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Exposure of aquatic environments to emerging contaminants is a global issue, special relevant in many estuaries due to impacts from anthropogenic activity. The aim of this work was to evaluate thirty-seven endocrine disruptor chemicals (EDCs) from four different classes (pesticides, bisphenols, polycyclic musks and UV-filters) in water and sediment samples collected during one-year in the estuaries of Tagus and Douro Rivers located into the NE Atlantic Ocean coast. EDCs analysis was achieved afterward validation of a gas-chromatography mass spectrometry (GC-MS) method using Dispersive Liquid-Liquid Microextraction (DLLME) as extraction procedure for water samples, and Quick, Easy, Cheap, Efficient, Rugged and Safe (QuEChERS) combined with DLLME for sediments. Tagus estuary presented higher levels of contamination with pesticide residues and bisphenols (BPs) than the Douro estuary in both water and sediment samples. Contrariwise, levels and frequency of polycyclic musks (PCMs) and UV-filters (UVF) were slightly higher in Douro estuary. Levels of pesticide residues in both sediment and water samples, and levels of PCMs and UVF in water samples were higher in warmer seasons (summer and spring) than in colder ones (winter and autumn). The opposite was found in what respect levels of BPs in water and sediment samples, and PCMs and UVF levels in sediment samples. Although the levels found for each contaminant are low, usually in the order of a few ng/mL(g), the presence of a high number of toxic compounds is a source of concern and requires constant monitoring., Competing Interests: Declaration of competing interest The authors declare that they have no known competing financial interests or personal relationships that could have appeared to influence the work reported in this paper., (Copyright © 2022 Elsevier B.V. All rights reserved.)

Especially added on many industrial and domestic products as flame retardants (FRs), polybrominated diphenyl ethers (PBDEs) are among the chemicals of high environmental concern because of their potential harmfulness for environmental and human health. Seafood consumption is considered the main source of PBDEs and their methoxylated congeners (MeO-BDEs) for humans. The present study aims to investigate the seasonal occurrence of six PBDEs and eight MeO-BDEs congeners using Douro river biota (different trophic levels) as sentinels, as well as to evaluate the human exposure risk to PBDEs through seafood consumption. Biota samples (n = 273) were collected from one of the most important Portuguese estuaries in the north-western coast of Portugal at four different seasons (2019-2020). The analyses were performed by an environmental-friendly extraction procedure followed by Gas Chromatography coupled to a triple quadrupole detector (GC-MS/MS). PBDEs were detected in all seafood samples analysed, with means ranging from 0.02 ng g -1 ww (flounder in autumn) to 3.75 ng g -1 ww (mussel in winter). Levels of lower-brominated PBDE congeners were significantly higher than higher-brominated ones in all seasons (p < 0.01). MeO-BDEs ranged from 0.001 ng g -1 ww (grey mullet in summer) to 5.66 ng g -1 ww (green crab in spring). Crabs and mussels presented the highest means of PBDEs and MeO-BDEs. Regarding the health risk assessment of the studied PBDE congeners (47, 99, and 153), consumption of Douro river fish is not a case of concern for consumers., Competing Interests: Declaration of competing interest The authors declare that they have no known competing financial interests or personal relationships that could have appeared to influence the work reported in this paper., (Copyright © 2021 Elsevier B.V. All rights reserved.)

This study documented seasonal levels of microplastics (MPs) and biomarkers (condition index, neurotoxicity, energy, oxidative stress) in mussels (Mytilus galloprovincialis), and water physico-chemical parameters in the Douro estuary (NE Atlantic coast), and estimated the human risk of MP intake (HRI) through mussels. Mussel stress was determined through the Integrated Biomarker Response (IBR). HRI was estimated from mussel MP concentrations and consumer habits. MPs were mainly micro-fibres (72 %) with varied chemical composition. Seasonal MP means (±SEM) in mussels ranged from 0.111 ± 0.044 (spring) to 0.312 ± 0.092 MPs/g (summer). Seasonal variations of mussel stress (IBR: 1.4 spring to 9.7 summer) and MP concentrations were not related. MeO-BDEs, PBDEs, temperature, salinity and other factors likely contributed to mussel stress variation. HRI ranged from 2438 to 2650 MPs/year. Compared to the literature, MP contamination in mussels is low, as well as the human risk of MP intake through their consumption. • Microplastic (MP) concentrations and biomarkers were studied in marine mussels. • Seasonal means (0.111 ± 0.044 to 0.312 ± 0.092 MPs/g) indicate low MP contamination. • Neurotoxicity and oxidative and energy stresses were observed (mainly in summer). • Seasonal variation trends of MP concentrations and stress levels were not related. • Human risk of MP intake through the consumption of Douro estuary mussels is low. [ABSTRACT FROM AUTHOR]

