Water temperature modulates mercury accumulation and oxidative stress status of common goby (Pomatoschistus microps).

Mercury (Hg) is a widespread pollutant across estuarine and coastal areas, raising concern on its potential impact on aquatic organisms. Hg may origin from natural and anthropogenic sources, being persistent and potentially toxic to biota, ultimately representing a serious risk to human health. Hg accumulation and toxicity may also induce reactive oxygen species (ROS) production in marine organisms, responsible for cell and tissue damage. Additionally, the temperature is undoubtedly an important environmental factor to consider regarding accumulation, due to its marked influence on the physiology and ecology of aquatic organisms. This study aimed to investigate the effect of different temperature scenarios (15, 20 and 25 °C) on the Hg accumulation in Pomatoschistus microps (Krøyer, 1838) liver and muscle, as well as on oxidative stress responses and energy metabolism, after short-term exposure to a naturally contaminated sediment with an environmentally relevant [Hg] (1.2 µg g-1). The results showed that Hg accumulation tends to increase along the temperature gradient with higher values of Hg accumulated in liver than in muscle tissue. The action of antioxidant enzymes and stress proteins seems to be effective in combating oxidative stress in the liver. Despite the action of antioxidant defences in the muscle, oxidative damage was observed at the protein level concomitantly with a decrease in aerobic energy production after exposure to Hg at higher temperatures. These findings are ecologically relevant and highlight the importance of further investigation of combined effects of Hg and other stressors, especially in a scenario of a changing climate where events leading to rapid alterations on water parameters are more frequent.

Mercury bioaccumulation in the long-fin squid Loligo forbesi near the Mid-Atlantic Ridge: Implications to human exposure.

Due to growing commercial interest as a fishing resource and its intermediate position in the marine trophic chains as both prey and predator, cephalopods can play an important role in mercury (Hg) transfer along the marine food webs, since they can bioaccumulate this metal in its tissues. Our study aims to analyze Hg accumulation in Loligo forbesi caught in the Azores Archipelago (Portugal) quantifying Hg in different tissues (mantle and stomach), as well in the squid stomach content, in order to evaluate the efficiency of Hg transfer from prey to predator. Hg data from the tissues was used to estimate the weekly tolerable Hg intake due to squid consumption. Overall data indicate that Hg measured in the stomach tissue (0.1 ± 0.01 µg g-1) was significantly higher than Hg levels found in the mantle (0.04 ± 0.001 µg g-1) and stomach contents (0.01 ± 0.001 µg g-1). BMF (bioaccumulation factor) was >1 for all the samples, indicating a biomagnification process from prey to predator. Hg concentration in the mantle tissue was correlated with mantle size; although females present higher Hg levels than males, the difference was found to be not related to gender but rather to the fact that females had larger bodies. Finally, considering the Hg concentration found in the mantle and the permitted Hg levels, it is advisable to consume up to 1050-1890g of squid per week, according to the regulatory agencies. Thus, our results indicate that, since these doses are respected, consumption of squids from the Azorean waters do not pose a risk to humans.

Effects of diclofenac and salicylic acid exposure on Lemna minor: Is time a factor?

The global occurrence of pharmaceuticals in the aquatic environment has been considered a particularly concerning problem with unknown consequences. Non-steroidal anti-inflammatory drugs (NSAIDs) including diclofenac (DCF) and salicylic acid (SA), are among the most frequently prescribed drugs in the world, being consequently commonly found in the aquatic environment. Prolonged experiments (with duration of exposure that surpass those recommended by already established testing guidelines) are important to obtain ecologically relevant data to address the issue of NSAIDs ecotoxicity, because by being more realistically (namely in terms of levels and durations of exposure), such tests may indicate realistic challenges posed to aquatic organisms. Among the most common test species that are used for assessing environmental quality, plants play a leading role. Lemna species are among the most important plants used for ecotoxicity testing. Therefore, the aim of this study was to evaluate the temporal effect of a prolonged exposure of DCF and SA on Lemna minor. To attain this purpose, L. minor plants were chronically exposed to 0, 4, 20, and 100 µg/L of both pharmaceuticals, and samplings were performed at 6, 10 and 14 days of exposure. The analyzed endpoints were: levels of chlorophyll a, b and total, carotenoids; and enzymatic biomarkers, such as catalase, ascorbate peroxidase and glutathione-S-transferases. Diclofenac was responsible for alterations in all analyzed parameters in different intervals of exposure. Salicylic acid exposure was not capable of causing alterations on pigment contents of L. minor, however, enzymatic biomarkers were altered at all sampling intervals. Thus, it is possible to conclude that both pharmaceuticals can cause damage on the tested macrophyte species, biochemical parameters being more sensitive than physiological ones. Additional prolonged experiments are required to understand the chronic effects of different pharmaceuticals in the aquatic environment, especially in plants.

