Contaminants of Concern and Spatiotemporal Metabolomic Changes in Quagga Mussels (Dreissena bugensis rostriformis) from the Milwaukee Estuary (Wisconsin, USA).

Environmental metabolomics has emerged as a promising technique in the field of biomonitoring and as an indicator of aquatic ecosystem health. In the Milwaukee Estuary (Wisconsin, USA), previous studies have used a nontargeted metabolomic approach to distinguish between zebra mussels (Dreissena polymorpha) collected from sites of varying contamination. To further elucidate the potential effects of contaminants on bivalve health in the Milwaukee Estuary, the present study adopted a caging approach to study the metabolome of quagga mussels (Dreissena bugensis rostriformis) deployed in six sites of varying contamination for 2, 5, or 55 days. Caged mussels were co-deployed with two types of passive sampler (polar organic chemical integrative samplers and semipermeable membrane devices) and data loggers. In conjunction, in situ quagga mussels were collected from the four sites studied previously and analyzed for residues of contaminants and metabolomics using a targeted approach. For the caging study, temporal differences in the metabolomic response were observed with few significant changes observed after 2 and 5 days, but larger differences (up to 97 significantly different metabolites) to the metabolome in all sites after 55 days. A suite of metabolic pathways were altered, including biosynthesis and metabolism of amino acids, and upmodulation of phospholipids at all sites, suggesting a potential biological influence such as gametogenesis. In the caging study, average temperatures appeared to have a greater effect on the metabolome than contaminants, despite a large concentration gradient in polycyclic aromatic hydrocarbons residues measured in passive samplers and mussel tissue. Conversely, significant differences between the metabolome of mussels collected in situ from all three contaminated sites and the offshore reference site were observed. Overall, these findings highlight the importance of contextualizing the effects of environmental conditions and reproductive processes on the metabolome of model organisms to facilitate the wider use of this technique for biomonitoring and environmental health assessments. Environ Toxicol Chem 2024;43:307-323. © 2023 The Authors. Environmental Toxicology and Chemistry published by Wiley Periodicals LLC on behalf of SETAC. This article has been contributed to by U.S. Government employees and their work is in the public domain in the USA.

Integrative biomarker response - Threshold (IBR-T): Refinement of IBRv2 to consider the reference and threshold values of biomarkers.

The Integrated Biomarker Response (IBR) is one of the most used index in biomonitoring, especially the IBRv2 integrating a reference condition. However, some limitations remain for its routine and large-scale use. The IBRv2 is proportional to the total number of biomarkers, is dependent on the nature of biomarkers and considers all biomarkers modulations, even small and biologically non-significant. In addition, IBRv2 relies on reference values but the references are often different between each study, making it difficult to compare results between studies and/or campaigns. To overcome these limitations, the present work proposed a new index called IBR-T ("Integrated Biomarker Response - Threshold") which considers the threshold values of biomarkers by limiting the calculation of the IBR value to biomarkers with significant modulations. The IBRv2 and the IBR-T were calculated and compared on four datasets from active biomonitoring campaigns using Dreissena polymorpha, a bivalve widely used in freshwater biomonitoring studies. The comparison between indices has demonstrated that the IBR-T presents a better correlation (0.907 < r2 < 0.998) with the percentage of biomarkers significantly modulated than the IBRv2 (0.002 < r2 < 0.759). The IBRv2 could not be equal to 0 (0.915 < intercept <1.694) because the value was dependent on the total number of biomarkers, whereas the IBR-T reached 0 when no biomarker was significantly modulated, which appears more biologically relevant. The final ranking of sites was different between the two index and the IBR-T ranking tends to be more ecologically relevant that the IBRv2 ranking. This IBR-T have shown an undeniable interest for biomonitoring and could be used by environmental managers to simplify the interpretation of large datasets, directly interpret the contamination status of the site, use it to decision-making, and finally to easily communicate the results of biomonitoring studies to the general public.

