Fate, subcellular distribution and biological effects of rare earth elements in a freshwater bivalve under complex exposure.

Rare earth elements (REE) are emerging contaminants due to their increased use in diverse applications including cutting-edge and green-technologies. Their environmental concerns and contradicting results concerning their biological effects require an extensive understanding of REE ecotoxicology. Thus, we have studied the fate, bioaccumulation and biological effects of three representative REE, neodymium (Nd), gadolinium (Gd) and ytterbium (Yb), individually and in mixture, using the freshwater bivalve Corbicula fluminea. The organisms were exposed for 96 h at 1 mg L-1 REE in the absence and presence of dissolved organic matter (DOM) reproducing an environmental contamination. Combined analysis of the fate, distribution and effects of REE at tissue and subcellular levels allowed a comprehensive understanding of their behaviour, which would help improving their environmental risk assessment. The bivalves accumulated significant concentrations of Nd, Gd and Yb, which were decreased in the presence of DOM likely due to the formation of REE-DOM complexes that reduced REE bioavailability. The accumulation of Nd, Gd and Yb differed between tissues, with gills > digestive gland ≥ rest of soft tissues > hemolymph. In the gills and in the digestive gland, Nd, Gd and Yb were mostly (>90 %) distributed among metal sensitive organelles, cellular debris and detoxified metal-rich granules. Gadolinium, Yb and especially Nd decreased lysosome size in the digestive gland and disturbed osmo- and iono-regulation of C. fluminea by decreasing Na concentrations in the hemolymph and Ca2+ ATPase activity in the gills. Individual and mixed Nd, Gd and Yb exhibited numerous similarities and some differences in terms of fate, accumulation and biological effects, possibly because they have common abiotic and biotic ligands but different affinities for the latter. In most cases, individual and mixed effects of Nd, Gd, Yb were similar suggesting that additivity approach is suitable for the environmental risk assessment of REE mixtures.

Spatial and seasonal use of biomarkers in dreissenids: implications for biomonitoring.

In addition to pollution, organisms are exposed to natural variations of the biotic and abiotic factors of their environment. A battery of sub-cellular biomarkers has been measured seasonally in several populations of both Dreissena polymorpha and Dreissena rostriformis bugensis. To observe and understand the variability associated with biomarker responses, water physicochemistry, sediment contamination, and internal concentrations of contaminants in soft tissues were also considered. Results evidenced seasonal, inter-specific, and inter-populational variability of the measured responses, highlighting the needs (1) to acquire long-term data on the studied populations and (2) to incorporate environmental parameters and contamination in the interpretation of biological responses. From a biomonitoring perspective, significant relationships were identified between biomarkers, internal concentrations of contaminants in soft tissues, and sediment contamination in D. r. bugensis and, to a lesser extent, in D. polymorpha. The detailed interpretation of each biomarker of the battery measured is complex, but a global analysis of all biomarkers at once allows to obtain this signature of the contamination of the studied sites.

Assessing the effects of silver nanoparticles on the ecophysiology of Gammarus roeseli.

Being part of the macrobenthic fauna, gammarids are efficient indicators of contamination of aquatic ecosystems by nanoparticles that are likely to sediment on the bottom. The present study investigates the effects of silver nanoparticles (nAg) on ecophysiological functions in Gammarus roeseli by using a realistic scenario of contamination. Indeed, an experiment was conducted during 72 h, assessing the effects of 5 silver nAg from 10 to 100 nm diluted at concentrations of maximum 5 µg L-1 in a natural water retrieved from a stream and supplemented with food. The measured endpoints in gammarids were survival, silver concentrations in tissues, consumption of oxygen and ventilation of gills. Additionally, a set of biomarkers of the energetic metabolism was measured. After a 72-h exposure, results showed a concentration-dependent increase of silver levels in G. roeseli that was significant for the smallest nAg size (10 nm). Ecophysiological responses in G. roeseli were affected and the most striking effect was a concentration-dependent increase in oxygen consumption especially for the smallest nAg (10 to 40 nm), whereas ventilation of gills by gammarids was not changed. The potential mechanisms underlying these findings are discussed. Thus, we demonstrated the very low exposure concentration of 0.5 µg L-1 for the small nAg size led to significant ecophysiological effects reinforcing the need to further investigate subtle effects on nanoparticles on aquatic organisms.