Multi-generational exposure to Pb in two monophyletic Daphnia species: Individual, functional and population related endpoints.

To better evaluate chemical damage in chronically contaminated habitats, a nine-generational exposure to Lead (Pb) was done with two monophyletic Daphnia species, from temperate (Daphnia magna) and tropical (Daphnia similis) environments. The multi-generational test consisted generally of a continuous Pb exposed set of organisms, plus an extra control set running simultaneously. To assess daphnids recovery after Pb exposure, some organisms from the sixth generation were transferred to clean media for three extra generations (recovery period; F6 to F9), while others were keep Pb exposed. All setups (control, Pb exposure and recovery period) were submitted to two different dietary regimes, the standard (3 × 105 cells/mL) and restricted food (1.5 × 105 cells/mL) regimes. To evaluate the effects of generational Pb exposure and food regimes, individual, functional and population related endpoints were assessed (number of offspring, body length and rate of population increase (r) and feeding rate (FR)). The tests were conducted on the first (F0) and last generations (F9). No differences were shown on number of offspring and feeding among F9 control and continuous Pb exposed D. magna, although a higher r was shown for F9 Pb exposed organisms. F9 Pb exposed D. similis also presented a higher r than F9 control, however, lethality was induced at high Pb exposure levels. At food restriction the patterns were opposite and D. magna died at high Pb exposure while Pb exposed D. similis was the only setup (compared to control and recovery period) to survive at high Pb exposure levels. Regarding the recovery period, D. magna (standard food) did not cope well with the Pb re-exposure and lethality was induced, while D. similis indicate a decreased Pb sensitivity (only setup that survived high Pb exposure levels). Under food restriction, both species presented a decreased Pb sensitivity and consequent failed recovery (possibly due to epigenetic changes). Both species presented similar patterns regarding generations. Organisms from F0 presented enhanced reproductive outputs in comparison to F9 and the contrary occurred to the FR (even in control organisms). Data show an acclimation under a generational Pb exposure, which could increase the population of adapted organisms in natural habitats. And, since there was not a full recovery after three generations in clean media, an indication of epigenetic changes for both species may also be considered.

Toxicity of lead and mancozeb differs in two monophyletic Daphnia species.

Lead and mancozeb are two important chemicals used for different human purposes and activities worldwide. Hazard assessment in different areas of the world is carried out with different but phylogenetically similar species, adapted to different climatic conditions, in order to increase relevance. This study evaluated the sensitivity of two monophyletic species, the tropical species Daphnia similis and the temperate species Daphnia magna, to the two chemicals lead and mancozeb. Standard acute and chronic ecotoxicological tests (reproduction and growth), as well as other sublethal measurements such as the intrinsic rate of population increase (r), feeding rate (FR) and O2 consumption, were recorded along with the analysis of the AChE activity to determine the neurotoxicity of both contaminants. Albeit their similar evolutionary status, D. magna generally presented a lower sensitivity to Pb in comparison to D. similis. Despite the differences in sensitivity, both species presented similar patterns of response under Pb exposure, with diminished reproductive outputs, feeding impairment, reduced O2 consumption and no effect on AChE activity. Mancozeb decreased the reproduction, rate of population increase and feeding rate, increased the AChE activity in both species and increased O2 consumption only in D. magna. While D. magna increased O2 consumption under mancozeb exposure, no effects were observed for D. similis. Thus, species may present different responses and sensitivities to different pollutants, regardless of their phylogeny. Therefore, the use of ecotoxicological assays with native species is crucial for a better ecological risk assessment in contaminated areas.

Influence of biochar particle size on biota responses.

