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Demonstration of the ability of the bacterial polysaccharide FucoPol to flocculate kaolin suspensions

Publication . Araújo, Diana; Concórdio-Reis, Patrícia; Marques, Ana C.; Sevrin, Chantal; Grandfils, Christian; Delgado Alves, Vitor

In this study, the flocculation properties of FucoPol, a bacterial extracellular polysaccharide, were investigated. FucoPol is a high molecular weight polymer and negatively charged due to the presence of glucuronic acid and the acyl groups succinyl and pyruvyl. High flocculation rate values (>70%) were achieved with a low bioflocculant dosage of 1 mg/L, for pH values in the range 3–5 and temperature within 15–20°C. The bioflocculant was also shown to be stable after freezing/thawing and heating up to 100°C. Given the polymer’s anionic character, the size of flocs formed and their surface profile, bridging seems to be the main flocculation mechanism of FucoPol. This study demonstrated that FucoPol is a promising natural, biodegradable and biocompatible alternative to the currently used synthetic or inorganic hazardous products, with potential to be used as a novel flocculation agent in several applications, such as water treatment, food or mining. Further studies will involve evaluating the reduction of cation dosage on flocculation efficiency, as well as testing the applicability of FucoPol to flocculate different types of suspended solids, such as, for example, activated carbons, soil solids or yeast cells

2020Journal article

Open access

Evaluation of thermal stress in tropical marine organisms in the context of climate warming

Publication . Madeira, Carolina; Vinagre, Catarina Maria Batista, 1978-; Diniz, Mário; Cabral, Henrique N., 1969-

The marine environment is already registering the impacts of climate change. The current increase in global temperature since pre-industrial times is disrupting life in the oceans, from the tropics to the poles. The key impacts for marine species as warming continues include shifting home ranges and altered life histories due to the direct effects of temperature on metabolism, life cycles and behaviour of organisms. These pervasive effects on species will also have a significant impact in the goods and services provided by the ocean to human society. However, the vulnerability of tropical marine species (e.g. reef fish and non-coral invertebrates) towards ocean warming is still far from clear, not only due to the gap of knowledge on general ecology and biology of most tropical species, but also because there is a lack of integrative approaches addressing physiological and molecular thermal compensation mechanisms, as well as a lack of adequate or optimized assessment tools for tropical habitats. Therefore, the aim of this thesis was to assess the vulnerability of marine species (fish, crustaceans and gastropods) from tropical reef environments to ocean warming (gradual temperature increase and heat waves), using molecular- (proteins and antioxidant enzymes), cellular (lipid peroxidation), tissue- (energy reserves), organism- (condition, critical thermal maxima) and sample population (acclimation rate, thermal safety margins, mortality) parameters to measure stress and performance. General methodologies included collecting animals from tropical shallow waters with hand nets, experimental assays testing acute and chronic thermal stress and thermal tolerance in the lab (following control and warming scenarios), periodic samplings of several tissue types and biomarker quantification (sample homogenization and posterior immunoassays, kinetic colorimetric assays and elemental analyses by isotope ratio mass spectrometry). Mathematical calculations were used for several performance parameters as well as stress indices. Multivariate (as well as multifactorial) statistical analyses were then performed for all datasets. Results revealed that chronically increased water temperatures (30˚C – 32˚C) elicited a time-dependent cellular stress response in all species tested, where chaperones and antioxidant enzymes showed the greatest fold-changes. The latter molecular responses were highly inducible in vital organs such as gills, liver and muscle, suggesting a relation to tissue function, metabolic- and oxygen diffusion rate which determine the flux of reactive oxygen species and damage potential in cells. Additionally, animals subjected to increased temperatures also showed poorer health status and decreased body condition with lower energy reserves when compared to control temperatures, although mortality levels remained unchanged. An acute exposure to increased thermal load revealed plastic upper thermal limits. However, thermal safety margins were generally low for shallow water species in reef environments, indicating a low ability to tolerate further future warming. This means that organisms will have to rely on subtidal habitats for thermal refugia as shallow water habitats (e.g. tide pools) become ecological traps where animals cannot escape heat. Overall, the results point to the importance of monitoring thermal change in the wild to provide decision makers with appropriate and detailed information to sustain conservation efforts in fragile tropical marine ecosystems.

2018Doctoral thesis

Open access

Novel hydrogels based on yeast chitin-glucan complex: Characterization and safety assessment

Publication . Araújo, Diana; Delgado Alves, Vitor; Lima, Sofia A.C.; Reis, Salette; Freitas, Filomena; Reis, Maria A.M.