Implications of speciation on rare earth element toxicity: A focus on organic matter influence in Daphnia magna standard test.

Rare earth elements (REE) have become essential in high- and green-technologies. Their increasing use lead to the release of anthropogenic REE into the environment including aquatic systems. The limited data available on the aquatic ecotoxicology of REE indicate their biological effects are highly dependent on their speciation, posing challenges for a reliable environmental risk assessment (ERA). The current study assessed the influence of speciation on the toxicity of neodymium (Nd), gadolinium (Gd) and ytterbium (Yb) in the Daphnia magna mobility inhibition test (ISO 6341:2012). REE toxicity was assessed individually and in ternary mixture, in the absence and presence of dissolved organic matter (DOM). Speciation was predicted by modeling and REE bioaccumulation by D. magna was measured to better understand the relationship between REE speciation and toxicity. DOM decreased significantly the toxicity of Nd, Gd and the mixture towards this freshwater crustacean. This was explained by a lower REE bioaccumulation in the presence of DOM due to REE-DOM complexation, which reduced REE bioavailability. DOM effects on Yb toxicity and bioaccumulation were limited because of Yb precipitation. We show that the way of expressing EC50 values (based on nominal, measured or predicted REE concentrations in solution) drastically changed REE toxicity assessment and that these changes were influenced by REE speciation. This study demonstrates for the first time that REE speciation, and especially REE-DOM complexation, significantly influences REE bioaccumulation and toxicity towards D. magna. Our results have implications for the subsequent ERA of REE.

Profiling metal contamination from ultramafic sediments to biota along the Albanian shoreline of Lake Ohrid (Albania/Macedonia).

Ultramafic sediments exhibit high levels of geogenically-derived and potentially toxic metals, with Ni, Cr and Co often exceeding benchmark values. As yet, a holistic understanding of the bioavailability, mobility, potential ecotoxicity and trophic transfer of trace elements in both benthic and pelagic food chains in aquatic ultramafic environments (UME) is lacking. We investigated potential environmental health issues due to metal contamination by jointly implementing chemical, ecological and toxicological tools, along the Albanian shoreline of Lake Ohrid. It is an aquatic system of worldwide importance, representative of temperate UME with a legacy of Ni and Cr contamination from mining activities. Levels of Ni, Cr, Cd, Cu, Co, Fe, Mn and Zn were determined in waters, sediments and native biota. The potential environmental mobility of sediment-bound elements was further assessed using CaCl2, EDTA and acetic acid extractions. Whole-sediment ecotoxicity tests were also carried out using ostracods and chironomids, according to standardized procedures. Despite Ni and Cr concentrations above the sediment quality guidelines for probable effect levels, we did not observe ecotoxic effects in laboratory tests. However, these elements were bioavailable to native organisms under field conditions, especially to benthic primary producers in direct contact with sediments (up to 139 mg Cr kg-1 and 785 mg Ni kg-1). Although biomagnification was not observed, further investigations of metal translocation, metabolism and elemental trophic transfer along benthic food webs appears to be a general research priority in the management of temperate UME. The present study shows that proper management of temperate UME requires not only the integration of data from different lines of evidence, but also laboratory vs. field approaches to understand the subtler, long-term effects of increased elemental body burdens in native organisms.

Sub-chronic effects of AgNPs and AuNPs on Gammarus fossarum (Crustacea Amphipoda): From molecular to behavioural responses.