Despite the increasing interest for biochar as a soil amendment, a knowledge gap remains on its impacts on non-target soil and aquatic species. We hypothesised that biochar particle size and application rate can play a role in the toxicity to biota. Pine woodchip biochar was incorporated in a clean soil at three particle size classes: small (<0.5 mm), medium (1-2 mm), and large (<4 mm), and at two concentrations: 1% and 6% w/w. A laboratory screening with earthworm Eisenia andrei avoidance behaviour bioassay was carried out to test the most adequate application rates, particle sizes and soil-biochar pre-incubation period. Thereafter, a 28-day greenhouse microcosm experiment was conducted as an ecologically more representative approach. Survival, vertical distribution and weight changes of E. andrei, and bait-lamina consumption were recorded. Soil leachates from the microcosms were collected to evaluate their impact on Daphnia magna immobilisation and Vibrio fischeri (Microtox®) bioluminescence. A feeding experiment with E. andrei was also performed to address earthworm weight changes and to conduct a screening of PAH-type metabolites in their tissue. The 6% <0.5 mm treatment pre-incubated for 96 h induced significant avoidance of the earthworms. Significantly lower bait-lamina consumption was observed in microcosms for the 6% <0.5 mm treatment. Moreover, particle size was a statistically significant factor regarding the loss of weight in the feeding experiment and higher concentration of naphthalene-type metabolites detected in E. andrei tissue, when exposed to <0.5 mm biochar particles. The leachates had no adverse effects on the aquatic species. The results suggest that particles <0.5 mm of pine woodchip biochar can pose sub-lethal effects on soil biota.

Effects of ivermectin on Danio rerio: a multiple endpoint approach: behaviour, weight and subcellular markers.

Ivermectin (IVM) is a broad acting antihelmintic used in various veterinary pharmaceuticals. It has been shown that IVM enters the aquatic compartment and adversely affects organisms including fish. This study is based on the hypothesis that long term exposure to IVM affects fish and thus, the main objective was to assess the chronic effects of 0.25 and 25 µg IVM/L to zebrafish using multiple endpoints representative of several levels of biological organization: weight, behaviour (swimming and feeding) and subcellular markers including biomarkers for oestrogenicity (vitellogenin-VTG), oxidative stress (catalase-CAT and glutathione-S-transferase-GST) and neurotransmission (cholinesterase-ChE). Concentrations as low as 0.25 µg IVM/L disrupted the swimming behaviour, causing fish to spend more time at the bottom of aquaria. Such reduction of the swimming performance affected the feeding ability which is likely responsible for the weight loss. The effects on weight were gender differentiated, being more pronounced in males (0.25 µg IVM/L) than in females (25 µg IVM/L). Fish exposed to 25 µg/L exhibited darker coloration and mild curvature of the spine. No effects on VTG and AChE were observed, but a reduction on CAT and GST levels was observed in fish exposed to 25 µg IVM/L, although these alterations probably only reflect the general condition of the fish which was significantly compromised at this concentration. Despite that predicted environmental concentrations of IVM are below 0.25 µg/L, the behavioural effects may be translated into important ecological impacts, e.g. at predator-prey interactions where fish competitive advantage can be decreased. Future work should address the link between behaviour disruption and population fitness. The current study was based on a one experiment and multiple endpoint (anchored) approach, allowing the results to be integrated and linked towards a mechanistic understanding.

Effects of the Pyrethroid Esfenvalerate on the Oligochaete, Lumbriculus variegatus.

Esfenvalerate is a neurotoxic pyrethroid insecticide widely used for agricultural and residential purposes and is considered toxic to nontarget organisms such as fish and aquatic invertebrates. In this study, we evaluated the toxicity of esfenvalerate on the aquatic oligochaete Lumbriculus variegatus. In the acute test, organisms showed visible signs of stress but no LC50 value could be determined. In the 28-day chronic test, a significant decrease in reproduction was observed with a NOEC value of 0.25 µg/kg and a LOEC value of 2.34 µg/kg. As for biomass per worm, a significant decrease was also observed with a NOEC value of 2.34 µg/kg and a LOEC value of 36.36 µg/kg. Reproductive impairment and reductions in biomass of L. variegatus exposed to environmentally realistic concentrations of esfenvalerate observed in laboratory tests suggests potential deleterious effects of this pyrethroid on oligochaete natural populations.

Potential risk of biochar-amended soil to aquatic systems: an evaluation based on aquatic bioassays.

