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- Cation-exchange resin applied to paralytic Shellfish toxins depuration from Bivalves exposed to Gymnodinium catenatumPublication . Leal, Joana F.; Bombo, Gabriel; Amado, Patrícia; Pereira, Hugo; Lurdes S. Cristiano, M.The accumulation of marine biotoxins in shellfish and their consumption causes serious food safety problems, threatening human health and compromising the availability of protein-based food. It is thus urgent to develop methodologies for the detoxification of live bivalves, avoiding their economic and nutritional devaluation. In this context, we tested an adsorption mechanism of paralytic shellfish toxins (PST) based on a cation-exchange resin. The first studies using cultures of Gymnodinium catenatum (natural producers of PST) showed a decrease of about 80% in overall toxicity after 48 h. Interestingly, we found that the toxins are adsorbed differently, with toxins’ structural features playing a part in the adsorption capacity via steric hindrance, electronic effects, or the extent of positive charge density (e.g., dcSTX). The positive effect of the resin in accelerating PST clearance from live mussels (Mytilus edulis) is not evident when compared to resin-free clearance; nevertheless, relevant information could be gathered that will facilitate further in vivo studies. Several factors appear to be at play, namely the competition of natural substances (e.g., salts, organic matter) for the same binding sites, the blocking of pores due to interactions between molecules, and/or difficulties in resin absorption by mussels. Additionally, the present work revealed the ability of mussels to neutralize pH and proposes bioconversion reactions among the PST molecules.
- Dunaliella viridis TAV01: A halotolerant, Protein-Rich Microalga from the Algarve CoastPublication . Bombo, Gabriel; Cristofoli, N. L.; Santos, Tamara; Schüler, Lisa M.; Maia, Inês Beatriz; Pereira, Hugo; Barreira, Luísa; Varela, JoãoTolerance to harsh environmental conditions, high growth rates and an amino acid profile adequate for human consumption are beneficial features observed in Dunaliella viridis TAV01, a novel strain isolated from a salt pond in the Algarve, Portugal. TAV01 was identified down to the species level by maximum likelihood and Bayesian phylogenetic analyses of the ribosomal internal transcribed spacers one and two regions (ITS1 and ITS-2) and was supported by ITS2 secondary structure analysis. The biochemical profile revealed high protein (35.7 g 100 g−1 DW; 65% higher than the minimum recommended by the World Health Organization) and lipid contents (21.3 g 100 g−1 DW), a relatively higher proportion of the polyunsaturated fatty acids (PUFAs), α-linolenic (26.3% of total fatty acids (TFA)) and linoleic acids (22.8% of TFA), compared to those of other Dunaliella strains, and a balanced essential amino acids profile containing significant levels of leucine, phenylalanine, valine, and threonine. The major carotenoid was lutein, making up over 85% of total carotenoids. The presence of high-quality natural products in D. viridis TAV01 offers the possibility of using this new strain as a valuable biological resource for novel feed or food products as ingredients or supplements.
- Isolation and selection of protein-rich mutants of chlorella vulgaris by fluorescence-activated cell sorting with enhanced biostimulant activity to germinate garden cress seedsPublication . Trovão dos Santos, Mafalda; Schüler, Lisa; Pedroso, Humberto; Reis, Ana; Santo, Gonçalo Espírito; Barros, Ana; Correia, Nádia; Ribeiro, Joana; Bombo, Gabriel; Gama, Florinda; Viana, Catarina; Costa, Monya; Ferreira, Sara; Cardoso, Helena; Varela, João; Silva, Joana; Freitas, Filomena; Pereira, HugoMicroalgae are a promising feedstock with proven biostimulant activity that is enhanced by their biochemical components (e.g., amino acids and phytohormones), which turns them into an appealing feedstock to reduce the use of fertilisers in agriculture and improve crop productivity and resilience. Thus, this work aimed to isolate protein-rich microalgal mutants with increased biostimulant activity. Random mutagenesis was performed with Chlorella vulgaris, and a selection of protein-rich mutants were sorted through fluorescence-activated cell sorting (FACS), resulting in the isolation of 17 protein-rich mutant strains with protein contents 19-34% higher than that of the wildtype (WT). Furthermore, mutant F4 displayed a 38%, 22% and 62% higher biomass productivity, growth rate and chlorophyll content, respectively. This mutant was then scaled up to a 7 L benchtop reactor to produce biomass and evaluate the biostimulant potential of this novel strain towards garden cress seeds. Compared to water (control), the germination index and the relative total growth increased by 7% and 19%, respectively, after the application of 0.1 g L-1 of this bioproduct, which highlights its biostimulant potential.