The aim of the present study was the assessment of the sub-chronic effects of silver (AgNPs) and gold nanoparticles (AuNPs) of 40 nm primary size either stabilised with citrate (CIT) or coated with polyethylene glycol (PEG) on the freshwater invertebrate Gammarus fossarum. Silver nitrate (AgNO3) was used as a positive control in order to study the contribution of silver ions potentially released from AgNPs on the observed effects. A multibiomarker approach was used to assess the long-term effects of AgNPs and AuNPs 40 nm on molecular, cellular, physiological and behavioural responses of G. fossarum. Specimen of G. fossarum were exposed for 15 days to 0.5 and 5 µgL-1 of CIT and PEG AgNPs and AuNPs 40 nm in the presence of food. A significant uptake of both Ag and Au was observed in exposed animals but was under the toxic threshold leading to mortality of G. fossarum. Silver nanoparticles (CIT-AgNPs and PEG-AgNPs 40 nm) led to an up-regulation of Na+K+ATPase gene expression. An up-regulation of Catalse and Chitinase gene expressions due to exposure to PEG-AgNPs 40 nm was also observed. Gold nanoparticles (CIT and PEG-AuNPs 40 nm) led to an increase of CuZnSOD gene expression. Furthermore, both AgNPs and AuNPs led to a more developed digestive lysosomal system indicating a general stress response in G. fossarum. Both AgNPs and AuNPs 40 nm significantly affected locomotor activity of G. fossarum while no effects were observed on haemolymphatic ions and ventilation.

Environmental transcriptomes of invasive dreissena, a model species in ecotoxicology and invasion biology.

Dreissenids are established model species for ecological and ecotoxicological studies, since they are sessile and filter feeder organisms and reflect in situ freshwater quality. Despite this strong interest for hydrosystem biomonitoring, omics data are still scarce. In the present study, we achieved full de novo assembly transcriptomes of digestive glands to gain insight into Dreissena polymorpha and D. rostriformis bugensis molecular knowledge. Transcriptomes were obtained by Illumina RNA sequencing of seventy-nine organisms issued from fifteen populations inhabiting sites that exhibits multiple freshwater contamination levels and different hydrosystem topographies (open or closed systems). Based on a recent de novo assembly algorithm, we carried out a complete, quality-checked and annotated transcriptomes. The power of the present study lies in the completeness of transcriptomes gathering multipopulational organisms sequencing and its full availability through an open access interface that gives a friendly and ready-to-use access to data. The use of such data for proteogenomic and targeted biological pathway investigations purpose is promising as they are first full transcriptomes for this two Dreissena species.

A sub-individual multilevel approach for an integrative assessment of CuO nanoparticle effects on Corbicula fluminea.

Because they are widely used, copper oxide nanoparticles (CuO NPs) are likely to enter the aquatic environment and then reach the sediment. We have examined the effect of CuO NPs in the freshwater endobenthic bivalve Corbicula fluminea. Some previous studies have investigated effects at biochemical and physiological levels, but molecular endpoints are still poorly studied despite they are sensitive in early detection of NPs effect. In the present study, we have investigated short-term effects (96 h) of CuO NP (12, 30 nm; 0, 20 and 100 µg/L) using molecular endpoints as well as more conventional biochemical and physiological markers. The expression of antioxidant (CuZnSOD, MnSOD, Cat, Se-GPx, Trxr) and antitoxic (GST-Pi, HSP70, MT, Pgp, MRP1) related genes was measured at the mRNA level while antioxidant (SOD, TAC) and antitoxic (GST, ACP) defenses, energetic reserves and metabolism (ETS, Tri, LDH), and cellular damages (LPO) were assessed using a biochemical approach. The filtration rate measured at 96 h provided information at the physiological scale. Gene expression and filtration rate were responsive to CuO NPs but the effects differed according to the NP size. The results suggest that defense mechanisms may have been set up following 30 nm-NP exposure. The response to 12 nm-NP was lower but still showed that exposure to 12 nm-NP led to activation of cellular elimination mechanisms. The lowering of the filtration rate may have protected the organisms from the contamination. However, this raised the question of further repercussions on organism biology. Together, the results (i) indicate that CuO NP may exert effects at different levels even after a short-term exposure and (ii) point out the precocity of molecular response.

Corbicula fluminea gene expression modulated by CeO2 nanomaterials and salinity.