The health status of eels (Anguilla anguilla) developing in three estuaries of the NW Portuguese coast with different types and levels of pollution was compared in relation to morphometric parameters, Fulton condition index (F index) and several biomarkers. Relatively to the reference population, glass eels from the Lima estuary had lower weight and length, cholinesterase (ChE) and lactate dehydrogenase (LDH) inhibition, and lower levels of some anti-oxidant parameters, while yellow eels also showed a decreased F index, and increased Na+/K+-ATPase and lipid peroxidation (LPO) levels. Relatively to the reference population, glass eels from the Douro estuary had increased Na+/K+-ATPase and glutathione-S-transferase activities and LDH inhibition, while yellow eels also had ChE inhibition and increased LPO. Overall, these results indicate that eels from polluted estuaries showed a poor health status than those from a reference estuary, and that adverse effects become more pronounced after spending several years in polluted estuaries. [Copyright &y& Elsevier]

The study presented here searched for the level of taxonomic resolution required to detect the effects of low-level chronic pollution on estuarine meiobenthic communities. Meiofauna from two sites, with special attention to harpacticoid copepods, was analysed at different taxonomic levels of aggregation using uni- and multivariate methods. Adaptation processes that could buffer biodiversity disruptions were also considered through the analysis of fitness-related and tolerance traits in the harpacticoid copepod Paronychocamptus nanus. Results showed that uni- and multivariate analyses could be inadequate when assessing subtle anthropogenic contamination. Instead, the assessment of inter-population differences in tolerance to the main source of stress rises as a required procedure if potential effects of this type of contamination are being investigated. Specifically, a 96 h acute toxicity test performed with populations from the affected site appears as a faster and reliable general tool to assess impacts of low-level chronic pollution in estuaries. [Copyright &y& Elsevier]

The aims of the present study were to compare the health status of yellow eels (Anguilla anguilla) developing in three estuaries of the NW Portuguese coast with different levels of pollution and their physiological responses to combined effects of environmental variation and pollution. For this, a field study was performed using a multi-parameter approach, including eels condition indexes and biomarkers, water quality variables and other environmental factors. Sixteen biological parameters were assessed, namely: hepatosomatic index (LSI), Fulton's condition index (K), lipid peroxidation (LPO), total glutathione (TG), reduced glutathione (GSH), oxidised glutathione (GSSG), GSH/GSSG, and the activity of the enzymes acetylcholinesterase (AChE), lactate dehydrogenase (LDH), sodium-potassium ATPase (Na?/ K?-ATPase), ethoxyresorufin O-deethylase (EROD), glutathione S-transferases (GST), catalase (CAT), superoxide dismutase (SOD), glutathione peroxidase (GPx) and glutathione reductase (GR). Ten environmental factors were also measured in water: temperature, salinity, pH, phosphates, nitrates, nitrites, ammonium, silica, phenol and hardness. Globally, the biomarkers indicate exposure and toxic effects of pollutants on eels living in contaminated estuaries. The relationships between biological and environmental variables were assessed through redundancy analysis. K and LSI indexes, AChE and Na?/K?-ATPase, total glutathione levels and the antioxidant enzymes CAT, GR, and SOD where the factors most discriminating reference (Minho River estuary) from contaminated estuaries (Lima and Douro Rivers estuaries). Moreover, the most striking outcomes of pollutants exposure on biological responses were observed during winter, probably due to a joint effect of cold weather and pollution stress. Altogether, the results indicate that the development of eels in the polluted estuaries of Lima and Douro rivers is interfering with physiological functions determinant for their survival and performance. This may increase the mortality rates during the continental life-phase of the species and decrease the percentage of animals able to successfully complete their oceanic migration and, thus, reduce the contribution of each generation to the next one. [ABSTRACT FROM AUTHOR]