Review and Development of Best Practices for Toxicity Tests with Dreissenid Mussels.

Since their introduction to North America in the 1980s, research to develop effective control tools for invasive mussels (Dreissena polymorpha and D. rostriformis bugensis) has been ongoing across various research institutions using a range of testing methods. Inconsistencies in experimental methods and reporting present challenges for comparing data, repeating experiments, and applying results. The Invasive Mussel Collaborative established the Toxicity Testing Work Group (TTWG) in 2019 to identify "best practices" and guide development of a standard framework for dreissenid mussel toxicity testing protocols. We reviewed the literature related to laboratory-based dreissenid mussel toxicity tests and determined the degree to which standard guidelines have been used and their applicability to dreissenid mussel testing. We extracted detailed methodology from 99 studies from the peer-reviewed and gray literature and conducted a separate analysis for studies using presettlement and postsettlement mussels. We identified specific components of methods and approaches that could be refined or standardized for dreissenid mussels. These components included species identification, collection methods, size/age class distinction, maintenance practices, testing criteria, sample size, response measures, reporting parameters, exposure methods, and mortality criteria. We consulted experts in the field of aquatic toxicology and dreissenid mussel biology on our proposed. The final recommendations contained in the present review are based on published standard guidelines, methods reported in the published and gray literature, and the expertise of TTWG members and an external panel. In addition, our review identifies research needs for dreissenid mussel testing including improved methods for early-life stage testing, comparative data on life stages and between dreissenid mussel species, inclusion of a reference toxicant, and additional testing of nontarget species (i.e., other aquatic organisms). Environ Toxicol Chem 2023;42:1649-1666. © 2023 His Majesty the King in Right of Canada. Environmental Toxicology and Chemistry published by Wiley Periodicals LLC on behalf of SETAC. Reproduced with the permission of the Minister of Environment and Climate Change Canada. This article has been contributed to by U.S. Government employees and their work is in the public domain in the USA.

Recreationist willingness to pay for aquatic invasive species management.

We estimated willingness to pay for local aquatic invasive species lake management in the form of a daily lake access fee by conducting summer lake surveys in Minnesota, USA. Similar pairs of lakes with differing infestations of zebra mussels, Dreissena polymorpha, and starry stonewort, Nitellopsis obtuse, were used as study sites to infer how being at an infested lake vs. being at an uninfested lake and different local species would impact responses. We also examined recreationists' visit motivation, and aquatic invasive species perceived risk, knowledge, and awareness of problem. We estimated mean willingness to pay about nine to ten dollars per day, which did not differ significantly by lake. Additionally, perceived risk, awareness of problem, and visit motivation were significant in predicting willingness to pay, which could have important ramifications for aquatic invasive species management.

Survival and infectivity of Toxoplasma gondii and Cryptosporidium parvum oocysts bioaccumulated by Dreissena polymorpha.

AIMS: The study was aimed to understand the depuration process of Cryptosporidium parvum and Toxoplasma gondii oocysts by zebra mussel (Dreissena polymorpha), to consider the use of the zebra mussel as a bioremediation tool. MATERIALS AND METHODS: Two experiments were performed: (i) individual exposure of mussel to investigate oocyst transfers between bivalves and water and (ii) in vivo exposure to assess the ability of the zebra mussel to degrade oocysts. RESULTS: (i) Our results highlighted a transfer of oocysts from the mussels to the water after 3 and 7 days of depuration; however, some oocysts were still bioaccumulated in mussel tissue. (ii) Between 7 days of exposure at 1000 or 10 000 oocysts/mussel/day and 7 days of depuration, the number of bioaccumulated oocysts did not vary but the number of infectious oocysts decreased. CONCLUSION: Results show that D. polymorpha can release oocysts in water via (pseudo)faeces in depuration period. Oocysts remain bioaccumulated and infectious oocyst number decreases during the depuration period in zebra mussel tissues. Results suggest a degradation of bioaccumulated C. parvum and T. gondii oocysts. SIGNIFICANCE AND IMPACT OF THE STUDY: This study highlighted the potential use of D. polymorpha as a bioremediation tool to mitigate of protozoan contamination in water resources.