Cerium dioxide nanomaterials (CeO2 NMs) are used in different fields and incorporated in daily products. Several studies highlighted their effects on organism physiology, although molecular studies remain scarce. NM behavior is strongly dependent on the environment but few data are available using complex exposure media, raising the question of its environmental impacts. The aim of the present work was to assess the toxic potential of three CeO2 NMs in Corbicula fluminea at a molecular level by RT-qPCR under a more realistic scenario of exposure, in a multistress context at two different salinities (1.5 and 15 psu). C. fluminea was exposed for 28 days to pulses of the three selected NMs (reference, manufactured, and aged manufactured). In bivalves, the gills and digestive gland are two key organs used for ecotoxicological studies. The expression change of 12 genes was measured in control organisms after 28 days in both organs, allowing us to clearly separate the responses for both organs and salinities. As gills come in contact with the environment first, we monitored gene the expression at intermediate time points (7, 14, and 21 days) for this organ in order to highlight clams responses to NM and salinity. Two genes (Se-GPx, MnSOD) had a salinity-dependent level of expression. HSP70, Se-GPx, and Trxr mRNAs presented significant changes in their expressions in the presence of NM. This study was completed using an integrated statistical approach. The exposed organisms differed more from control at field salinity than those exposed to hyper-saline conditions. At 15 psu, salinity pressure seems to cause the first molecular impact. At 1.5 psu, gene expression patterns allowed the effect of each NM to separate clearly. These results confirmed the usefulness of gene expression studies. Moreover, we highlighted the necessity to assess the environmental toxicity of the different forms of manufactured NM.

Changes in protein expression in mussels Mytilus galloprovincialis dietarily exposed to PVP/PEI coated silver nanoparticles at different seasons.

Potential toxic effects of Ag NPs ingested through the food web and depending on the season have not been addressed in marine bivalves. This work aimed to assess differences in protein expression in the digestive gland of female mussels after dietary exposure to Ag NPs in autumn and spring. Mussels were fed daily with microalgae previously exposed for 24 h to 10 µg/L of PVP/PEI coated 5 nm Ag NPs. After 21 days, mussels significantly accumulated Ag in both seasons and Ag NPs were found within digestive gland cells and gills. Two-dimensional electrophoresis distinguished 104 differentially expressed protein spots in autumn and 142 in spring. Among them, chitinase like protein-3, partial and glyceraldehyde-3-phosphate dehydrogenase, that are involved in amino sugar and nucleotide sugar metabolism, carbon metabolism, glycolysis/gluconeogenesis and the biosynthesis of amino acids KEGG pathways, were overexpressed in autumn but underexpressed in spring. In autumn, pyruvate metabolism, citrate cycle, cysteine and methionine metabolism and glyoxylate and dicarboxylate metabolism were altered, while in spring, proteins related to the formation of phagosomes and hydrogen peroxide metabolism were differentially expressed. Overall, protein expression signatures depended on season and Ag NPs exposure, suggesting that season significantly influences responses of mussels to NP exposure.

Energy allocation in two dreissenid species under metal stress.

Measurements of biological responses on living organisms are essential in aquatic biomonitoring. In freshwaters, Dreissena polymorpha is an invasive bivalve commonly used in ecotoxicological studies and considered as a model organism. However, D. polymorpha abundances are declining while another species colonizes most of the freshwaters: Dreissena rostriformis bugensis. This species has already been studied in ecophysiology but there is still a lack of data concerning its responses to stressors before its use as a bioindicator of environmental pollution. This study aims to compare the responses of the two species exposed to metal stress. Responses at different levels of biological organization were targeted with measurement of sub-cellular and individual biomarkers following an exposure of up to 7 days to cadmium at 10 µg.L-1. At the individual level, the scope for growth (SFG) was measured. It corresponds to the energy allocated to growth and reproduction. D. polymorpha exhibits variations in biomarker measurements as well as in the SFG in presence of Cd. D. r. bugensis shows no variation in its responses at the different targeted levels. According to the present results, energy metabolism seems to have an essential role for these species when facing a metal stress. Different energy allocation strategies were evidenced between the two species, although the link with biochemical biomarkers is more evident for D. polymorpha than for D. r. bugensis.

Identification of reference genes for RT-qPCR data normalization in Gammarus fossarum (Crustacea Amphipoda).