It is vital to address potential risks to aquatic ecosystems exposed to runoff and leachates from biochar-amended soils, before large scale applications can be considered. So far, there are no established approaches for such an assessment. This study used a battery of bioassays and representative aquatic organisms for assessing the acute toxicity of water-extractable fractions of biochar-amended soil, at reported application rates (80 t ha(-1)). Biochar-amended aqueous soil extracts contained cadmium (Cd), cobalt (Co), chromium (Cr), copper (Cu), manganese (Mn), zinc (Zn), nickel (Ni), lead (Pb), arsenic (As) and mercury (Hg) (Σmetals 96.3 µg l(-1)) as well as the 16 priority PAHs defined by the U.S. Environmental Protection Agency (Σ16PAHs 106 ng l(-1)) at contents in the range of current EU regulations for surface waters. Nevertheless, acute exposure to soil-biochar (SB) extracts resulted in species-specific effects and dose-response patterns. While the bioluminescent marine bacterium Vibrio fischeri was the most sensitive organism to aqueous SB extracts, there were no effects on the growth of the microalgae Pseudokirchneriella subcapitata. In contrast, up to 20 and 25% mobility impairment was obtained for the invertebrate Daphnia magna upon exposure to 50 and 100% SB extract concentrations (respectively). Results suggest that a battery of rapid and cost-effective aquatic bioassays that account for ecological representation can complement analytical characterization of biochar-amended soils and risk assessment approaches for surface and groundwater protection.

Cholinesterase activity on Echinogammarus meridionalis (Pinkster) and Atyaephyra desmarestii (Millet): characterisation and in vivo effects of copper and zinc.

Metals are released into freshwater ecosystems from natural and anthropogenic sources, compromising their structural and functional equilibrium. As early warning tools, cholinesterases (ChEs) are usually used to assess the effects of organophosphate and carbamate pesticides, but are also known to be inhibited by metals. The objectives of this work were to characterise the activity of ChE present in the amphipod Echinogammarus meridionalis and the shrimp Atyaephyra desmarestii and to evaluate the in vivo effects of the metals copper and zinc in their ChE activity. To achieve this, firstly the activity of ChE forms were characterised using different in vitro assays with substrates and selective inhibitors. Then, the in vivo effects of 48 h exposures to increasing concentrations of copper and zinc on ChE activity were determined. The ChE form present in both species was acetylcholinesterase (AChE) since both revealed preference for the acetylthiocholine iodide substrate, total inhibition with eserine, the inhibitor of ChEs, and with 1,5-bis(4-allyldimethylammoniumphenyl)-pentan-3-one dibromide, the specific inhibitor of AChE, and presented insensitivity to iso-OMPA, a specific inhibitor of butyrylcholinesterase. The activity of ChEs was inhibited by zinc exposures in the amphipod species, but was not affected by copper. Exposure to copper and zinc did not affect ChEs activity in the shrimp at the concentrations tested. This work is a relevant contribution as foundation for the use of AChE in freshwater crustaceans in further studies including biomonitoring campaigns in different contamination scenarios.

Water-extractable priority contaminants in LUFA 2.2 soil: back to basics, contextualisation and implications for use as natural standard soil.

The natural LUFA 2.2 standard soil has been extensively used in hazard assessment of soil contaminants, combining representation with ecological relevance for accurate risk evaluation. This study revisited the water-extractable fraction of LUFA 2.2 soil, through consecutive soil wet-dry cycles and discusses implications of use as standard substrate in derivation of ecotoxicological data and toxicity thresholds. Potentially bioavailable contents of metals (177.9-888.7 µg/l) and the 16 polycyclic aromatic hydrocarbons (PAHs; 0.064-0.073 µg/l) were dependent on the number of soil wetting-drying cycles applied. Such contents were screened based on current EU guidelines for surface waters and reported toxicological benchmarks for aquatic organisms. Aqueous concentrations generally fit within recommended Environmental Quality Standards (EQS), except for Hg (0.13-0.22 µg/l; >Maximum Allowable Concentration-MAC-of 0.07 µg/l) and for the sum of benzo(g,h,i)perylene and indeno(1,2,3-cd)pyrene (0.005 µg/l; >double the Annual Average of 0.002 µg/l). Further, aqueous As, Zn, Cd, Ni and Cr concentrations exceeded 'lower benchmark' values for aquatic organisms, possibly reflecting an inadequate derivation for ecotoxicological data. In turn, PAHs in LUFA 2.2 soil aqueous extracts, whilst individually, are not likely to constitute a hazard to test biota exposed to its aqueous fractions. This study urges for potentially bioavailable fractions of reference and standard natural soils to be adequately characterized and addressed as part of the research aim, experimental approach and design, as well as the expected scope of the outcomes.