Temperature-Related Responses of an Invasive Mussel and 2 Unionid Mussels to Elevated Carbon Dioxide.

Zebra mussels (Dreissena polymorpha) have exacerbated the decline of native freshwater mussels (order Unionida) in North America since their arrival in the 1980s. Options for controlling invasive mussels, particularly in unionid mussel habitats, are limited. Previously, carbon dioxide (CO2 ) showed selective toxicity for zebra mussels, relative to unionids, when applied in cool water (12 °C). We first determined 96-h lethal concentrations of CO2 at 5 and 20 °C to zebra mussels and responses of juvenile plain pocketbook (Lampsilis cardium). Next, we compared the time to lethality for zebra mussels at 5, 12, and 20 °C during exposure to partial pressure of CO2 (PCO2 ) values of 110 to 120 atm (1 atm = 101.325 kPa) and responses of juvenile plain pocketbook and fragile papershell (Leptodea fragilis). We found efficacious CO2 treatment regimens at each temperature that were minimally lethal to unionids. At 5 °C, plain pocketbook survived 96-h exposure to the highest PCO2 treatment (139 atm). At 20 °C, the 96-h lethal concentration to 10% of animals (LC10) for plain pocketbook (173 atm PCO2 , 95% CI 147-198 atm) was higher than the LC99 for zebra mussels (118 atm PCO2 , 95% CI 109-127 atm). Lethal time to 99% mortality (LT99) of zebra mussels in 110 to 120 atm PCO2 ranged from 100 h at 20 °C to 300 h at 5 °C. Mean survival of both plain pocketbook and fragile papershell juveniles exceeded 85% in LT99 CO2 treatments at all temperatures. Short-term infusion of 100 to 200 atm PCO2 at a range of water temperatures could reduce biofouling by zebra mussels with limited adverse effects on unionid mussels. Environ Toxicol Chem 2020;39:1546-1557. Published 2020. This article is a U.S. Government work and is in the public domain in the USA. Environmental Toxicology and Chemistry published by Wiley Periodicals LLC on behalf of SETAC.

Identification of Optimal Calcium and Temperature Conditions for Quagga Mussel Filtration Rates as a Potential Predictor of Invasion.

Quagga mussels (Dreissena rostriformis burgensis) are a highly invasive aquatic species to North America, capable of filtering large volumes of water and causing severe ecological and economic impacts. Their range has been expanding since they first invaded the Great Lakes in the 1980s. To predict their spread, it is crucial to understand environmental parameters, which facilitate their range expansion. Two factors likely to influence their distribution include calcium and temperature, because the former is vital for shell development and the latter for metabolic activity. When these factors are optimal for mussels' fitness, the filtration rate has the potential to be maximized if other environmental conditions are also favorable, thus enabling mussels to exploit their growth potential. Deviations from optimal conditions likely result in filtration-rate decline. We identify calcium concentrations and temperatures that maximize the mussel filtration rate for 2 phytoplankton species: Ankistrodesmus facaltus, a common food source for quagga mussels, and a less palatable Microcystis icthyoblabe. In laboratory experiments, filtration rates were measured through cell counts after 24 h of filtration when exposed to a range of temperatures between 2 and 30 °C, and calcium concentrations between 0 and 180 mg/L. Response surface methodology was used to identify a maximum filtration rate, which occurred at 22 mL/mg/h at 137 mg/L of calcium carbonate and 26 °C when fed Ankistrodesmus. To establish a quagga mussel population in a new water source, optimum conditions are required; thus, this information can be used to rank the relative susceptibility of water bodies to invasion by quagga mussels. Environ Toxicol Chem 2020;39:410-418. © 2019 SETAC.

