Browsing by Author "Barata, Carlos"
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- Asparagopsis armata exudate cocktail: the quest for the mechanisms of toxic action of an invasive seaweed on marine invertebratesPublication . Silva, Carla; Simões, Tiago; Félix, Rafael; A.M.V.M., Soares; Barata, Carlos; Novais, Sara C.; Lemos, Marco F.L.The seaweed Asparagopsis armata exhibits a strong invasive behavior, producing halogenated compounds with effective biological effects. This study addresses the biochemical responses to sublethal concentrations of A. armata exudate on the marine snail Gibbula umbilicalis whole body and the shrimp Palaemon elegans eyes and hepatopancreas. Antioxidant defenses superoxide dismutase (SOD) and glutathione-S-transferase (GST), oxidative damage endpoints lipid peroxidation (LPO) and DNA damage, the neuronal parameter acetylcholinesterase (AChE), and the fatty acid profile were evaluated. Results revealed different metabolic responses in both species. Despite previous studies indicating that the exudate affected G. umbilicalis’ survival and behavior, this does not seem to result from oxidative stress or neurotoxicity. For P. elegans, the inhibition of AChE and the decrease of antioxidant capacity is concomitant with the increase of LPO, suggesting neurotoxicity and oxidative stress as contributor mechanisms of toxicity for this species. Fatty acid profile changes were more pronounced for P. elegans with a general increase in polyunsaturated fatty acids (PUFAs) with the exudate exposure, which commonly means a defense mechanism protecting from membrane disruption. Nonetheless, the omega-3 PUFAs arachidonic acid (ARA) and docosapentaenoic acid (DPA) increased in both invertebrates, indicating a common regulation mechanism of inflammation and immunity responses
- Assessment of metal contamination levels and stress responses of endangered sea turtles of São Tomé and PrincipePublication . Morão, Inês F.; Novais, Sara C.; Vieira, Sara; Barata, Carlos; Piña, Benjamin; Lemos, Marco F.L.São Tomé Island harbors important sea turtle nesting and feeding sites. However, insufficient enforcement of environmental laws to avoid illegal take of nesting females and eggs, associated with a great lack of knowledge about how these species interact with their environment and how human activities impact their survival in the region, constitute significant challenges for sea turtle conservation. Through current local conservation projects, some information on genetics and nutrition of sea turtle populations is being unveiled but very little is known about how ocean pollution is impacting these turtles. The main objective of this study was to assess the metal concentrations accumulated by one species of S. Tomé sea turtles (Chelonia mydas) and infer about possible impacts of such contamination on their general stress responses and health status. More specifically, the final goal was to find correlations between metal concentrations and the expression of key genes involved in detoxification/sequestration and metal transport, antioxidant responses and oxidative stress, immunological responses, mitochondrial respiratory and energy production, among others, which could be indicative of these organisms health and future viability. To achieve these goals, nesting female turtles were sampled for blood, immediately after egg laying in their well-documented spawning sites in S. Tomé. Blood samples were withdrawn from the external jugular vein, a portion was stored in EDTA treated tubes at -20ºC until analysis of metal concentrations, and another portion was stored in RNAlater at -20°C until RNA extraction and gene expression analysis using quantitative real-time PCR (qPCR). Additionally, body mass and carapace length were also recorded, along with all information regarding egg spawning and its success. Results showed significant correlations between expression of some genes and metal contaminant levels, pinpointing some candidate genes to be used as biomarkers of interest for biomonitoring campaigns, which worrying function highlights the need for a close follow-up of these organisms. This study represents the first attempt to address pollutant levels and the biological impairments of such stressors in these turtle species nesting in S. Tomé which, given their classification as endangered species (IUCN red list), is of paramount importance to contribute for conservation measures and management.