Wild fish (Cyprinus carpio, Mugil cephalus, Platichthys flesus) from an estuary of the NE Atlantic coast were investigated for plastic contamination (N = 128). From the 1289 particles recovered from fish samples, 883 were plastics. Among these, 84% were fibres and 97% were microplastics. Thirty-six polymers were identified. The number of microplastics (mean ± SD) per individual fish (MP/fish) was 8 ± 6 in C. carpio , 10 ± 9 in M. cephalus and 2 ± 2 in P. flesus. The means of MP/fish per body site were 6 ± 7 in gastrointestinal tract, 0.5 ± 1.1 in gills, 0.3 ± 0.7 in liver and 0.6 ± 1.2 in muscle samples. A few large fibres in liver (≤ 4841 μm) and muscle (≤ 5810 μm) samples were found. The results evidence the existence of high fish contamination by microplastics and reinforce the need of further research on plastic pollution in estuaries. [Display omitted] • Fish (flounders, mullets, carps) were studied for plastic (PL) contamination. • 883 PL were recovered, of which 97% were microplastics (MP). • 94% of fish had PL with a mean of 8 ± 8 PL per fish. • PL in the gastrointestinal tract, gills & muscle, and MP in the liver were found. • Fibres had size ≤ 4841 μm in the liver, and ≤ 5810 μm in the dorsal muscle. [ABSTRACT FROM AUTHOR]

Given the urgent need for ecologically relevant toxicity tools specific to estuarine systems, the main goal of the present study was to develop and evaluate the potential of an in situ algal assay for estuarine sediment-overlying water toxicity evaluations using the microalga Phaeodactylum tricornutum immobilized in alginate beads. With its simple and cost-effective design, the assay apparatus, coupled with the use of microalgae immobilized in alginate beads, allowed rapid and easy deployment as well as retrieval of assay chambers. The effectiveness of the proposed in situ assay was investigated by deploying it at two reference and six contaminated Portuguese estuaries. By deploying open chambers both with and without an artificial nutrient source and closed chambers with an artificial seawater medium, it was possible to discriminate toxic effects from those caused by differences in temperature, light, and nutrient conditions across sites. A reduction in algal growth was found at contaminated sites relative to reference sites. The strontium alginate beads used for microalgae immobilization were found to be appropriate to perform in situ assays, showing no signs of disruption or dissolution after up to 8 d of exposure in estuarine sediment-overlying waters. The proposed in situ assay was shown to be a useful tool for estuarine sediment-overlying water toxicity testing to provide relevant data concerning primary production to estuarine ecological risk assessment studies. [ABSTRACT FROM AUTHOR]

This study evaluated a short-term sublethal endpoint for in situ toxicity assays for estuarine sediments, based on postexposure feeding of the polychaete Hediste (Nereis) diversicolor Müller. A method for precisely quantifying postexposure feeding rates of H. diversicolor was established under laboratory conditions using Artemia franciscana Kellog nauplii. The sensitivity of the postexposure feeding response to copper was investigated by comparing postexposure feeding rates to growth and lethality. The 48-h and 96-h median lethal concentration (LC50) of copper were 241 and 125 µg/L, respectively, whereas the 48-h median inhibitory concentration (IC50) for postexposure feeding and the 20-d IC50 for growth were 52 and 25 µg/L of copper, respectively. The influence of different exposure conditions (substrate, temperature, salinity, food availability, and light) on H. diversicolor postexposure feeding was assessed; temperature and salinity were found to influence significantly postexposure feeding. The effectiveness of the proposed in situ assay was investigated by deploying it at two reference and six contaminated Portuguese estuaries. A 48-h exposure period was followed by a 1-h postexposure feeding period. High organism recoveries (89-100%) were obtained. Postexposure feeding was depressed significantly (17-90%) at all contaminated sites relatively to reference sites. The proposed in situ assay with H. diversicolor was shown to be a potential useful tool for estuarine sediment toxicity testing. [ABSTRACT FROM AUTHOR]