Dreissenid (quagga and zebra mussel) veligers are adversely affected by bloom forming cyanobacteria.

Quagga (Dreissena rostriformis bugensis) and zebra (D. polymorpha) mussels are broadcast spawners that produce planktonic, free swimming veligers, a life history strategy dissimilar to native North American freshwater bivalves. Dreissenid veligers require highly nutritious food to grow and survive, and thus may be susceptible to increased mortality rates during harsh environmental conditions like cyanobacteria blooms. However, the impact of cyanobacteria and one of the toxins they can produce (microcystin) has not been evaluated in dreissenid veligers. Therefore, we exposed dreissenid veligers to eleven distinct cultures (isolates) of cyanobacteria representing Anabaena, Aphanizomenon, Dolichospermum, Microcystis, and Planktothrix species and the cyanotoxin microcystin to determine the lethality of cyanobacteria on dreissenid veligers. Six-day laboratory bioassays were performed in microplates using dreissenid veligers collected from the Detroit River, Michigan, USA. Veligers were exposed to increasing concentrations of cyanobacteria and microcystin using the green algae Chlorella minutissima as a control. Based on dose response curves formulated from a Probit model, the LC50 values for cyanobacteria used in this study range between 15.06 and 135.06 µg/L chlorophyll-a, with the LC50 for microcystin-LR at 13.03 µg/L. Because LC50 values were within ranges observed in natural waterbodies, it is possible that dreissenid recruitment may be suppressed when veliger abundances overlap with seasonal cyanobacteria blooms. Thus, the toxicity of cyanobacteria to dreissenid veligers may be useful to include in models forecasting dreissenid mussel abundance and spread.

Cyanobacteria reduce motility of quagga mussel (Dreissena rostriformis bugensis) sperm.

The temporal expansion of harmful algal blooms, primarily associated with cyanobacteria, may impact aquatic organisms at vulnerable life-history stages. Broadcast spawning species release gametes into the water column for external fertilization, directly exposing sperm to potential aquatic stressors. To determine if cyanobacteria can disrupt reproduction in freshwater broadcast spawners, we evaluated sublethal effects of cyanobacteria exposure on quagga mussel (Dreissena rostriformis bugensis) sperm. In laboratory studies, sperm were collected after inducing mussels to spawn using serotonin and exposed to 11 cultures of cyanobacteria including Anabaena flos-aquae, Aphanizomenon flos-aquae, Dolichospermum lemmermannii, Gloeotrichia echinulata, 5 cultures of Microcystis aeruginosa, M. wesenbergii, and Planktothrix suspensa. Sperm motility, using endpoints of cumulative distance traveled and mean velocity, was calculated for a minimum of 10 individual sperm using a novel optical biotracking assay method. The distance and velocity at which sperm traveled decreased when exposed to Aphanizomenon flos-aquae and 2 M. aeruginosa cultures. Our findings indicate that cyanobacteria impede the motility of quagga mussel sperm, which can potentially result in reproductive impairments to mussels and potentially other broadcast spawning species. Environ Toxicol Chem 2019;38:368-374. © 2018 SETAC.

Toxic effect and physiological disruption of sodium phosphate to the quagga mussel (Dreissena bugensis).