- Compounds altering fat storage in Daphnia magnaPublication . Jordão, Rita; Garreta, Elba; Campos, Bruno; Lemos, Marco; Soares, Amadeu M.V.M.; Tauler, Romà; Barata, CarlosThe analysis of lipid disruptive effects in invertebrates is limited by our poor knowledge of the lipid metabolic pathways. A recent study showed that tributyltin activated the ecdysteroid, juvenile hormone and retinoic X receptor signaling pathways, and disrupted the dynamics of neutral lipids in the crustacean Daphnia magna impairing the transfer of triacylglycerols to eggs and hence promoting their accumulation in post-spawning females. Tributyltin disruptive effects correlated with lower fitness for offspring and adults. The present study aims to addresses effects of existing compounds on storage lipids in post-spawning females and their health effects. D. magna individuals were exposed 12 chemicals that included vertebrate obesogens (tributyltin, triphenyltin, bisphenol A, nonylphenol, di-2-ethylhexyl phthalate), other contaminants known to affect arthropods (pyriproxyfen, fenarimol, methoprene, emamectin benzoate and fluoxetine), as well as the natural hormones methyl farnesoate and 20-hydroxyecdysone. Reproductive effects were also assessed. Quantitative changes in storage lipids accumulated in lipid droplets were studied using Nile red staining, which showed a close relationship with whole organism levels of triacylglycerols. Ten compounds altered storage lipids in a concentration related manner enhancing (tributyltin, bisphenol A, methyl farnesoate, pyriproxyfen and 20-hydroxyecdysone) or decreasing (nonylphenol, fenarimol, emamectin benzoate, methoprene and fluoxetine) their levels in post-spawning females. Eight compounds that altered lipid levels also had detrimental effects on growth and/or reproduction.
- Fatty acid profile of the sea snail Gibbula umbilicalis as a biomarker for coastal metal pollutionPublication . Silva, Carla; Simões, Tiago; Novais, Sara C.; Pimparel, Inês; Granada, Luana; Soares, Amadeu M. V. M.; Barata, Carlos; Lemos, Marco F. L.Metals are among the most common environmental pollutants with natural or anthropogenic origin that can be easily transferred through the food chain. Marine gastropods are known to accumulate high concentrations of these metals in their tissues. Gibbula umbilicalis ecological importance and abundant soft tissues, which enables extent biochemical assessments, makes this particular organism a potentially suitable species for marine ecotoxicological studies. Fatty acids are carbon-rich compounds that are ubiquitous in all organisms and easy to metabolize. Their biological specificity, relatively well-studied functions and importance, and the fact that they may alter when stress is induced, make fatty acids prospect biomarkers. This work aimed to assess fatty acid profile changes in the gastropod G. umbilicalis exposed to three metal contaminants. After a 168 h exposure to cadmium, mercury, and nickel, the following lipid related endpoints were measured: total lipid content; lipid peroxidation; and fatty acid profile (FAP). The analysis of the FAP suggested an alteration in the fatty acid metabolism and indicated a link between metals exposure and homeoviscous adaptation and immune response. In particular, five fatty acids (palmitic, eicosatrienoic, arachidonic, eicosapentaenoic, and docosahexaenoic acids), demonstrated to be especially good indicators of G. umbilicalis responses to the array of metals used, having thus the potential to be used as biomarkers for metal contamination in this species. This work represents a first approach for the use of FAP signature as a sensitive and informative parameter and novel tool in environmental risk assessment (ERA) of coastal environments, using G. umbilicalis as model species.