Chitin-glucan complex (CGC) was used for the first time for the preparation of hydrogels. Alkali solvent systems, NaOH and KOH solutions, either at 1 or 5 mol/L, were used for CGC dissolution using a freeze-thaw procedure (freezing at −20 °C and thawing at room temperature; four cycles). The CGC solutions thus obtained were subjected to dialysis that induced the spontaneous gelation of the biopolymer, yielding translucid hydrogels with a yellowish coloration. Although all CGC hydrogels exhibited porous microstructures, high water content (above 97%) and good mechanical properties, their morphology, viscoelastic properties and texture were influenced by the type of solvent system used for CGC dissolution, as well as by their ionic strength. The K-based hydrogels presented a less compact network with larger pores and exhibited lower elastic properties. The Na-based hydrogels, on the other hand, exhibited a denser structure with smaller pores and a stiffer gel structure. These results show that it is possible to prepare CGC hydrogels with differing characteristics that can be suitable for different applications. Furthermore, all hydrogels were non-cytotoxic towards L929 fibroblasts and HaCaT keratinocytes. This study demonstrates CGC can be used to prepare biocompatible hydrogels with properties render them promising biomaterials

2020Journal article

Open access

Epitope mapping by NMR of a novel anti-Aβ antibody (STAB-MAb)

Publication . Posado-Fernández, Adrián; Afonso, Cláudia F.; Dória, Gonçalo; Flores, Orfeu; Cabrita, Eurico J.

Alzheimer´s Disease (AD) is one of the most common neurodegenerative disorders worldwide. Excess of β-amyloid (Aβ), a peptide with a high propensity to misfold and self-aggregate, is believed to be the major contributor to the observed neuronal degeneration and cognitive decline in AD. Here, we characterize the epitope of a novel anti-Aβ monoclonal antibody, the STAB-MAb, which has previously demonstrated picomolar affinities for both monomers (KD = 80 pM) and fibrils (KD = 130 pM) of Aβ(1-42) and has shown therapeutic efficacy in preclinical mouse models of AD. Our findings reveal a widespread epitope that embraces several key Aβ residues that have been previously described as important in the Aβ fibrillation process. Of note, STAB-MAb exhibits a stronger affinity for the N-terminus of Aβ and stabilizes an α-helix conformation in the central to N-terminal region of the peptide, in addition to disrupting a characteristic salt-bridge of a hairpin structure present in fibrils. The NMR derived epitope supports the observed results from ThT-monitored fluorescence and electron microscopy experiments, in which STAB-MAb was shown to inhibit the formation of aggregates and promote disruption of pre-formed fibrils. In combination with the published in vitro and in vivo assays, our study highlights STAB-MAb as a rare and versatile antibody with analytical, diagnostic and therapeutic efficacy.

2019Journal article

Open access

Neonatal cholestasis : development of a diagnostic decision algorithm from multivariate predictive models

Publication . Santos Silva, Ermelinda; Moreira Silva, Helena; Catarino, Cristina; Dias, Cláudia Camila; Santos Silva, Alice; Lopes, Ana Isabel

Despite the recent advances involving molecular studies, the neonatal cholestasis (NC) diagnosis still relays on the expertise of medical teams. Our aim was to develop models of etiological diagnosis and unfavourable prognosis which may support a rationale diagnostic approach. We retrospectively analysed 154 patients born between January 1985 and October 2019. The cohort was divided into two main groups: (A) transient cholestasis and (B) other diagnosis (with subgroups) and also in two groups of outcomes: (I) unfavourable and (II) favourable. Multivariate logistic regression analysis identified the lower gestational age as the only variable independently associated with an increased risk of transient cholestasis and signs and/or symptoms of sepsis with infectious or metabolic diseases. Gamma-glutamyl transferase serum levels > 300 IU/L had a positive predictive value for both diagnosis of biliary atresia and for alpha-1-antitrypsin deficiency (A1ATD) and for unfavourable prognosis. A model of diagnosis for A1ATD (n = 34) showed an area under the ROC curve = 0.843 [confidence interval (CI): 0.773-0.912].Conclusion: This study identified some predictors of diagnosis and prognosis which helped to build a diagnostic decision algorithm. The unusually large subgroup of patients with A1ATD in this cohort emphasizes its predictive diagnostic model. What Is Known • The etiological diagnosis of neonatal cholestasis (NC) requires a step-by-step guided approach, and diagnostic models have been developed only for biliary atresia. • Current algorithms neither address the epidemiology changes nor the application of the new molecular diagnostic tools. What Is New • This study provides diagnostic predictive models for patients with A1ATD, metabolic/infectious diseases, and transient cholestasis, and two models of unfavourable prognosis for NC. • A diagnostic decision algorithm is proposed based on this study, authors expertise and the literature.

2021Journal article

Restricted access