DNA integrity assessment in hemocytes of soft-shell clams (Mya arenaria) in the Saguenay Fjord (Québec, Canada).

The purpose of this study was to examine the effects of pollution on DNA integrity in the feral soft-shell clam (Mya arenaria) in the Saguenay Fjord. Intertidal clams were collected downstream and upstream of the fjord at sites under anthropogenic pollution. DNA integrity was assessed by following changes in single- and double-stranded breaks, variation in DNA content and micro-nuclei (MN) incidence in hemocytes. The results revealed that clams collected at polluted sites had reduced DNA strand breaks (lower DNA repair activity), increased DNA content variation and MN frequency in hemocytes. The data revealed that DNA content variation was closely related to MN frequency and negatively with DNA strand breaks formation. Water conductivity was also related to reduced MN frequency and DNA content variation, indicating that, in addition to the effects of pollution, the gradual dilution of saltwater could compromise mussel health.

Ecotoxicological impacts of effluents generated by oil sands bitumen extraction and oil sands lixiviation on Pseudokirchneriella subcapitata.

The exploitation of Athabasca oil sands deposits in northern Alberta has known an intense development in recent years. This development has raised concern about the ecotoxicological risk of such industrial activities adjacent to the Athabasca River. Indeed, bitumen extraction generated large amounts of oil sands process-affected water (OSPW) which are discharged in tailing ponds in the Athabasca River watershed. This study sought to evaluate and compare the toxicity of OSPW and oil sands lixiviate water (OSLW) with a baseline (oil sands exposed to water; OSW) on a microalgae, Pseudokirchneriella subcapitata, at different concentrations (1.9, 5.5, 12.25, 25 and 37.5%, v/v). Chemical analyses of water-soluble contaminants showed that OSPW and OSLW were enriched in different elements such as vanadium (enrichment factor, EF=66 and 12, respectively), aluminum (EF=64 and 15, respectively), iron (EF=52.5 and 17.1, respectively) and chromium (39 and 10, respectively). The toxicity of OSPW on cells with optimal intracellular esterase activity and chlorophyll autofluorescence (viable cells) (72h-IC 50%<1.9%) was 20 times higher than the one of OSW (72h-IC 50%>37.5%, v/v). OSLW was 4.4 times less toxic (IC 50%=8.5%, v/v) than OSPW and 4.5 times more toxic than OSW. The inhibition of viable cell growth was significantly and highly correlated (<-0.7) with the increase of arsenic, beryllium, chromium, copper, lead, molybdenum and vanadium concentrations. The specific photosynthetic responses studied with JIP-test (rapid and polyphasic chlorophyll a fluorescence emission) showed a stimulation of the different functional parameters (efficiency of PSII to absorb energy from photons, size of effective PSII antenna and vitality of photosynthetic apparatus for energy conversion) in cultures exposed to OSPW and OSLW. To our knowledge, our study highlights the first evidence of physiological effects of OSPW and OSLW on microalgae.

Differential changes in gene expression in rainbow trout hepatocytes exposed to extracts of oil sands process-affected water and the Athabasca River.

The oil sands region of northern Alberta represents the world's largest reserves of bitumen, and the accelerated pace of industrial extraction activity has raised concern about the possible impacts on the Athabasca River and its tributaries. An ecotoxicogenomic study was undertaken on Oncorhynchus mykiss trout hepatocytes exposed to extracts of water samples near the oil sand development area, as well as to oil sands process-affected water (OSPW) extracts using the quantitative reverse transcriptase polymerase chain reaction technique. The expression of the following genes (mRNA) was monitored to track changes in xenobiotic biotransformation (CYP1A1, CYP3A4, glutathione S-transferase, multi-drug resistance transporter), estrogenicity (estrogen receptor and vitellogenin), oxidative stress (superoxide dismutase and metallothionein) and DNA repair activity (DNA ligase). The extent of DNA-aromatic hydrocarbon adducts was also determined in cells by immuno-staining. A comparative analysis of gene expression between the river/lake and OSPW samples revealed that CYP3A4, metallothioneins, DNA ligase and GST genes, were specifically expressed by OSPW. Cells exposed to OSPW, commercial naphthenic acids, and benzo(a)pyrene showed increased polyaromatic hydrocarbon DNA-adducts, as determined by cell immunofluorescence analysis. Other genes were induced by all types of water samples, although the induction potential was stronger in OSPW most of the time (e.g., VTG gene was expressed nearly 15-fold by surface waters from the lake and river samples but increased to a maximum of 31-fold in OSPW). A multivariate discriminant function analysis revealed that the lake and river water samples were well discriminated from the OSPW. The CYP3A4 gene was the most highly expressed gene in cells exposed to OSPW and responded less to the lake or river water in the Athabasca River area. This study identified a suite of gene targets that responded specifically to OSPW extracts, which could serve as toxicogenomic fingerprints of OSPW contamination.

