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- The interplay between membrane lipids and phospholipase A family members in grapevine resistance against Plasmopara viticolaPublication . Laureano, Gonçalo; Figueiredo, Joana; Cavaco, Ana Rita; Duarte, Bernardo; Caçador, Isabel; Malhó, Rui; Sousa Silva, Marta; Matos, Ana Rita; Figueiredo, AndreiaGrapevine downy mildew, caused by the biotrophic oomycete Plasmopara viticola, is one of the most important diseases in modern viticulture. The search for sustainable disease control measure is of extreme importance, thus becoming imperative to fully characterize the mechanisms leading to an incompatible interaction. We have previously shown that lipid signalling events play an important role in grapevine's response to this pathogen, namely through changes in linolenic acid content, lipid peroxidation and jasmonic acid synthesis. Here, we have characterized the modulation of lipid metabolism in leaves from two V. vinifera cultivars (resistant and susceptible to P. viticola) in the first hours after pathogen inoculation. Prior to pathogen inoculation both genotypes present an inherently different fatty acid composition that is highly modulated in the resistant genotype after pathogen challenge. Such changes involve modulation of phospholipase A activity suggesting that the source of lipids mobilized upon pathogen infection are the chloroplast membranes. This work thus provides original evidence on the involvement of lipid signalling and phospholipases in grapevine immune responses to pathogen infection. The results are discussed considering the implications on the plant's physiological status and the use of discriminating lipid/fatty acids pattern in future selection procedures of cultivars.
- Leaf fatty acid remodeling in the salt-excreting halophytic grass Spartina patens along a salinity gradientPublication . Duarte, Bernardo; Matos, Ana Rita; Marques, João Carlos; Caçador, IsabelSpartina patens is a highly dispersed halophytic grass invader in Mediterranean marshes. It is also characterized by having a high degree of resistance to salinity, one of the main drivers of plant zonation in salt marshes. Nevertheless, the physiological basis behind the extreme resistance of S. patens requires more detailed studies. In the present work, we aimed to study how membrane fatty acid remodeling could contribute to the resistance of this plant to salt. Spartina patens individuals exposed to increasing levels of salinity and its leaf fatty acid profile under lipid peroxidation products evaluated under all tested concentrations. A significant increase in the relative amounts of the saturated fatty acids (SFA) was observed, namely palmitic acid (C16:0), essential for PS II functioning, and stearic (C18:0) acid. The chloroplastidial trans-hexadecenoic acid (C16:1t) as well as the polyunsaturated linoleic (C18:2) and linolenic (C18:3) acids showed significant decreases in all the salt treatments. These changes led to a reduction in the double bond index in salt-treated plants which reflects reduction of the fluidity of the chloroplast membranes, which could contribute to maintain the membrane impermeable to the toxic exogenous Na. Despite the decrease observed in the total fatty acid contents in plants exposed to high salt concentrations the amounts of lipid peroxidation products decreased highlighting the resistance of this species towards toxic exogenous salt concentrations. Membrane fatty acid remodeling could represent an efficient mechanism to maintain the photosynthetic machinery of S. patens highly efficient under salt stress.
- Halophyte fatty acids as biomarkers of anthropogenic-driven contamination in Mediterranean marshes: Sentinel species survey and development of an integrated biomarker response (IBR) indexPublication . Duarte, Bernardo; Carreiras, João; Pérez-Romero, Jesús Alberto; Mateos-Naranjo, Enrique; Redondo-Gómez, Susana; Matos, Ana Rita; Marques, J. C.; Caçador, IsabelEstuarine environments are extremely affected by anthropogenic-driven contamination, namely heavy metals. In the recent years, several organisms have been studied to be used as sentinel species providing a wide range of biomarkers for estuarine contamination. Only recently non-traditional biomarkers, such as fatty acids, were included in animal ecotoxicology and impact assessment studies. Nevertheless, there is a wide lack of knowledge regarding to its application in estuarine plants. Considering this the present work aimed to evaluate the application of fatty acid profiling in Mediterranean halophytic species chronically exposed to different degrees of metal contamination, while incorporating these fatty acid biomarkers into an unifying index. From the fatty acids analysis two different types of sentinel halophytes could be identified: the contamination sensitive Halimione portulacoides (sea purslane), Sarcocornia fruticosa (glasswort) and Spartina patens (saltmeadow cordgrass), and the contamination-tolerant Spartina maritima (small cordgrass). In sensitive species the most evident changes in the fatty acid profiles were the decrease in the 18:3 and the increase in the 16:0 fatty acid relative concentrations, while the inverse trend was recorded in the tolerant S. maritima under chronic contamination. Beyond the evident physiological importance, these shifts in the halophyte fatty acid profiles provide some understanding on their use as biomarkers of metal contamination. After application of an integrated biomarker response (IBR) index it was possible to conclude that in sensitive halophytes (H. portulacoides, S. fruticosa and S. patens) the samples collected from the contaminated site produce had higher IBR values than the tolerant species S. maritima being these lower values linked with an adaptation mechanism towards contamination. Furthermore, the fatty acid-based IBR index was intrinsically connected with the bioavailable metals concentrations in the rhizosediments, which make that it could be used in future impact assessment and/or ecotoxicology studies. In summary, the data showed that two of the most abundant halophytes in the Mediterranean eco-region (S. fruticose and H. portulacoides) are potential sentinel species of metal contamination, whereas its fatty acid profile is an efficient biomarker of the degree of environmental contamination.
