Proteome changes in muscles, ganglia, and gills in Corbicula fluminea clams exposed to crude oil: Relationship with behavioural disturbances.

The use of online remote control for 24/7 behavioural monitoring can play a key role in estimating the environmental status of aquatic ecosystems. Recording the valve activity of bivalve molluscs is a relevant approach in this context. However, a clear understanding of the underlying disturbances associated with behaviour is a key step. In this work, we studied freshwater Asian clams after exposure to crude oil (measured concentration, 167 ± 28 µg·L-1) for three days in a semi-natural environment using outdoor artificial streams. Three complementary approaches to assess and explore disturbances were used: behaviour by high frequency non-invasive (HFNI) valvometry, tissue contamination with polycyclic aromatic hydrocarbons (PAH), and proteomic analysis. Two tissues were targeted: the pool adductor muscles - retractor pedal muscle - cerebral and visceral ganglia, which is the effector of any valve movement and the gills, which are on the frontline during contamination. The behavioural response was marked by an increase in valve closure-duration, a decrease in valve opening-amplitude and an increase in valve agitation index during opening periods. There was no significant PAH accumulation in the muscle plus nervous ganglia pool, contrary to the situation in the gills, although the latter remained in the low range of data available in literature. Major proteomic changes included (i) a slowdown in metabolic and/or cellular processes in muscles plus ganglia pool associated with minor toxicological effect and (ii) an increase of metabolic and/or cellular processes in gills associated with a greater toxicological effect. The nature of the proteomic changes is discussed in terms of unequal PAH distribution and allows to propose a set of explanatory mechanisms to associate behaviour to underlying physiological changes following oil exposure. First, the first tissues facing contaminated water are the inhalant siphon, the mantle edge and the gills. The routine nervous activity in the visceral ganglia should be modified by nervous information originating from these tissues. Second, the nervous activity in the visceral ganglia could be modified by its own specific contamination. Third, a decrease in nervous activity of the cerebral ganglia close to the mouth, including some kind of narcosis, could contribute to a decrease in visceral ganglia activity via a decrease or blockage of the downward neuromodulation by the cerebro-visceral connective. This whole set of events can explain the decrease of metabolic activity in the adductor muscles, contribute to initiate the catch mechanism and then deeply modify the valve behaviour.

Asiatic clam Corbicula fluminea exhibits distinguishable behavioural responses to crude oil under semi-natural multiple stress conditions.

Aquatic ecosystems are subject to many anthropogenic disturbances, and understanding their possible impacts is a real challenge. Developing approaches based on the behaviour of bivalve mollusks, an integrating marker of the state of the organisms, and therefore of their environment, is relevant, whether within a natural ecosystem or an ecosystem subject to industrial activities. The main objective of this study was to identify by HFNI Valvometry a reliable and reproducible clam behavioural response in the presence of crude oil in a multistress context. To closely replicate actual field conditions, Corbicula fluminea was exposed in outdoor artificial streams that were subject to natural variations and were continuously fed by fresh water from the Gave de Pau (S.W. France). After a period of 26 days in these artificial streams, the clams (n = 14-16 per condition) were separately exposed for 10 days to crude oil alone, crude oil and barium, crude oil and noise pollution, crude oil and turbidity pulses, barium alone, noise pollution alone, turbidity pulses alone or natural changes alone. The secondary objective was to characterize the accumulation of polycyclic aromatic hydrocarbons (PAH) in 3 tissues (gills, adductor muscles and foot) in clams exposed for 10 days to crude oil alone or under multistress conditions (n = 5 clams per condition) and then to compare the accumulation and behaviour of clams under these conditions. The response of clams to crude oil alone or under multistress conditions was visually and statistically significant and not confounded by the other disturbances tested, despite large variations in water temperature. In the presence of crude oil, the behaviour of clams was characterized by an increase in valve-closure duration, a decrease in valve-opening amplitude and an increase in valve agitation index. In the presence of crude oil, the clam behaviour showed no direct relationship with PAH accumulation in the gills, adductor muscles or foot, although hypothetical mechanisms are discussed. This work supports the growing interest in studying the behaviour of bivalve mollusks in the context of biomonitoring of the aquatic environment surrounding oil facilities.

Feeding inhibition in Corbicula fluminea (O.F. Muller, 1774) as an effect criterion to pollutant exposure: Perspectives for ecotoxicity screening and refinement of chemical control.