- Induction of multixenobiotic defense mechanisms in resistant Daphnia magna clones as a general cellular response to stressPublication . Jordão, Rita; Campos, Bruno; Lemos, Marco; Soares, Amadeu M.V.M.; Tauler, Romà; Barata, CarlosMultixenobiotic resistance mechanisms (MXR) were recently identified in Daphnia magna. Previous results characterized gene transcripts of genes encoding and efflux activities of four putative ABCB1 and ABCC transporters that were chemically induced but showed low specificity against model transporter substrates and inhibitors, thus preventing us from distinguishing between activities of different efflux transporter types. In this study we report on the specificity of induction of ABC transporters and of the stress protein hsp70 in clones selected to be genetically resistant to ABCB1 chemical substrates. Clones resistant to mitoxantrone, ivermectin and pentachlorophenol showed distinctive transcriptional responses of transporter protein coding genes and of putative transporter dye activities. Expression of hsp70 proteins also varied across resistant clones. Clones resistant to mitoxantrone and pentachlorophenol showed high constitutive levels of hsp70. Transcriptional levels of the abcb1 gene transporter and of putative dye transporter activity were also induced to a greater extent in the pentachlorophenol resistant clone. Observed higher dye transporter activities in individuals from clones resistant to mitoxantrone and ivermectin were unrelated with transcriptional levels of the studied four abcc and abcb1 transporter genes. These findings suggest that Abcb1 induction in D. magna may be a part of a general cellular stress response.
- Linking cholinesterase inhibition with behavioural changes in the sea snail Gibbula umbilicalis: Effects of the organophosphate pesticide chlorpyrifosPublication . Silva, Carla O.; Novais, Sara C.; Alves, Luís M. F.; Soares, Amadeu M. V. M.; Barata, Carlos; Lemos, Marco F. L.Inhibition of acetylcholinesterase (AChE) activity has been widely used to assess the exposure and effects of anticholinergic environmental contaminants in several species. The aim of this study was to investigate if sublethal concentrations of the organophosphorous pesticide chlorpyrifos (CPF), a well-known AChE inhibitor, would also affect cholinesterases (ChE) in Gibbula umbilicalis and if this inhibition would result in an alteration of its behaviour, in an attempt to link the effects observed at the cellular level with effects at higher levels of ecological relevance. The biochemical properties of ChEs in this species were first characterized through the assessment of different enzymatic forms present in the sea snail, using different substrates and selective inhibitors. The results suggest that G. umbilicalis possess ChEs with characteristics of typical AChE, which should be the main form present. Additionally, in vitro and in vivo effects of CPF on AChE activity were investigated, along with effects on snails' behaviour: the ability of the snails to move/turn after exposure to the contaminant (flipping test). As expected, CPF inhibited AChE activity both in vitro and in vivo conditions. Moreover, the link between AChE activity inhibition and adverse effects on behavioural changes was established: AChE inhibition was positively correlated with the flipping test, indicating a mechanistic relationship between the two endpoints determined in in vivo exposures. This study highlights the importance of linking biochemical endpoints such as AChE activity with higher level endpoints like behavioural alterations, increasing the ecological relevance of the effects observed.
- Obesogens beyond Vertebrates: Lipid Perturbation by Tributyltin in the Crustacean Daphnia magna.Publication . Jordão, Rita; Casas, Josefina; Fabrias, Gemma; Campos, Bruno; Piña, Benjamín; Lemos, Marco; Soares, Amadeu M V M; Tauler, Romà; Barata, CarlosThe analysis of obesogenic effects in invertebrates is limited by our poor knowledge of the regulatory pathways of lipid metabolism. Recent data from the crustacean Daphnia magna points to three signaling hormonal pathways related to the molting and reproductive cycles [retinoic X receptor (RXR), juvenile hormone (JH), and ecdysone] as putative targets for exogenous obesogens. Objective: The present study addresses the disruptive effects of the model obesogen tributyltin (TBT) on the lipid homeostasis in Daphnia during the molting and reproductive cycle, its genetic control, and health consequences of its disruption. Methods: D. magna individuals were exposed to low and high levels of TBT. Reproductive effects were assessed by Life History analysis methods. Quantitative and qualitative changes in lipid droplets during molting and the reproductive cycle were studied using Nile red staining. Lipid composition and dynamics were analyzed by ultra-performance liquid chromatography coupled to a time-of-flight mass spectrometer. Relative abundances of mRNA from different genes related to RXR, ecdysone, and JH signaling pathways were studied by qRT-PCR. Results and Conclusions: TBT disrupted the dynamics of neutral lipids, impairing the transfer of triacylglycerols to eggs and hence promoting their accumulation in adult individuals. TBT’s disruptive effects translated into a lower fitness for offspring and adults. Co-regulation of gene transcripts suggests that TBT activates the ecdysone, JH, and RXR receptor signaling pathways, presumably through the already proposed interaction with RXR. These findings indicate the presence of obesogenic effects in a nonvertebrate species. © 2015, Public Health Services, US Dept of Health and Human Services. All rights reserved.