Abstract: Biochemical biomarkers in common estuarine species, such as the brown shrimp Crangon crangon, have the potential to provide early warning of contaminant exposure from field collected samples and through the development of in situ tests. The biomarkers acetylcholinesterase (AChE), lactate dehydrogenase (LDH) and glutathione S-transferase (GST) have been shown to provide evidence of exposure to contaminants in a number of species and field situations. As they may naturally respond to the marked physicochemical changes found in estuaries (thus confounding contaminant-induced effects), this work aims to determine the effects of salinity, temperature and handling stressors on these biomarkers in C. crangon. AChE recovery in field-collected shrimp transplanted to clean laboratory conditions suggests the presence of inhibiting factors at the sampling site (River Minho estuary). Maintenance time in stock tanks had effects that led to the choice of a minimum 15-day maintenance period of C. crangon in the laboratory before subsequent use of the enzymes as effect criterions in toxicity assays. Field levels of biomarker activity were unaffected following field–laboratory transportation of C. crangon, making this factor unlikely to jeopardize detection of contaminant associated effects. LDH levels were significantly lower under conditions that mimic a diurnal salinity fluctuation, increasing under low salinity conditions; this potentially indicates increased energy costs associated with raised osmoregulatory demands. It is recommended that a lower limit to field exposure of in situ tests should be based on salinity. Higher temperatures led to higher AChE activities and this is in agreement with the existing evidence of increases of endogenous AChE levels as a function of temperature (within a certain range). To avoid misinterpretation of biomarker responses, studies such as this are an important contribution to the establishment of reference activity levels against which biomarker changes can be estimated and are therefore essential preliminary steps in the development of in situ bioassays using biomarkers. [Copyright &y& Elsevier]

This work was focused on evaluating the occurrence of endocrine-disrupting compounds (EDCs) in fish muscles, such as bisphenol analogues, personal care products, including some UV-filters and musk fragrances, and selected pesticides. A total of 238 muscle samples of three fish species (Platichthys flesus , Mugil cephalus, and Dicentrarchus labrax) of an important estuary ending in the North East Atlantic Ocean (Douro River estuary, Portugal), in different seasons of the year, was analysed. M. cephalus was the species with a greater occurrence of contaminants, mainly in fishes collected during the spring. Bisphenol A was the main bisphenol analogue detected with an average content of 23.1 μg/kg wet weight (w.w.). Among chemicals used in personal care products, galaxolide, tonalide, and 2-ethylhexyl salicylate, were the most often found, being present in 15.9% (2.2–538.4 μg/kg w. w.), 4.6% (1.1–57.8 μg/kg w. w.) and 3.4% (3.9–56.1 μg/kg w. w.) of the samples, respectively. About 14% of the samples contained residues of at least one pesticide, being alachlor, aldrin, p,p' -DDT, permethrin, and prochloraz the main chemicals observed, varying from 0.1 μg/kg w. w. (p,p' -DDT) to 37.8 μg/kg w. w. (prochloraz). The daily intake estimates of the most frequent EDCs found in the fish muscles suggested that there are no health concerns, based on the recommended weekly consumption of fish for adults. • Levels of EDCs in fishes collected during one year in the Douro River estuary. • About 25% of the samples (n = 238) contained at least one endocrine disruptor. • Galaxolide, tonalide, and p,p' -DDT were frequently detected in fish muscles. • There are no health concerns at the dietary exposure levels estimated for the EDCs. [ABSTRACT FROM AUTHOR]