Pesticide residue analyses and biomarker responses of native Costa Rican fish of the Poeciliidae and Cichlidae families to assess environmental impacts of pesticides in Palo Verde National Park.

Pesticide chemical residues in water samples and biomarker responses in transplanted fish were used to monitor environmental hazards of pesticides in Palo Verde National Park (Costa Rica). The Costarican fish, Parachromis dovii (Ciclhidae) and Poecilia gillii (Poecillidae), were selected as sentinel species. Contaminant analyses detected up to 15 different pesticide residues in water with hexachlobenzene (2261 ng l(-1)), phorate (473 ng l(-1)), epoxiconazole (314) and bromacil (117 ng l(-1)) being the compounds found in higher concentrations. Biomarker responses evidenced impacts on cholinesterase activities in transplanted fish at Barbudal site probably due to the presence of organophosphate insecticides such as phorate. High enzyme activities of glutathione S-transferase and catalase and elevated levels of lipid peroxides were also observed at a site impacted by rice fields (Cabuyo); those effects could be associated with the presence of hexachloro benzene and triazole fungicides. In general, P. dovii biomarkers were affected to a greater extent than those of P. gillii in fish transplanted to sites associated with agriculture, which suggests the former species is a good candidate for future surveys.

Soil habitat function after innovative nanoagriproducts application: Effect of ageing on the avoidance behaviour of the soil invertebrates Enchytraeus crypticus and Folsomia candida.

Research on nanotechnology with applications in agriculture has been gathering attention because it may achieve a good balance between agricultural production and environmental integrity. Among the vast nanomaterials, layered double hydroxides (LDHs) are a promising solution for supplying crops with macro- and/or micronutrients. Still, little is known about their safety implications for non-target organisms, such as soil invertebrates. The habitat function of soils might be impacted by potential stressors, which can be assessed through avoidance behaviour tests. This study aimed to assess the effect of two innovative agriproducts, Zn-Al-NO3 LDH and Mg-Al-NO3 LDH, on the avoidance behaviour of the enchytraeid Enchytraeus crypticus and the collembolan Folsomia candida, over time. Simultaneously, Zn and Mg potential release from LDHs to soil was evaluated. Overall, the behaviour of soil invertebrates differed between species, with enchytraeids being more sensitive to LDHs-treated soils than collembolans, possibly explained by their different physiological traits. The behaviour of soil organisms also depended on the LDH structural composition and was time-variable. Soil treated with Zn-Al-NO3 LDH was perceived as less favourable compared to Mg-Al-NO3 LDH, which was preferred to clean soil at most tested concentrations. LDHs toxicity was partly, but not exclusively, related to Zn and Mg release. Cations release over time was demonstrated in the chemical assessment. Still, LDHs toxicity to soil invertebrates decreased as increasing AC50 values were derived over time. Slower dissolution over time might explain the decrease in toxicity. Our study demonstrates that both soil invertebrates could sense LDHs in soil and eventually adapt their behaviour by avoiding or preferring, according to the type and level of LDH present.

Boric acid as reference substance: pros, cons and standardization.

Boric acid (BA) has been successfully used as reference substance in some standard test guidelines. Due to the fact that previously selected reference substances present a significant risk to human health and/or are banned for environmental reasons, BA is being discussed for broader adoption in OECD or ISO guidelines. To provide input on BA data and contribute to the discussion on its suitability as a reference substance, in the present study BA was tested with two standard soil organisms, Enchytraeus albidus and Folsomia candida, in terms of survival, reproduction and avoidance. Additionally, published data on other organisms was analysed to derive the most sensitive soil dwelling invertebrate (hazard concentration-HC5). Results showed that BA affected the tested organisms, being two times more toxic for collembolans (LC50 = 96; EC50 = 54 mg/kg) than for enchytraeids (LC50 = 325; EC50 = 104 mg/kg). No avoidance behaviour occurred despite the fact that BA affects earthworms. Actually, it is the recommended reference substance for the earthworm avoidance test. Clearly, the suitable performance of BA in one species should not be generalized to other species. Absolute toxicity is not an important criterion for the selection of a reference substance, but it has been proposed that effects should occur within a reasonable range, i.e. <1,000 mg/kg. We could confirm, compiling previous data that for most soil invertebrates, the EC50 is expected to be below 1,000 mg/kg. From these data it could be derived that the most sensitive soil dwelling invertebrate (HC5, 50%) is likely to be affected (EC10) at 28 (8-53) mg H(3)BO(3)/Kg, equivalent to 4.6 (1.4-8.7) mg boron/kg.