- Stress response markers in the blood of São Tomé green sea turtles (Chelonia mydas) and their relation with accumulated metal levelsPublication . Morão, Inês F.C.; Lemos, Marco F.L.; Félix, Rafael; Vieira, Sara; Barata, Carlos; Novais, Sara C.Metals are persistent worldwide being harmful for diverse organisms and having complex and combined effects with other contaminants in the environment. Sea turtles accumulate these contaminants being considered good bioindicator species for marine pollution. However, very little is known on how this is affecting these charismatic animals. Sâo Tomé and Príncipe archipelago harbours important green sea turtle (Chelonia mydas) nesting and feeding grounds. The main goal of this study was to determine metal and metalloid accumulation in the blood of females C. mydas nesting in São Tomé Island, and evaluate the possible impacts of this contamination by addressing molecular stress responses. Gene expression analysis was performed in blood targeting genes involved in detoxification/sequestration and metal transport (mt, mtf and fer), and in antioxidant and oxidative stress responses (cat, sod, gr, tdx, txrd, selp and gclc). Micronuclei analysis in blood was also addressed as a biomarker of genotoxicity. Present results showed significant correlations between different gene expressions with the metals evaluated. The best GLM models and significant relationships were found for mt expression, for which 78% of the variability was attributed to metal levels (Al, Cu, Fe, Hg, Pb and Zn), followed by micronuclei count (65% - Cr, Cu, Fe, Hg, Mn and Zn), tdx expression (52% - Cd, Fe, Mn, Pb and Se), and cat expression (52% - As, Fe, Se and Cd x Hg). Overall, this study demonstrates that these green sea turtles are trying to adapt to the oxidative stress and damage produced by metals through the increased expression of antioxidants and other protectors, which raises concerns about the impacts on these endangered organisms’ fitness. Furthermore, promising biomarker candidates associated to metal stress were identified in this species that may be used in future biomonitoring studies using C. mydas’ blood, allowing for a temporal follow-up of the organisms.
- Two-generational effects of contaminants in Daphnia magna : Effects of offspring qualityPublication . Campos, Bruno; Jordão, Rita; Rivetti, Claudia; Lemos, Marco; Soares, Amadeu; Tauler, Roma; Barata, CarlosThe authors set up a protocol to perform a 2-generational ring test using the existing guidelines for the Daphnia magna reproduction test. It is well known in ecology that size and quality of offspring vary across the first clutches in D. magna and that certain chemicals affect offspring quality. Therefore, the origin of the second generation is an important factor to consider. Two-generational effects across first, second, and third clutches were evaluated using 4-nonylphenol; those across first and third clutches were evaluated using tributyltin, and those across the third clutch were evaluated using piperonyl butoxide. The compound showing the greatest aggravation of toxic effects between the parental and second generations was piperonyl butoxide, followed by 4-nonylphenol, whereas intergenerational effects of tributyltin varied across experiments. The studied chemicals affected the quantity and quality of the offspring produced by exposed females of the parental generation, those effects being greater in third-clutch neonates. Therefore, when third-clutch offspring were further exposed, they turned out to be more sensitive than the parental generation. The results are in line with those obtained in multigenerational studies using mammalian tests, which showed that, in many cases, effects on the second generation can be predicted by evaluating the quality of the offspring produced.