Ecotoxicity of a brominated flame retardant (tetrabromobisphenol A) and its derivatives to aquatic organisms.

The large use of tetrabromobisphenol A (B(4)BPA) in common products (plastics, electric and electronic equipments) has raised concern about its ecotoxicity. Physical and bio-degradations may lead to the formation of tetrabromobisphenol A derivatives like tri- (B(3)BPA), di- (B(2)BPA), monobromobisphenol A (B(1)BPA) and bisphenol A (BPA). However, little is known about the toxicity of these brominated derivatives. An appraisal on the ecotoxicity of B(4)BPA and its derivatives was carried out with several bioassays representing organisms (bacteria, algae, micro-invertebrates and fish) of different taxonomic groups present in aquatic ecosystems. Endpoint values showed that B(4)BPA was significantly less toxic than the other chemicals when tested with the Microtox and algal asssays. A similar trend was observed with other bioassays for BPA. One of the brominated derivatives was particularly toxic: B(2)BPA. The LuminoTox assay and the rainbow trout hepatocytes assay reported the most significant toxicity for this derivative. Its toxicity was also significantly higher than the other compounds barring B(3)BPA when tested with the micro-crustacean test.

Sensitivity of freshwater periphytic diatoms to agricultural herbicides.

The biomonitoring of pesticide pollution in streams and rivers using algae such as diatoms remains difficult. The responses of diatom communities to toxic stress in stream water are disturbed by the variations of environmental parameters. In this study, periphytic algae collected in situ were exposed under controlled conditions to two major herbicides used in French agriculture (isoproturon and s-metolachlor). Three exposure regimes were tested: 5 and 30 microg L(-1) for 6 days and 30 microg L(-1) for 3 days followed by a recovery period of 3 days. The algal biomasses were assessed from pigment concentrations (chlorophyll a and c) and from live cell density. The highest concentration (30 microg L(-1)) of isoproturon inhibited the biomass increase statistically significantly. In periphyton exposed to 5 and 30 microg L(-1) of s-metolachlor, chlorophyll c concentration and live cell density were also statistically significantly lower than in the control. Periphyton left to recover after reduced exposure duration (3 days) showed higher growth rates after treatment with s-metolachlor than with isoproturon. Taxonomic identifications showed that species like Melosira varians, Nitzschia dissipata and Cocconeis placentula were not affected by the herbicide exposure. Other species like Eolimna minima and Navicula reichardtiana were more sensitive. Studying diatoms according to their trophic mode showed that facultative heterotroph species were statistically significantly favoured by isoproturon exposure at the highest concentration. Results obtained with s-metolachlor exposure showed a disturbance of cell multiplication rather than that of photosynthesis. These results suggest that photosynthesis inhibitors like isoproturon favour species able to survive when the autotroph mode is inhibited.

Herbicide effects on freshwater benthic diatoms: induction of nucleus alterations and silica cell wall abnormalities.

Benthic diatoms are well known bio-indicators of river pollution by nutrients (nitrogen and phosphorus). Biological indexes, based on diatom sensitivity for non-toxic pollution, have been developed to assess the water quality. Nevertheless, they are not reliable tools to detect pollution by pesticides. Many authors have suggested that toxic agents, like pesticides, induce abnormalities of the diatom cell wall (frustule). High abnormal frustule abundances have been reported in natural diatom communities sampled in streams contaminated by pesticides. However, no direct link was found between the abundances of abnormal frustules in these communities and the pesticide concentrations in stream water. In the present study, a freshwater benthic diatom community, isolated from natural biofilm and cultured under controlled conditions, was treated with a known genotoxic herbicide, maleic hydrazide (MH). Cells were exposed to three concentrations of MH (5x10(-6), 10(-6), 10(-7)M) for 6h followed by a 24h-recovery time. After MH treatments, nucleus alterations were observed: abnormal nucleus location, micronucleus, multinuclear cell or disruption of the nuclear membrane. A dose-dependent increase of nuclear alterations was observed. The difference between the control (9.65 nuclear alterations per 1000 cells observed (9.65 per thousand), S.D.=4.23) and the highest concentrations (29.40 per thousand, S.D.=8.49 for 10(-6)M and 35.96 per thousand, S.D.=3.71 for 5x10(-6)M) was statistically significant (Tukey test, P<0.05). Diatoms also exhibited frustules with deformed morphology and abnormal ornamentation. Significantly increased abundances of abnormal frustules were observed for the highest concentrations (10(-6) and 5x10(-6)M; Tukey test, P<0.05). These two parameters tended to increase together (Pearson correlation=0.702, P<0.05). The results suggest that the induction of abnormal frustules could be associated with the genotoxic effects of MH. The alterations observed could be related to the effects of MH on the synthesis of the proteins involved in frustule formation or in the regulation of the cytoskeleton of the diatom cells.