- Enhancement of heat tolerance by salt stress in Tetraselmis striata CTP4: impacts on HSP gene expression, pigments, and proximal compositionPublication . Santos, Tamara; Pereira, Hugo; Maia, Inês Beatriz; Jacinto, Rita; Bombo, Gabriel; Pinheiro, Filipa Maria Ribeiro; Barreira, Luísa; Varela, JoãoAs the world average temperature is on the rise and heat waves are becoming more prevalent, microalgal producers have been facing significant challenges regarding the time periods during which they are able to grow less thermotolerant microalgae in outdoor production facilities. Therefore, it is urgent to explore how microalgae cope with thermal stress and under which growth conditions tolerance to temperatures close to or higher than their maximum thermal threshold is induced. For this purpose, Tetraselmis striata CTP4, a euryhaline microalga known for its thermotolerance, was selected. Tetraselmis striata belongs to the Chlorodendrophyceae, a clade that branched off early from other "core chlorophyte" clades, usually comprised of microalgae able to colonise freshwater habitats. Here, we present compelling evidence that the ability of this microalga to withstand otherwise lethal thermal upshifts to 40 degrees C is induced by exposure to higher salinity (35 ppt). In contrast, this response is abrogated at lower salinities. Concomitantly, the expression of genes encoding HSP70 and HSP100, two heat shock proteins known to mediate thermotolerance and tolerance to other stresses (e.g., salt stress) in fungi, animals and plants, was enhanced when exposed to both heat stress and higher salinities. This suggests that cross-protective mechanisms against abiotic stress appeared early during the evolution of the core chlorophytes and of Archaeplastida in general. This knowledge can be used to select novel strains and growth conditions that promote thermotolerance in microalgae that are grown in outdoor industrial production facilities in environments where heat waves are expected.
- Toxin profile of two Gymnodinium catenatum strains from Iberian Coastal WatersPublication . Leal, Joana F.; Bombo, Gabriel; Pereira, Hugo; Vicente, Bernardo; Amorim, Ana; Cristiano, Maria L. S.Gymnodinium catenatum has been the main species responsible for paralytic shellfish poisoning events along the Portuguese coast (Iberian Peninsula), causing bans on bivalve harvesting that result in huge economic losses. This work presents the characterization of two novel isolates of G. catenatum regarding their growth and toxin profiles. Laboratory growth experiments revealed that, although low growth rates were obtained during cultivation, the cell yields were high compared to those reported in the literature. Evaluation of the toxin profiles, by HPLC-FLD, essentially confirmed the typical composition of toxins of this regional population (Iberian Peninsula), namely, the absence or low representation of the toxins dcNEO, GTX1,4 and NEO and a higher ratio of the toxins C1,2, GTX6 and GTX5. However, the percentage of the identified toxins varied among the strains of this study (under the same isolation, growth, and analysis conditions), and also differed from that of other strains described in the literature. Interestingly, we found a comparatively high abundance of dcSTX in both strains, relative to the other toxins, and an unquantifiable amount of C3,4 toxins. In addition to the geographic relationship between toxin profiles, chemical conversions among toxins may explain some differences encountered in the toxin profiles of G. catenatum strains.