Bivalves are commonly used in biomonitoring programs to track pollutants. Several features, including its filter-feeding abilities, cumulatively argue in favour of the use of the Asian clam (Corbicula fluminea) as a biosentinel and an ecotoxicological model. Filtration in bivalves is very sensitive to external stimuli and its control is dictated by regulation of the opening/closure of the valves, which may be used as an avoidance defence against contaminants. Here, we investigate the filter-feeding behaviour of the Asian clam as an endpoint for assessing exposure to pollutants, driven by two complementary goals: (i) to generate relevant and sensitive toxicological information based on the ability of C. fluminea to clear an algal suspension, using the invasive species as a surrogate for native bivalves; (ii) to gain insight on the potential of exploring this integrative response in the refinement of chemical control methods for this pest. Clearance rates and proportion of algae removed were measured using a simple and reproducible protocol. Despite some variation across individuals and size classes, 50-90% of food particles were generally removed within 60-120 min by clams larger than 20 mm. Removal of algae was sensitive to an array of model contaminants with biocide potential, including fertilizers, pesticides, metals and salts: eight out of nine tested substances were detected at the µg l-1 or mg l-1 range and triggered valve closure, decreasing filter-feeding in a concentration-dependent manner. For most toxicants, a good agreement between mortality (96 h - LC50 within the range 0.4-5500 mg l-1) and feeding (2 h - IC50 within the range 0.005-2317 mg l-1) was observed, demonstrating that a 120-min assay can be used as a protective surrogate of acute toxicity. However, copper sulphate was very strongly avoided by the clams (IC50 = 5.3 µg l-1); on the contrary, dichlorvos (an organophosphate insecticide) did not cause feeding depression, either by being undetected by the clams' chemosensors and/or by interfering with the valve closure mechanism. Such an assay has a large potential as a simple screening tool for industry, environmental agencies and managers. The ability of dichlorvos to bypass the Asian clam's avoidance strategy puts it in the spotlight as a potential agent to be used alone or combined with others in eradication programs of this biofouler in closed or semi-closed industrial settings.

[Impacts of Asian clams (Corbicula fluminea) on lake sediment properties and phosphorus movement].

To examine the impact of Corbicula fluminea on sediment properties and phosphorus dynamics across sediment-water interface in lake, the microcosm experiment was carried out with sediment and lake water from the estuary of Dapu River, a eutrophic area in Taihu Lake. Rhizon samplers were used to acquire pore water, and soluble reactive phosphorus (SRP) flux across sediment-water interface and sediment properties were determined. The activity of C. fluminea destroyed the initial sediment structure, mixed sediment in different depths, increased oxygen penetration depth, sediment water content, and total microbial activity in sediment. The downward movement of overlying water was enhanced by the activity of C. fluminea, which decreased Fe2+ in pore water by oxidation. The production of ferric iron oxyhydroxide adsorbed SRP from pore water and decreased SRP concentration in pore water, and this increased iron bound phosphorus in corresponding sediment. The emergence of C. fluminea accelerated SRP release from sediment to overlying water, and enhanced SRP flux increased with the rise of introduced C. fluminea density. Metabolization of C. fluminea might play an important role in accelerating SRP release.

Sediment toxicity and bioaccumulation of nano and micron-sized aluminum oxide.

Nano-aluminum oxide (Al(2)O(3)) is used commercially in coatings and abrasives. Nano-Al(2)O(3) can also be generated through the oxidation of nano-aluminum in military propellants and energetics. The purpose of the present study was to assess toxicity and bioaccumulation of nano-Al(2)O(3) to a variety of sediment organisms (Tubifex tubifex, Hyalella azteca, Lumbriculus variegatus, and Corbicula fluminea). The bioaccumulation and toxicity of nano-Al(2)O(3) was compared with that of micron-sized Al(2)O(3) to investigate potential size-related effects. Results of the present study show species-specific differences in relative bioaccumulation of nano and micron-sized Al(2)O(3). Significant toxic effects (survival and growth) were observed in H. azteca testing, but only at high concentrations unlikely to be found in the environment. Nano-Al(2)O(3) was found to be more toxic than micron-sized Al(2)O(3) to H. azteca survival in a 14-d study in which organisms were in direct contact with a thin layer of 625 or 2,500 mg of Al(2)O(3) dispersed on the surface of either sediment or sand. A significant growth effect was also observed for nano but not micron-sized Al(2)O(3) at the highest treatment level tested (100 g/kg Al(2)O(3)) in a 10-d H. azteca bioassay in which Al(2)O(3) was homogenized with sediment. However, differences in measured sediment Al concentrations (micron-sized = 55.1 [+/-0.6] g/kg Al; nano-sized = 66.2 [+/-0.6] g/kg Al) in the nano and micron-sized Al(2)O(3) preclude direct comparison of the toxicity of these two treatments based on particle size.

Accumulation of trace elements and growth responses in Corbicula fluminea downstream of a coal-fired power plant.

Lentic organisms exposed to coal-fired power plant (CFPP) discharges can have elevated trace element concentrations in their tissues, but this relationship and its potential consequences are unclear for lotic organisms. To explore these patterns in a lotic environment, we transplanted Corbicula fluminea from a reference stream to a stream receiving CFPP discharge. We assessed trace element accumulation and glutathione concentration in clam tissue, shell growth, and condition index at five sites along a contamination gradient. Clams at the most upstream and contaminated site had the highest growth rate, condition index, glutathione concentrations, and concentrations of arsenic (7.85+/-0.25 microg/g [dry mass]), selenium (17.75+/-0.80 microg/g), and cadmium (7.28+/-0.34 microg/g). Mercury concentrations declined from 4.33+/-0.83 to 0.81+/-0.11 microg/g [dry mass] in clams transplanted into the selenium-rich environment nearest the power plant, but this effect was not as evident at less impacted, downstream sites. Even though dilution of trace elements within modest distances from the power plant reduced bioaccumulation potential in clams, long-term loading of trace elements to downstream depositional regions (e.g., slow moving, silty areas) is likely significant.