Toxicity and genotoxicity of organic and inorganic nanoparticles to the bacteria Vibrio fischeri and Salmonella typhimurium.

The present work aimed at evaluating the toxicity and genotoxicity of two organic (vesicles composed of sodium dodecyl sulphate/didodecyl dimethylammonium bromide-SDS/DDAB and of monoolein and sodium oelate-Mo/NaO) and four inorganic (titanium oxide-TiO2, silicon titanium-TiSiO4, Lumidot-CdSe/ZnS, and gold nanorods) nanoparticles (NP), suspended in two aqueous media (Milli Q water and American Society for Testing and Materials (ASTM) hardwater), to the bacteria Vibrio fischeri (Microtox test) and Salmonella typhimurium-his⁻ (Ames test with strains TA98 and TA100). Aiming a better understanding of these biological responses physical and chemical characterization of the studied NP suspensions was carried out. Results denoted a high aggregation state of the NP in the aqueous suspensions, with the exception of SDS/DDAB and Mo/NaO vesicles, and of nanogold suspended in Milli Q water. This higher aggregation was consistent with the low values of zeta potential, revealing the instability of the suspensions. Regarding toxicity data, except for nano TiO2, the tested NP significantly inhibited bioluminescence of V. fischeri. Genotoxic effects were only induced by SDS/DDAB and TiO2 for the strain TA98. A wide range of toxicity responses was observed for the six tested NP, differing by more than 5 orders of magnitude, and suggesting different modes of action of the tested NP.

Caffeine as a contaminant of concern: A review on concentrations and impacts in marine coastal systems.

Caffeine has been identified as emerging contaminant of concern due to its widespread occurrence in the aquatic environment and potential to be biologically active. Recently, these concerns have been translated in an increasing research on its occurrence and effects on biota. However, there is still a limited knowledge on seawater matrices and the implications of caffeine presence in coastal and marine ecosystems are not fully known. The present review aims to fill these knowledge gaps, analysing the existing literature regarding the occurrence, effects and potential risks of caffeine residues to coastal ecosystems, contributing to the risk assessment of this psychoactive drug in the aquatic environment. The analysed literature reported caffeine concentrations in the coastal ecosystems, raising high concerns about the potential adverse impacts on the ecological safety and human health. Caffeine has been found in tissues from coastal and marine biota including microalgae, coral reefs, bivalves and fish due to bioaccumulation after chronic, long-term exposures in a contaminated environment. Additionally, caffeine residues had been demonstrated to have adverse impacts on aquatic organisms, at environmentally realistic concentrations, inducing oxidative stress and lipid peroxidation, neurotoxicity, changing energy reserves and metabolic activity, affecting reproduction and development and, in some cases, causing mortality. Considering the increasing adverse impacts of caffeine pollution in the coastal environment, this review highlights the urgent need to minimize the increasing load of caffeine to the aquatic ecosystems; being imperative the implementation of scientific programs and projects to classify effectively the caffeine as a high-priority environmentally hazardous emerging pollutant.

Single and combined effects of ultraviolet radiation and triclosan during the metamorphosis of Solea senegalensis.

Ultraviolet radiation (UV) and triclosan (TCS) affect the early development of marine fish; however, the corresponding molecular mechanisms are still not fully understood. Therefore, this work aims to study the effects of the single and combined exposure to these stressors during the thyroid-regulated metamorphosis of the flatfish Solea senegalensis. Sub-lethal exposure (5.89 kJ m-2 UV and/or 0.546 and 1.090 mg L-1 TCS for 48 h) was performed at the beginning of metamorphosis (13 days after hatching, dah), followed by a period in clean media until complete metamorphosis (24 dah). Malformations, metamorphosis progression, length, behavior and the expression of thyroid axis-related genes were studied. TCS induced malformations, decreased swimming performance, and induced metamorphosis acceleration at 15 dah, followed by a significant metamorphosis delay. Such effects were more noticeable in the presence of UV. The down-regulation of five thyroid axis-related genes occurred after exposure to TCS (15 dah), and after 9 days in clean media two genes were still down-regulated. UV exposure increased the effect of TCS by further down-regulating gene expression immediately after the exposure. Since several effects persisted after the period in clean media, implications of these stressors (mainly TCS) on the ecological performance of the species are suggested.