Gene expression profiling via RT-qPCR is a robust technique increasingly used in ecotoxicology. Determination and validation of optimal reference genes is a requirement for initiating RT-qPCR experiments. To our best knowledge, this study is the first attempt of identifying a set of reference genes for the freshwater crustacean Gammarus fossarum. Six candidate genes (Actin, TUB, UB, SDH, Clathrin and GAPDH) were tested in order to determine the most stable ones in different stress conditions and to increase the robustness of RT-qPCR data. SDH and Clathrin appeared as the most stable ones. A validation was performed using G. fossarum samples exposed for 15 days to AgNO3, silver nanoparticles (AgNPs) 40 nm and gold nanoparticles (AuNPs) 40 nm. Effects on HSP90 were evaluated and data normalized using Clathrin and SDH. A down-regulation of HSP90 was observed when G. fossarum were exposed to AuNPs 40 nm whereas no effects were observed when G. fossarum were exposed to AgNPs 40 nm. This study highlights the importance of the preliminary determination of suitable reference genes for RT-qPCR experiments. Additionally, this study allowed, for the first time, the determination of a set of valuable genes that can be used in other RT-qPCR studies using G. fossarum as model organism.

Do the pristine physico-chemical properties of silver and gold nanoparticles influence uptake and molecular effects on Gammarus fossarum (Crustacea Amphipoda)?

The specific and unique properties of silver nanoparticles (AgNPs) and gold nanoparticles (AuNPs), make them of high interest for different scientific and industrial applications. Their increasing use will inevitably lead to their release in the environment and aquatic ecosystems where they may represent a threat to aquatic organisms. Being a widespread and important component of the aquatic macroinvertebrate assemblage, amphipods and more specifically Gammarus fossarum will certainly be exposed to AgNPs and AuNPs. For these reasons, G. fossarum was selected as model organism for this study. The aim of the present work was the evaluation of the influence of both size (20, 40 and 80 nm) and surface coating (citrate CIT, polyethylene glycol PEG) on the acute toxicity of AgNPs and AuNPs on G. fossarum. We investigated the effects of AgNPs and AuNPs on the uptake by G. fossarum, NP tissue distribution and the expression of stress related genes by the use of ICP-MS, NanoSIMS50, Cytoviva®, and Rt-qPCR, respectively. Ag and Au bioaccumulation revealed a significant surface-coating dependence, with CIT-AgNPs and CIT-AuNPs showing the higher bio-accumulation potential in G. fossarum as compared to PEG-NPs. Opposite to that, no size-dependent effects on the bioaccumulation potential was observed. SIMS imaging and CytoViva® revealed an influence of the type of metal on the tissue distribution after uptake, with AgNPs detected in the cuticle and the gills of G. fossarum, while AuNPs were detected in the gut area. Furthermore, AgNPs were found to up-regulate CuZnSOD gene expression while AuNPs led to its down-regulation. Modulation of SOD may indicate generation of reactive species of oxygen and a possible activation of antioxidant defence in order to prevent and defend the organism from oxidative stress. However, further investigations are still needed to better define the mechanisms underlying the observed AgNPs and AuNPs effects.

Impact of multiple stressors on biomarker responses in sympatric dreissenid populations.

Dreissenid mussels, well-known invaders of the northern hemisphere, also constitute good biomonitors for freshwater quality assessment. Whereas the oldest, Dreissena polymorpha, is relatively well-known, the new invasive, Dreissena rostriformis bugensis, has been very little characterized. The aim of this study was to compare subcellular biomarker responses within these species, by taking into account inter- and intra-specific variability in biomarker responses during a multi-stress assessment. Indeed, due to local environmental conditions and genetic background, biomarker responses can be different between populations of a given species, not only between species. To evaluate these differential responses, we sampled mussels on two sites where they are both present in sympatry (plus another D. r. bugensis population), and we exposed these populations in laboratory to different stressors: thermal stress (12 or 17 °C), dietary stress (fed or unfed), contamination (nickel at 0, 20 and 500 µg L-1) and exposure duration (0, 4 or 8 days). Results mainly evidenced strong inter-species and inter-population differences, underlining the need to discriminate correctly between the two species and to know well the populations used in biomonitoring. Results also evidenced thermal and food stress-related effects. The numerous data obtained during this multi-stress experiment also highlight the complexity of working on several stressors and analysing the associated results.

Genotoxicity and physiological effects of CeO2 NPs on a freshwater bivalve (Corbicula fluminea).