Phosphorous is an essential nutrient for all forms of life; however, the question of toxicity to aquatic species remains largely unanswered, despite many systems that exceed natural phosphorus loads. This study determined the ecotoxicological threshold concentration of phosphorus to the freshwater bivalve Dreissena bugensis using a 96-h bioassay. Sublethal, medial lethal, and lethal levels of sodium phosphate to D. bugensis were found to be 125, 260, and 476 ppm. Physiological biomarkers such as the oxygen consumption and filtration rate were estimated by exposing D. bugensis to five different sublethal concentrations (25, 50, 75, 100, and 125 ppm) of sodium phosphate for 96 h. Both oxygen consumption and filtration rate gradually declined with increasing exposure concentrations and durations, which was significant (α < 0.05) for 75, 100, and 125 ppm of sodium phosphate concentrations. Based on the feeding rate and oxygen consumption endpoints, the no-observed effect concentration and the low observed effect concentration were 25 and 75 ppm, respectively. Maximum acceptable toxicant concentration of sodium phosphate was 43.3 ppm. Measured environmental concentration (MEC) of total phosphorus (0.015 ppm; n = 6) was obtained from seasonal field assessments in Saginaw Bay during the years 2008 to 2010. An assessment factor of 1000 was used for calculating the predicted no effect concentration (PNEC) of 0.025 ppm. Risk quotient (RQ) of "0.6" was therefore established using MEC/PNEC (real risk) ratio. Binary ecological classification (RQ < 1) suggested that there is no appreciable risk of phosphorus to D. bungensis in the Saginaw Bay of Lake Huron of Laurentian Great Lakes.

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.

Ecological change alters the evolutionary response to harvest in a freshwater fish.

Harvesting can induce rapid evolution in animal populations, yet the role of ecological change in buffering or enhancing that response is poorly understood. Here, we developed an eco-genetic model to examine how ecological changes brought about by two notorious invasive species, zebra and quagga mussels, influence harvest-induced evolution and resilience in a freshwater fish. Our study focused on lake whitefish (Coregonus clupeaformis) in the Laurentian Great Lakes, where the species supports valuable commercial and subsistence fisheries, and where the invasion of dreissenid (zebra and quagga) mussels caused drastic shifts in ecosystem productivity. Using our model system, we predicted faster rates of evolution of maturation reaction norms in lake whitefish under pre-invasion ecosystem conditions when growth and recruitment of young to the population were high. Slower growth rates that occurred under post-invasion conditions delayed when fish became vulnerable to the fishery, thus decreasing selection pressure and lessening the evolutionary response to harvest. Fishing with gill nets and traps nets generally selected for early maturation at small sizes, except when fishing at low levels with small mesh gill nets under pre-invasion conditions; in this latter case, evolution of delayed maturation was predicted. Overall, the invasion of dreissenid mussels lessened the evolutionary response to harvest, while also reducing the productivity and commercial yield potential of the stock. These results demonstrate how ecological conditions shape evolutionary outcomes and how invasive species can have a direct effect on evolutionary responses to harvest and sustainability.

Species identity matters when interpreting trophic markers in aquatic food webs.

In aquatic systems, food web linkages are often assessed using diet contents, stable isotope ratios, and, increasingly, fatty acid composition of organisms. Some correlations between different trophic metrics are assumed to be well-supported; for example, particular stable isotope ratios and fatty acids seem to reflect reliance on benthic or pelagic energy pathways. However, understanding whether the assumed correlations between different trophic metrics are coherent and consistent across species represents a key step toward their effective use in food web studies. To assess links among trophic markers, we compared relationships between major diet components, fatty acids, and stable isotope ratios in three fishes: yellow perch (Perca flavescens), round goby (Neogobius melanostomus), and spottail shiner (Notropis hudsonius) collected from nearshore Lake Michigan. Yellow perch and spottail shiner are native in this system, while round goby are a relatively recent invader. We found some evidence for agreement between different trophic metrics, especially between diet components, n-3:n-6 fatty acid ratios, and stable isotope ratios (δ13C and δ15N). However, we also observed significant variation in observed relationships among markers and species, potentially due to taxonomic variation in the specific diet items consumed (e.g., chydorid microcrustaceans and Dreissena mussels) and species-specific biochemical processes. In many of these latter cases, the invasive species differed from the native species. Understanding the effects of taxonomic variation on prey and predator signatures could significantly improve the usefulness of fatty acids in food web studies, whereas diet contents and stable isotopes appear to be reliable indicators of trophic niche in aquatic food webs.

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.