Abstract: Pisidium amnicum is an important indigenous species in the freshwater tidal portion of the River Minho estuary and until the introduction of the non-indigenous invasive species Corbicula fluminea was the most abundant bivalve in this ecosystem. Between January 2005 and August 2006 monthly samples were collected in three sites to study the abundance, biomass, growth and production of P. amnicum. Results indicate that P. amnicum has continuous growth throughout its life span and growth rates were higher during the early phases of development, coincident with the spring and summer months. Life span estimated was 24 months. The annual 2005 growth production of P. amnicum estimated was 2.339g AFDWm−2 year−1, and the mean annual biomass was 1.594g AFDWm−2, resulting in a ratio of 1.47year−1 and a turnover time of 248.7 days. The annual 2005 elimination production estimated was 7.541g AFDWm−2 year−1, resulting in an ratio of 4.73year−1. It is concluded that P. amnicum continues to play an important role in the food web in areas where it coexists with C. fluminea. Previous studies showed declines in the indigenous population after the introduction of C. fluminea and currently, areas of high abundance are restricted to small patches in the upper limit of the tidal influence. Consequently, P. amnicum is now under considerable pressure and this situation requires ecological and conservational attention. [Copyright &y& Elsevier]

Abstract: The identification of different species inside the Corbicula genus is complicated due to the high variation of shell shape, colour and sculpture of the individuals. The species Corbicula fluminea has been present in the River Minho estuary (NW Portugal) at least since 1989. More recently, individuals of the same genus colonized an adjacent estuary (River Lima estuary). Although appearing also to be C. fluminea, the individuals of the Lima estuary differ from those of Minho estuary in the colour and shape of the shell. Therefore, the two populations were compared by conventional morphometric measures (shell length, width and height), geometric morphometric methods (landmarks analysis using the interior of the shell) and genetic analysis (based on the mitochondrial cytochrome c oxidase subunit I gene sequence). Genetic analysis showed an identical mtCOI sequence indicating that both populations belong to the species C. fluminea. However, results of conventional and geometric morphometric analysis showed significant differences in shell shape between individuals from the two populations. These differences may be due to (a) phenotypical plasticity in response to different environmental and/or ecological conditions existing in the two estuaries, (b) different origins of the populations and/or distinct routes until reaching the two estuaries and (c) inter-population genetic differences caused by processes occurring after the introduction of the species in the two estuaries (e.g. differential selection). [Copyright &y& Elsevier]

Abstract: This study aimed at integrating postexposure feeding and some biochemical parameters in the responses of the estuarine polychaeta, Hediste diversicolor, to controlled laboratory exposure conditions and to in situ exposures scenario of sediment contamination. Since H. diversicolor feeding may be considered as a major rate-limiting step in the processing of detritus in European estuaries, a reduction in feeding activity may have implications not only at the individual and population level of the species but also in detritus processing and in organic matter decomposition rates at the ecosystem level. The biochemical parameters were chosen as indicators of four key physiological functions: neurotransmission, metabolic condition, detoxification processes and antioxidant defences. The Mira and Sado estuaries, located in the Southwest coast of Portugal and classified as undisturbed and impacted, respectively, were selected as sites for this study. A significant depression in H. diversicolor postexposure feeding (from 30 to 70%) was consistently detected in all impacted sediments, supporting the sensitivity and responsiveness of feeding as a sublethal toxicity endpoint. Alongside with a reduced energy intake, an increased rate of organisms’ anaerobic metabolism, as evidenced by an enhancement of lactate dehydrogenase activity (up to 1.5-fold), suggested a rapid need of additional energy to ameliorate chemical stress. Moreover, oxidative stress was shown to be an important mechanism of toxicity of the impacted sediments in H. diversicolor, as evidenced by a marked reduction in the glutathione redox status (up to 6.5-fold) and an increase in lipid peroxides levels (up to 2.3-fold) in organisms exposed to the most impacted sediments. Results of the in situ assay, conducted to assess the ecological relevance of sediment laboratory toxicity estimates and their application to make valid field extrapolations, revealed a lack of agreement in the response of catalase in organisms exposed to moderate impacted sediments. Our results support the utility of integrating responses at individual and sub-individual level to evaluate potential toxicant-induced changes in key physiological functions of H. diversicolor and to interpret their potential ecological consequences. [Copyright &y& Elsevier]