The rapid development of nanotechnology and the increased use of nanomaterials in products used in everyday life have raised the question of the potential release of nanoparticles into the aquatic environment. Their fate and effects in natural ecosystems are not currently well understood but harmful effects of nanoparticles have been demonstrated at low concentrations on some freshwater and marine species. Cerium dioxide nanoparticles (CeO2 NPs) are produced in large quantities and used in products in many different fields, such as automotives or optics. Because of their widespread use in daily products, CeO2 NPs are included in the OECD priority list of manufactured nanomaterials for human and environmental assessment. Indeed some studies have been conducted to assay various enzymatic biomarkers, which showed the CeO2 NPs potential to modify anti-oxidative defenses and cellular membrane stability. Nevertheless, only a few studies were performed on their genotoxic potential. The aim of this work was to evaluate the genotoxic and physiological effects of CeO2 NPs on a widespread freshwater bivalve Corbicula fluminea by using comet assay and a multi-enzymatic biomarker approach. Exposure to two CeO2 NP concentrations during a short term experiment (6 days) was set up. The first one (10 µg/L) was chosen in order to work with low but measurable concentrations whereas the second one was ten times higher (100 µg CeO2 NPs/L). DNA damage was significantly more pronounced compared with control for both concentrations tested as early as two days of exposure and seemed to increase with time. Some enzymatic biomarkers of anti-oxidative defenses (total antioxidant capacity, catalase activity), anti-toxic mechanisms (glutathione-S-transferase activity, caspase-3 activity) or metabolism (lactate dehydrogenase activity) tended to increase after 6 days of exposure but only the induction of caspase pathway and DNA damages appeared significant for exposed organisms. In this study, time and concentration effects of CeO2 NPs were highlighted by coupling genotoxic and cellular biomarker assessments.

Differential tolerance to nickel between Dreissena polymorpha and Dreissena rostriformis bugensis populations.

Differential tolerance to stress is partly responsible for the heterogeneity of biomarker responses between populations of a sentinel species. Although currently used for freshwater biomonitoring, studies concerning inter-populational variability in tolerance to contaminants for the zebra mussel (Dreissena polymorpha) are scarce. Moreover, this well-known invader is currently replaced by another, the quagga mussel (Dreissena rostriformis bugensis). To evaluate the differential tolerance between dreissenids, several populations of both species were exposed to a high concentration of nickel. A LT50 (time when 50% of individuals were dead) was established for each population. Biomarker responses and internal nickel concentration were also measured, to link tolerance with physiological status. Results evidenced that D. polymorpha populations are more heterogeneous and more tolerant than D. r. bugensis ones. For D. polymorpha populations only, LT50 values were positively correlated with the nickel contamination in situ, with higher anti-oxidative defences and a higher Integrated Biomarker Response value in the field. Such findings may be explained by local adaptation and invasion dynamic within each species. The significance of this differential tolerance when using biomarker responses for biomonitoring purposes is thus discussed.

Combined influence of oxygenation and salinity on aggregation kinetics of the silver reference nanomaterial NM-300K.

The combined influence of oxygenation and salinity on agglomeration and/or aggregation kinetics of the silver (Ag) nanomaterial NM-300K was investigated, and the relationship between its physicochemical fate and toxicity toward an estuarine bivalve was established. The results showed that the presence of NaCl under certain oxygen conditions (8.5 ppm) promoted the formation of AgCl aggregates that could be linked to toxicity effects on aquatic organisms. Environ Toxicol Chem 2018;37:1007-1013. © 2017 SETAC.

Integrated multi-biomarker responses in two dreissenid species following metal and thermal cross-stress.