Will extreme weather events influence the toxic impacts of caffeine in coastal systems? Comparison between two widely used bioindicator species.

In the recent years, marine heatwaves (MHWs) have caused devastating impacts on marine life. The understanding of the combined effects of these extreme events and anthropogenic pollution is a vital challenge. In particular, the combined effect of MHWs on the toxicity of pharmaceuticals to aquatic life remains unclear. To contribute to these issues, the main goal of the present investigation was to evaluate how MHWs may increase caffeine (CAF) toxicity on the clam Ruditapes philippinarum and the mussel Mytilus galloprovincialis. Bioaccumulation levels and changes on oxidative stress, metabolic capacity and neurotoxic status related biomarkers were investigated. The obtained results revealed the absence of CAF accumulation in both species. However, the used contaminant generated in both bivalve species alteration on neurotransmission, detoxification mechanisms induction as well as cellular damage. The increase of antioxidant defence mechanisms was complemented by an increase of metabolic activity and decrease of energy reserves. The obtained results seemed magnified under a simulated MHWs, suggesting to a climate-induced toxicant sensitivities' response. On this perspective, understanding of how toxicological mechanisms interact with climate-induced stressors will provide a solid platform to improve effect assessments for both humans and wildlife.

Effects of imidacloprid exposure on Chironomus riparius Meigen larvae: linking acetylcholinesterase activity to behaviour.

Imidacloprid (IMI) is an insecticide that interferes with the transmission of stimuli in the nervous system of insects. It is neurotoxic by mimicking nicotine through its binding to the nicotinic acetylcholine receptor. In this work, experiments comprising 96 h exposure followed by 48 h in clean medium were conducted to evaluate the toxicity of IMI to Chironomus riparius and its potential recovery. Behavioural parameters and AChE activity were assessed. After 96 h exposure to IMI, AChE activity, and the behaviour parameters ventilation and locomotion were reduced. There were no signs of recovery after removal to clean water for 48 h. Ventilation behaviour was the most sensitive parameter and the one with the highest correlation to AChE activity. Despite the possibility that IMI might be having an indirect effect on AChE activity, the behavioural endpoint showed a higher sensitivity than the biochemical response itself. This work highlights the importance of linking parameters with ecological relevance at individual level (behavioural parameters) with biochemical responses, to unravel xenobiotics mode of action.

Biochemical characterization of cholinesterases in Enchytraeus albidus and assessment of in vivo and in vitro effects of different soil properties, copper and phenmedipham.

Enchytraeus albidus are important organisms of the soil biocenosis, used as standard test species in environmental risk assessment. The inhibition of cholinesterases (ChE) activity of several species has been widely used to assess the exposure and effects of anti-cholinesterase environmental contaminants. Several studies have shown the association between ChE activity inhibition and adverse effects on behaviour and survival. Extensive studies addressing survival and behavioural endpoints, as well as other biomarkers, have been done in E. albidus with different types of soil contaminants. The main objectives of this study were: (1) to characterize biochemically the ChE present in the soluble post-mitochondrial fraction of E. albidus whole body homogenates, using different substrates and selective inhibitors; (2) to assess the in vivo effects of copper, phenmedipham and different soil properties (pH, organic matter, clay) on the ChE activity; (3) to assess the in vitro effects of copper and phenmedipham on the ChE activity. The results suggest the presence of one ChE in the soluble post-mitochondrial fraction of E. albidus whole body homogenates, which displays properties of both acetylcholinesterase and pseudocholinesterase considering the typical mammalian enzymes. It is also shown that ChE activity is not inhibited by exposure to different soil properties and that copper and phenmedipham inhibited ChE activity both in in vivo and in in vitro conditions and therefore ChE inhibition seems to be a robust biomarker for this herbicide and this heavy metal. This study showed that ChE activity in E. albidus might be correlated to previously determined higher level effects like survival and reproduction, as well as avoidance behaviour.