Neuronal Development in the Larvae of the Invasive Biofouler Dreissena polymorpha (Mollusca: Bivalvia), with Special Attention to Sensory Elements and Swimming Behavior.

Although understanding of the neuronal development of Trochozoa has progressed recently, little attention has been paid to freshwater bivalves, including species with a strong ecological impact, such as the zebra mussel (Dreissena polymorpha). Therefore, an important question might concern how the developing nervous system is involved in the formation of the rapid and successful invasive behavior of this species. Our aim was to reveal the neuronal development of trochophore and veliger larvae of Dreissena, with special attention to the organization of sensory structures and their possible involvement in detecting environmental cues. After applying serotonin and FMRFamide immunocytochemistry, the first serotonin immunoreactive sensory elements appeared 16-18 hours after fertilization, whereas the first FMRFamide immunoreactive sensory cell was seen only at 32 hours of development (trochophore stage). Later, sensory elements were found in three parts of the larval body, including the apical organ, the posterior region, and the stomach. Although differences in the timing of appearance and the morphology of cells were observed, the two signaling systems showed basic similarity in their organization pattern until the end of the veliger stage. Pharmacological, physiological, and quantitative immunocytochemical investigations were also performed, suggesting the involvement of both the serotoninergic system and the FMRFamidergic system in sensomotor processes. Manipulation of the serotonin synthesis by para-chloroplenylalanine and 5-hydroxytryptophane, as well as application of increased salinity, influenced larval swimming activity, both accompanied by changes in immunofluorescence intensity. We concluded that these two early sensory systems may play an important role in the development of settlement competency of this biofouling invasive bivalve, Dreissena.

Lessons learned from studies with the freshwater mussel Dreissena polymorpha exposed to platinum, palladium and rhodium.

The platinum group elements (PGE) platinum, palladium and rhodium gain increasing (eco-)toxicological interest due to their cumulative introduction into ecosystems. So far, most PGE exposure studies investigating biological availability, uptake and bioaccumulation of PGE as well as their effects on different toxicological endpoints were performed under non-standardized conditions which occasionally make an interpretation and comparison of the results difficult. Here we compare the results of different PGE exposure studies with zebra mussels (Dreissena polymorpha) showing influences due to the PGE source, the exposure medium, the exposure concentration and period as well as the test system. Problems associated to the performance and evaluation of these studies were identified and recommendations as well as needs for future studies are given. As nominal exposure concentrations often did not reflect real exposure conditions the reference for exposure concentration has to be chosen with caution, i.e. nominal versus quantified aqueous concentrations. The determination of bioaccumulation factors can be problematic when PGE concentrations in the exposure medium and in the test organism did not reach steady state even after several weeks of exposure. For future studies it would be advantageous to regularly correlating PGE bioaccumulation and biomarker responses to increase the knowledge on potential adverse effects of these metals, preferably using environmentally relevant PGE concentration scenarios. Many aspects discussed in the present study for zebra mussels and PGE can be transferred to other aquatic animals and other metals, respectively.

Field biomonitoring using the zebra mussel Dreissena polymorpha and the quagga mussel Dreissena bugensis following immunotoxic reponses. Is there a need to separate the two species?

The zebra mussel, Dreissena polymorpha constitutes an extensively used sentinel species for biomonitoring in European and North American freshwater systems. However, this invasive species is gradually replaced in freshwater ecosystem by Dreissena bugensis, a closely related dreissenid species that shares common morphological characteristics but possess some physiological differences. However, few are known about differences on more integrated physiological processes that are generally used as biomarkers in biological monitoring studies. Declining of zebra mussel populations raises the question of the sustainability of using one or both species indifferently to maintain the quality of environmental pollution monitoring data. In our study, we performed a field comparative study measuring immune-related markers and bioaccumulation of PCBs, PAHs and PBDEs in sympatrically occurring mussel populations from three sites of the St. Lawrence River. For tested organisms, species were identified using RFLP analysis. Measurement of bioaccumulated organic compounds indicated a higher accumulation of PCBs and PBDEs in D. bugensis soft tissues compared to D. polymorpha while no differences were noticed for PAHs. Results of hemocytic parameters highlighted that differences of hemocyte distributions were associated to modulations of phagocytic activities. Moreover, marked differences occurred in measurement of hemocytic oxidative activity, indicating divergences between the two species for ROS regulation strategies. This physiological characteristic may deeply influence species responses facing environmental or pollution related stress and induce bias if the two species are not differentiated in further biomarker or bioaccumulation measurement-based studies.