With current global changes, the combination of several stressors such as temperature and contaminants may impact species distribution and ecosystem functioning. In this study, we evaluated the combined impact of two metals (Ni and Cr) with a thermal stress (from 12 to 17 °C) on biomarker responses in two bivalves, Dreissena rostriformis bugensis and Dreissena polymorpha. Biomarkers are informative tools to evaluate exposure and effects of stressors on organisms. The set of 14 biomarkers measured here was representative of both physiologic (filtration activity) and cellular antioxidant and detoxification mechanisms. Our aim was to study the response pattern of both species, and its meaning in terms of invasive potential. The implications for the use of these mussels in environmental monitoring are also discussed. Results evidenced that the two species do not respond to multiple stressors in the same way. Indeed, the effects of contamination on biomarker responses were more marked for D. polymorpha, especially under nickel exposure. While we cannot conclude as to the effect of temperature, invasiveness could be influenced by species sensitivity to contaminants. The physiological and cellular differences between D. polymorpha and D. r. bugensis might also be of concern for environmental risk assessment. The two species present differential bioaccumulation patterns, filtration activity and cellular biomarker responses. If D. polymorpha populations decline, their substitution by D. r. bugensis for biomonitoring or laboratory studies will not be possible without a deeper understanding of biomarker responses of the new invasive.

Gammarus fossarum (Crustacea, Amphipoda) as a model organism to study the effects of silver nanoparticles.

Amphipods are one of the most important components of freshwater ecosystems. Among them, gammarids are the most widespread group in Europe and are often used as bioindicators and model organisms in ecotoxicology. However, their use, especially of Gammarus fossarum for the study of the environmental impact of nanoparticles, has been rather limited so far. G. fossarum was selected to assess effects of well-characterized chemically synthesized silver nanoparticles (AgNPs 20nm and 200nm) and "green" laboratory synthetized (from plant leaf extracts) AgNPs (AgNPs 23nm and 27nm). AgNO3 was used as a positive control to compare AgNPs effects and silver ions effects. A multibiomarker approach was used to investigate the sub-lethal effects of AgNPs on physiological and behavioural responses of G. fossarum. Two different experiments were carried out. In a preliminary experiment, two populations of G. fossarum (G.f1 and G.f2) were tested for sensitivity differences and the most sensitive one was exposed, in a final experiment, to sub-lethal concentrations of AgNO3 and the most toxic AgNPs. AgNO3 and AgNPs 23nm led to a significant decrease in survival rates, osmoregulation and locomotor activity. Ag internalisation, performed with Secondary Ion Mass Spectrometry (SIMS), showed the presence of silver in gills of G.f2 exposed to AgNPs 23 and 27nm. This study highlighted the influence of method of synthesis on ion release, uptake and toxic effects of AgNPs on G. fossarum. Osmoregulation appeared to be an effective biomarker indicating the physiological health status of G. fossarum. Locomotor activity, which was the most impacted response, reflects the potential effects of released ions from AgNPs 23nm at the population level as locomotion is necessary for foraging, finding mates and escaping from predators. Therefore, we propose G. fossarum as a suitable model for environmental nanotoxicology, providing information both at individual and population levels.

The influence of salinity on the fate and behavior of silver standardized nanomaterial and toxicity effects in the estuarine bivalve Scrobicularia plana.

Because of their antibacterial properties, silver (Ag) engineered nanomaterials are included in many products. The present study used a standardized Ag nanomaterial (NM-300K, 20 nm) supplied with a stabilizing agent. The aim was to investigate the behavior of Ag nanomaterial in an estuarine-like medium at 2 salinities (15 psu and 30 psu). Uptake as well as sublethal effects of Ag nanomaterial (10 µg Ag/L), its stabilizing agent, and AgNO3 (10 µg Ag/L) were assessed in the clam Scrobicularia plana, after 7 d of exposure. The release of soluble Ag from Ag nanomaterial in the experimental media was quantified by using diffusive gradient in thin films and ultrafiltration. A multibiomarker approach was employed to reveal responses of clams at subindividual and individual levels. The bioaccumulation of Ag was significantly greater at 15 psu versus 30 psu, which could be explained by differences in Ag speciation. In conclusion, the present study showed different impacts of Ag nanomaterial that were not always explained by the release of Ag ions in clams at both salinities; such impacts were particularly characterized by induction of oxidative stress, cell damage, and impairment of energetic levels. Burrowing of clams was affected by the stabilizing agent depending on the salinity tested, with stronger effects at 15 psu. Finally, the present study highlighted salinity-dependent changes in the physiology of estuarine bivalves. Environ Toxicol Chem 2016;35:2550-2561. © 2016 SETAC.