shown to reduce AD pathology in mouse models by promoting anti-inflammatory processes and slowing the progression of AD. While AD currently effects over 6 million people in the United States, individuals with DS are disproportionately affected by early onset AD. Therefore, investigating the efficacy of MSC-EVs as a potential therapeutic to mitigate AD like pathology in DS is critical. Accordingly, the current study aims to explore the application of EVs on 3D human brain models of DS, ischemia, and oxygen glucose deprivation (OGD). We generated human oligocortical spheroids (OLS) containing neurons, astrocytes and oligodendrocytes allowing investigation of the effects of the EVs in human, physiologically relevant conditions. First, with OLS in ischemic conditions results were insufficient in demonstrating the recapitulation of cell death and oxidative damage associated with ischemia in vivo. Consequently, the inconsistency of the model prevented us from comprehensive evaluation of the therapeutic potential of EVs in OGD model in OLS. However, we next used DS-derived OLS generated from isogenic induced pluripotent stem cell (iPSCs) lines to evaluate the efficacy of EV treatment in DS. Trisomic OLS display significantly higher levels of amyloid beta (Aβ40 and Aβ42) depositions in both the soluble and insoluble fractions. Additionally, trisomic OLS are consistently smaller than their euploid counterparts, and have elevated levels of cleaved-caspase 3 (CC3) detection indicating more cell death. When treated with EVs, trisomic OLS demonstrated greater preserved cortical volume, significantly decreased levels of Aβ40 and Aβ42 in both fractions, and significant reduction in cell death compared to the untreated trisomic OLS. These results suggest that EVs alleviated the AD-related pathology in DS-derived OLS. Evaluation of the markers of cortical layer neurons demonstrated significantly higher counts of neurons expressing deep and superficial layer markers, suggesting that EVs contributed to greater preserved cortical volume of trisomic OLS by promoting neurogenesis and alleviating trisomy-induced deficits. Our studies show for the first time the efficacy of MSC-EVs in mitigating DS and AD-related cellular phenotypes and pathological depositions in human OLS. Furthermore, oligocortical spheroids present a unique tool for a target validation of potential therapeutics

    The Genetics of Pain: An exploration of gene-by-environment interactions and their effects on pain

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    The findings presented in this dissertation are part of the bigger SYMBIOME project which aims to use the biopsychosocial model of pain to develop a prognostic clinical phenotype for people that experience musculoskeletal (MSK) trauma. Chapter 2 presents an exploratory analysis to assess the relationships between genetic polymorphisms and pain severity and interference. Early childhood trauma was also explored as a moderator between genetic polymorphisms and pain outcomes. For pain severity, major allele carriers (A/A and G/A) of FKBP5 rs9394314 reported significantly higher scores than minor allele carriers (G/G). Further, major allele carriers who had at least one adverse childhood experience (ACE) reported significantly higher scores than minor allele carriers with at least one ACE. For pain interference, minor allele carriers (G/G) of CNR2 rs2501431 scored significantly higher than major allele carriers (A/A and G/A). Chapter 3 presents a cluster analysis that combines genotypes of FKBP5 rs9394314 and CNR2 rs2501431 to explore meaningful relationships with pain and trauma-related distress. ACE was also explored as a moderator of these relationships. Three clusters were identified where the second cluster characterized by major allele carriers of rs9394314 and minor allele carriers of rs2501431 reported significantly higher pain-related functional interference scores. Participants in the second cluster with at least one ACE reported higher pain interference and traumatic distress scores compared to the third cluster, while participants in the first cluster with at least one ACE reported higher pain severity compared to the first cluster. Chapter 4 presents genomic structural equation models (SEM) that explore the relationships of genotypes with trauma-related distress using the traumatic injuries distress scale (TIDS), ACE, and recovery outcomes. The results demonstrate a relationship between TIDS and recovery outcomes, and an indirect relationship between FKBP5 rs9394314 and recovery outcomes exist which is mediated by TIDS. Major allele carriers of FKBP5 rs9394314 reported higher TIDS scores, which was also demonstrated for participants that had at least one ACE. Major allele carriers that scored higher on the TIDS were predicted to be in the none-recovered category. These results support the notion that gene-x-environment interactions may play an important role in pain and recovery