- Preliminary diversity assessment of an undervalued tropical bean (Lablab purpureus (L.) Sweet) through fatty acid profilingPublication . Vidigal, Patrícia; Duarte, Bernardo; Cavaco, Ana Rita; Caçador, Isabel; Figueiredo, Andreia; Matos, Ana Rita; Viegas, Wanda; Monteiro, FilipaSeveral large-scale metabolic profiling studies have been directed to prospect crops with a major focus on yield-related traits and, ultimately, with the definition of specific markers for plant selection in breeding programs. However, some of these technologies are expensive, time-consuming and not easily feasible for a quick approach. Fatty acid profiling was described as reliable biomarkers and as a chemotaxonomic tool allowing to study not only the diversity in germplasm collections but also to discriminate their geographic origin. We have used fatty acids profiling for a preliminary assessment of Lablab purpureus (L.) Sweet (hyacinth bean) diversity and landraces discrimination. Hyacinth bean displays an enormous variability of agro-morphological traits, probably linked to the multi-purpose uses in different regions, i.e. as pulse, or as food with nutraceutical potential (Africa and Asia), forage (Africa and Australia) and ornamental (Europe and USA). Only two forage cultivars are widely marketed, cv. Rongai and cv. Highworth, with several landraces remaining to be addressed in terms of diversity. We show that fatty acids profiling was able to distinguish landraces, which display shared fatty acids with cultivars from the center of hyacinth bean diversity origin (East Africa). We propose that fatty acid profiling is a tool that may be used not only for nutritional value assessment but also as a chemodiversity tool in crop research.
- Ecotoxicity of the lipid-lowering drug bezafibrate on the bioenergetics and lipid metabolism of the diatom Phaeodactylum tricornutumPublication . Duarte, Bernardo; Prata, Diogo; Matos, Ana Rita; Cabrita, Maria Teresa; Caçador, Isabel; Marques, J. C.; Cabral, Henrique; Reis Santos, Patrick; Fonseca, VanessaPharmaceutical residues impose a new and emerging threat to the marine environment and its biota. In most countries, ecotoxicity tests are not required for all pharmaceutical residues classes and, even when mandatory, these tests are not performed using marine primary producers such as diatoms. These microalgae are among the most abundant class of primary producers in the marine realm and key players in the marine trophic web. Blood-lipid-lowering agents such as bezafibrate and its derivatives are among the most prescribed drugs and most frequently found human pharmaceuticals in aquatic environments. The present study aims to investigate the bezafibrate ecotoxicity and its effects on primary productivity and lipid metabolism, at environmentally relevant concentrations, using the model diatom Phaeodactylum tricornutum. Under controlled conditions, diatom cultures were exposed to bezafibrate at 0, 3, 6, 30 and 60 μg L-1, representing concentrations that can be found in the vicinity of discharges of wastewater treatment plants. High bezafibrate concentrations increased cell density and are suggested to promote a shift from autotrophic to mixotrophic metabolism, with diatoms using light energy generated redox potential to breakdown bezafibrate as carbon source. This was supported by an evident increase in cell density coupled with an impairment of the thylakoid electron transport and consequent photosynthetic activity reduction. In agreement, the concentrations of plastidial marker fatty acids showed negative correlations and Canonical Analysis of Principal coordinates of the relative abundances of fatty acid and photochemical data allowed the separation of controls and cells exposed to bezafibrate with high classification efficiency, namely for photochemical traits, suggesting their validity as suitable biomarkers of bezafibrate exposure. Further evaluations of the occurrence of a metabolic shift in diatoms due to exposure to bezafibrate is paramount, as ultimately it may reduce O2 generation and CO2 fixation in aquatic ecosystems with ensuing consequences for neighboring heterotrophic organisms.
- Investigating the physiological mechanisms underlying Salicornia ramosissima response to atmospheric CO2 enrichment under coexistence of prolonged soil flooding and saline excessPublication . Pérez-Romero, Jesús Alberto; Duarte, Bernardo; Barcia-Piedras, Jose-Maria; Matos, Ana Rita; Redondo-Gómez, Susana; Caçador, Isabel; Mateos-Naranjo, EnriqueA 45-days long climatic chamber experiment was design to evaluate the effect of 400 and 700 ppm atmospheric CO2 treatments with and without soil water logging in combination with 171 and 510 mM NaCl in the halophyte Salicornia ramosissima. In order to ascertain the possible synergetic impact of these factors associate to climatic change in this plant species physiological and growth responses. Our results indicated that elevated atmospheric CO2 concentration improved plant physiological performance under suboptimal root-flooding and saline conditions plants. Thus, this positive impact was mainly ascribed to an enhancement of energy transport efficiency, as indicated the greater PG, N and Sm values, and the maintaining of carbon assimilation capacity due to the higher net photosynthetic rate (AN) and water use efficiency (iWUE). This could contribute to reduce the risk of oxidative stress owing to the accumulation of reactive oxygen species (ROS). Moreover, plants grown at 700 ppm had a greater capacity to cope with flooding and salinity synergistic impact by a greater efficiency in the modulation in enzyme antioxidant machinery and by the accumulation of osmoprotective compounds and saturated fatty acids in its tissues. These responses indicate that atmospheric CO2 enrichment would contribute to preserve the development of Salicornia ramosissima against the ongoing process of increment of soil stressful conditions linked with climatic change.