Evaluation of uptake and chronic toxicity of virgin polystyrene microbeads in freshwater zebra mussel Dreissena polymorpha (Mollusca: Bivalvia).

Microplastics (MPs), plastic debris smaller than 5mm, are widely found in both marine and freshwater ecosystems. However, few studies regarding their hazardous effects on inland water organisms, have been conducted. For this reason, the aim of our research was the evaluation of uptake and chronic toxicity of two mixtures (MIXs) of virgin polystyrene microbeads (PMs) of 10µm and 1µm in size (MIX 1, with 5×105 of 1µmsizePMs/L and 5×105 of 10µmsizePMs/L, and MIX 2 with 2×106 of 1µmsizePMs/L and 2×106 of 10µmsizePMs/L) on freshwater zebra mussel Dreissena polymorpha (Mollusca: Bivalvia) during 6 exposure days. The PM uptake in the mussel body and hemolymph was assessed using confocal microscopy, while the chronic toxicity of PMs was evaluated on exposed mussels using a comprehensive battery of biomarkers of cellular stress, oxidative damage and neuro- genotoxicity. Confocal microscopy analyses showed that MPs concentrated in the gut lumen of exposed mussels, absorbed and transferred firstly in the tissues and then in the hemolymph. The results revealed that PMs do not produce oxidative stress and genetic damage, with the exception of a significant modulation of catalase and glutathione peroxidase activities in mussels exposed to MIX 1. Regarding neurotoxicity, we observed only a significant increase of dopamine concentration in mussels exposed to both MIXs, suggesting a possible implication of this neurotransmitter in an elimination process of accumulated PMs. This research represents a first study about the evaluation of virgin MP toxicity in zebra mussel and more research is warranted concerning the long term neurological effects of virgin MPs.

Genotoxic and Cytotoxic Effects on the Immune Cells of the Freshwater Bivalve Dreissena polymorpha Exposed to the Environmental Neurotoxin BMAA.

The environmental neurotoxin ß-N-Methylamino-l-alanine (BMAA) has been pointed out to be involved in human neurodegenerative diseases. This molecule is known to be bioaccumulated by bivalves. However, little data about its toxic effects on freshwater mussels is available, particularly on the hemolymphatic compartment and its hemocyte cells involved in various physiological processes such as immune defenses, digestion and excretion, tissue repair, and shell production. Here we exposed Dreissena polymorpha to dissolved BMAA, at the environmental concentration of 7.5 µg of /mussel/3 days, during 21 days followed by 14 days of depuration in clear water, with the objective of assessing the BMAA presence in the hemolymphatic compartment, as well as the impact of the hemocyte cells in terms of potential cytotoxicity, immunotoxicity, and genotoxiciy. Data showed that hemocytes were in contact with BMAA. The presence of BMAA in hemolymph did not induce significant effect on hemocytes phagocytosis activity. However, significant DNA damage on hemocytes occurred during the first week (days 3 and 8) of BMAA exposure, followed by an increase of hemocyte mortality after 2 weeks of exposure. Those effects might be an indirect consequence of the BMAA-induced oxidative stress in cells. However, DNA strand breaks and mortality did not persist during the entire exposure, despite the BMAA persistence in the hemolymph, suggesting potential induction of some DNA-repair mechanisms.

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.