Impact of hypoxia on hemolymph contamination by uranium in an aquatic animal, the freshwater clam Corbicula fluminea.

Multi-stress situations are a major question and low-oxygenated waters (hypoxia) are a growing problem. Importantly, hypoxia stimulates the ventilatory flow rate in aquatic animals and this increases gill exposure to contaminants. Surprisingly, in the freshwater clam Corbicula fluminea, this is associated with increased bioaccumulation of uranium in gills but not in deep tissues. We searched for an explanation by analyzing hemolymph U-transport in Corbicula exposed to 0.36 microM dissolved uranium at various O2-levels for 10 days. In hypoxia, one observed an increased U concentration in the arterial hemolymph flowing from gills to tissues but this was not associated with an increased U concentration in the venous hemolymph nor in the other tissues. We conclude that the cardiac flow rate must have decreased to explain this absence of over-accumulation. In addition to its already known deleterious effects, uranium can thus deeply impair cardiac flow rate in exposed aquatic animals during multi-stress exposures.

Selenium bioaccumulation in Chlamydomonas reinhardtii and subsequent transfer to Corbicula fluminea: role of selenium speciation and bivalve ventilation.

The uptake of Se by the freshwater alga Chlamydomonas reinhardtii and the subsequent transfer to the Asiatic clam Corbicula fluminea was investigated. The objective was to investigate the bioavailability of algal-bound Se for C. fluminea while taking into account Se speciation and bivalve ventilation. First, uptake rates of waterborne Se (selenite, selenate, and selenomethionine) in the algae during a 1-h exposure period were determined for a range of concentrations up to 2,000 microg/L. Fluxes for selenite uptake were constant in the range of concentrations tested, whereas fluxes for selenate and selenomethionine uptake decreased with increasing concentrations, suggesting a saturated transport system at high concentrations (approximately 1,000 microg/L for selenate and 100 microg/L for selenomethionine). These data were used to set the algal contamination for the study of trophic transfer to the clam. Three parameters were studied: The Se form, the algal density, and the Se burden in the algae. The results show that for a fixed algal density, an Se-contaminated algal diet does not modify ventilation. In this case, the driving factor for ventilation is the algal density, with ventilation being enhanced for low algal densities. On the basis of ventilatory flow rate measurements and Se burdens in algae, it was found that bioaccumulation of Se in C. fluminea was proportional to the total quantity of Se passing through the whole organism, but with a lesser extraction coefficient for selenomethionine than for the inorganic forms. These results underline the importance of both physiological factors and speciation in understanding the trophic transfer of Se.

Modulation of uranium bioaccumulation by hypoxia in the freshwater clam Corbicula fluminea: induction of multixenobiotic resistance protein and heat shock protein 60 in gill tissues.

The influence of hypoxia on the bioaccumulation of uranium in the clam Corbicula fuminea was investigated in ecologically relevant conditions. The cellular impact at the gill-tissue level was assessed by analyzing the induction of multixenobiotic resistance protein (MXR) and heat shock protein 60. Analyses were performed at three biological levels. First, at the organism level, uranium induced a significant decrease in the valve open duration under normoxia, but not under hypoxia, in which oxygen drive imposed an increase of the valve open duration. Second, at the tissue level, the uranium bioaccumulation rate in the gills was higher under hypoxia than under normoxia. Third, at the cellular level, MXR was induced by uranium but not by hypoxia. The threshold of tissular uranium concentration triggering MXR induction was between 3 and 5 nmol/g. On the contrary, Hsp60 was induced by hypoxia but not by uranium.

Laboratory and field assessment of uranium trophic transfer efficiency in the crayfish Orconectes limosus fed the bivalve C. fluminea.

At present, ecotoxicological information regarding the impact of natural uranium (U) on freshwater ecosystems via the trophic contamination route is scarce. We generated an experimental trophic food chain involving the prey species, Corbicula fluminea, and a predator, Orconectes limosus, for a 10-day and a 30-day feeding periods (food ration: one whole soft body/day/crayfish). We studied the efficiency of U trophic transfer and the distribution of U in the predator. During the test, we varied the quantity of dietary U (from beforehand contaminated bivalves at concentrations ranging from 0.9+/-0.1 to 20.2+/-9 microg/g fw provided to each crayfish over the 10 days) applying a daily feeding rate equal to 3.9+/-0.8% fw. The efficiency of U trophic transfer from clams to crayfish varied between 1 and 13% depending on the prey exposure modalities. Accumulation of U was observed in the digestive gland but also in gills, in the muscle, and in the molt of the crayfish after trophic exposure treatments. Under high-level exposure conditions, the digestive gland was the main target-organ, however a significant accumulation was also observed in the stomach. With regard to low levels of trophic exposure, accumulation of U in gills, in the stomach, and in the digestive gland was of the same order of magnitude. Longer exposure period which incorporated a crayfish molt, resulted in a decrease of trophic transfer ratio and a modified U tissue distribution.