- Phytoplankton community-level bio-optical assessment in a naturally mercury contaminated Antarctic ecosystem (Deception Island)Publication . Duarte, Bernardo; Cabrita, Maria Teresa; Vidal, Tânia; Pereira, Joana Luísa; Pacheco, Mário; Pereira, Patrícia; Canário, João; Gonçalves, Fernando J.M.; Matos, Ana Rita; Rosa, Rui; Marques, J. C.; Caçador, Isabel; Gameiro, C.Mercury naturally contaminated environments, like Deception Island (Antarctica), are field labs to study the physiological consequences of chronic Hg-exposure at the community level. Deception Island volcanic vents lead to a continuous chronic exposure of the phytoplanktonic communities to potentially toxic Hg concentrations. Comparing Hg-contaminated areas (Fumarolas Bay - FB, Gabriel de Castilla station - GdC station), no significant differences in chlorophyll a concentrations were detected, indicating that biomass production was not impaired by Hg-exposure despite the high Hg levels found in the cells. Moreover, the electron transport energy, responsible for energy production, also presented rather similar values in phytoplankton from both locations. Regarding FB communities, although the cells absorbed and trapped lower amounts of energy, the effect of Hg was not relevant in the photochemical work produced by the electronic transport chain. This might be due to the activation of alternative internal electron donors, as counteractive measure to the energy accumulated inside the cells. In fact, this alternative electron pathway, may have allowed FB communities to have similar electron transport energy fluxes without using respiration as photoprotective measure towards excessive energy. Hg-exposed cells also showed a shift from the energy flux towards the PS I (photosystem I), alleviating the excessive energy accumulation at the PS II (photosystem II) and preventing an oxidative burst. Our findings suggest a higher energy use efficiency in the communities exposed to volcanic Hg, which is not observable in cultured phytoplankton species grown under Hg exposure. This may constitute a metabolic adaptation, driven from chronic exposure allowing the maintenance of high levels of primary productivity under the assumingly unfavourable conditions of Deception Island.
- Heat wave impacts on the model diatom Phaeodactylum tricornutum: Searching for photochemical and fatty acid biomarkers of thermal stressPublication . Feijão, Eduardo; Gameiro, C.; Franzitta, Marco; Duarte, Bernardo; Caçador, Isabel; Cabrita, Maria Teresa; Matos, Ana RitaGlobal warming is increasing the frequency and intensity of extreme thermal events, with inevitable consequences for marine ecosystems and organisms. Phytoplankton is at the base of marine food webs and diatoms are major contributors to global primary production. Therefore, environmental changes, such as heat, influencing growth, physiology and biochemical composition of diatoms, impact other organisms at higher trophic levels. The model diatom Phaeodactylum tricornutum, particularly rich in the long chain omega-3 eicosapentaenoic acid (EPA), and able to accumulate substantial amounts of storage lipids, has recently been the object of numerous works, regarding fundamental aspects of lipid metabolism and exploring its biotechnological potential for biodiesel and aquaculture purposes. The aim of this study was to use P. tricornutum, growing under controlled conditions, to examine the effects of a heat wave, in order to identify heat stress biomarkers. The photosynthetic and respiratory metabolism was investigated by Chlorophyll a fluorescence and by O2 evolution and discussed in connection with changes observed in the composition of photosynthetic pigments and fatty acids. Phaeodactylum tricornutum cells exposed to 26 °C displayed lower photosynthetic O2 production, but similar respiratory rate, comparing to cells at control temperature (18 °C), which is likely related to the biomass decrease observed under heat stress. Heat wave exposed cells also showed a less efficient PSII, higher energy dissipation and higher chlorophyll a and fucoxanthin concentrations, suggesting a heat-induced amplification of the light energy absorption capacity. Heat wave exposed cells showed lower relative EPA contents and double bond indexes, whereas the parameter inversely related to nutritional value, omega 6/omega 3 ratio, increased. Moreover, the analysis of the fatty acid profiles also suggested that heat exposure negatively impacted thylakoid lipids, in agreement with the decrease observed in photosynthesis. Results obtained highlight the negative impact of heat waves on diatom photosynthesis and nutritional value, as well as on their capacity to oxygenate ocean water. Furthermore, physiological parameters as well as fatty acids and photosynthetic pigments signatures, were identified, that could represent expedite biomarkers of thermal stress in future studies.