    Discovering the hidden structure of financial markets through bayesian modelling

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    Understanding what is driving the price of a financial asset is a question that is currently mostly unanswered. In this work we go beyond the classic one step ahead prediction and instead construct models that create new information on the behaviour of these time series. Our aim is to get a better understanding of the hidden structures that drive the moves of each financial time series and thus the market as a whole. We propose a tool to decompose multiple time series into economically-meaningful variables to explain the endogenous and exogenous factors driving their underlying variability. The methodology we introduce goes beyond the direct model forecast. Indeed, since our model continuously adapts its variables and coefficients, we can study the time series of coefficients and selected variables. We also present a model to construct the causal graph of relations between these time series and include them in the exogenous factors. Hence, we obtain a model able to explain what is driving the move of both each specific time series and the market as a whole. In addition, the obtained graph of the time series provides new information on the underlying risk structure of this environment. With this deeper understanding of the hidden structure we propose novel ways to detect and forecast risks in the market. We investigate our results with inferences up to one month into the future using stocks, FX futures and ETF futures, demonstrating its superior performance according to accuracy of large moves, longer-term prediction and consistency over time. We also go in more details on the economic interpretation of the new variables and discuss the created graph structure of the market.Open Acces

    Targeting Fusion Proteins of HIV-1 and SARS-CoV-2

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    Viruses are disease-causing pathogenic agents that require host cells to replicate. Fusion of host and viral membranes is critical for the lifecycle of enveloped viruses. Studying viral fusion proteins can allow us to better understand how they shape immune responses and inform the design of therapeutics such as drugs, monoclonal antibodies, and vaccines. This thesis discusses two approaches to targeting two fusion proteins: Env from HIV-1 and S from SARS-CoV-2. The first chapter of this thesis is an introduction to viruses with a specific focus on HIV-1 CD4 mimetic drugs and antibodies against SARS-CoV-2. It discusses the architecture of these viruses and fusion proteins and how small molecules, peptides, and antibodies can target these proteins successfully to treat and prevent disease. In addition, a brief overview is included of the techniques involved in structural biology and how it has informed the study of viruses. For the interested reader, chapter 2 contains a review article that serves as a more in-depth introduction for both viruses as well as how the use of structural biology has informed the study of viral surface proteins and neutralizing antibody responses to them. The subsequent chapters provide a body of work divided into two parts. The first part in chapter 3 involves a study on conformational changes induced in the HIV-1 Env protein by CD4-mimemtic drugs using single particle cryo-EM. The second part encompassing chapters 4 and 5 includes two studies on antibodies isolated from convalescent COVID-19 donors. The former involves classification of antibody responses to the SARS-CoV-2 S receptor-binding domain (RBD). The latter discusses an anti-RBD antibody class that binds to a conserved epitope on the RBD and shows cross-binding and cross-neutralization to other coronaviruses in the sarbecovirus subgenus.</p

    Increased lifetime of Organic Photovoltaics (OPVs) and the impact of degradation, efficiency and costs in the LCOE of Emerging PVs

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    Emerging photovoltaic (PV) technologies such as organic photovoltaics (OPVs) and perovskites (PVKs) have the potential to disrupt the PV market due to their ease of fabrication (compatible with cheap roll-to-roll processing) and installation, as well as their significant efficiency improvements in recent years. However, rapid degradation is still an issue present in many emerging PVs, which must be addressed to enable their commercialisation. This thesis shows an OPV lifetime enhancing technique by adding the insulating polymer PMMA to the active layer, and a novel model for quantifying the impact of degradation (alongside efficiency and cost) upon levelized cost of energy (LCOE) in real world emerging PV installations. The effect of PMMA morphology on the success of a ternary strategy was investigated, leading to device design guidelines. It was found that either increasing the weight percent (wt%) or molecular weight (MW) of PMMA resulted in an increase in the volume of PMMA-rich islands, which provided the OPV protection against water and oxygen ingress. It was also found that adding PMMA can be effective in enhancing the lifetime of different active material combinations, although not to the same extent, and that processing additives can have a negative impact in the devices lifetime. A novel model was developed taking into account realistic degradation profile sourced from a literature review of state-of-the-art OPV and PVK devices. It was found that optimal strategies to improve LCOE depend on the present characteristics of a device, and that panels with a good balance of efficiency and degradation were better than panels with higher efficiency but higher degradation as well. Further, it was found that low-cost locations were more favoured from reductions in the degradation rate and module cost, whilst high-cost locations were more benefited from improvements in initial efficiency, lower discount rates and reductions in install costs
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