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Numerous actions have been undertaken by farmers to attenuate the impact of agricultural activities on aquatic ecosystems. The identification of biomarkers that respond quickly to water quality improvement could facilitate the assessment of adopted alternative practices and help maintain mobilization among stakeholders. We evaluated the potential of the comet assay, a biomarker of genotoxic effects, using a freshwater mussel, Elliptio complanata, as a model animal. The frequency of DNA damage was assessed in hemocytes of mussels collected from a pristine habitat and caged for 8 weeks in the Pot au Beurre River, a tributary of the fluvial Lake St.-Pierre (Quebec, Canada) impacted by agricultural activities. We found that the level of DNA damage naturally induced in mussel hemocytes was low and showed very limited variations over time. Compared with these baseline levels and to laboratory controls, we observed a doubling in DNA alterations in mussels exposed to agricultural runoff in the third branch of the Pot au Beurre River. The genotoxic response was significantly lower in mussels caged in the first branch of the Pot au Beurre River, where longer stretches of shoreline have been restored as buffer strips. Glyphosate, mesotrione, imazethapyr, and metolachlor were the main discriminant pesticides between these two branches. Metolachlor was found in sufficient concentrations to induce DNA damage, but it is more likely that the observed genotoxicity was the result of a "cocktail effect," that is, the cumulative contribution of coexisting genotoxicants including the above-mentioned herbicides and ingredients in their formulation. Our findings suggest that the comet assay is a sensitive tool for the early detection of changes in water toxicity following the adoption of agricultural beneficial practices. Environ Toxicol Chem 2023;42:2201-2214. © 2023 Crown copyright and The Authors. Environmental Toxicology and Chemistry published by Wiley Periodicals LLC on behalf of SETAC. This article is published with the permission of the Controller of HMSO and the King's Printer for Scotland., (© 2023 Crown copyright and The Authors. Environmental Toxicology and Chemistry published by Wiley Periodicals LLC on behalf of SETAC. This article is published with the permission of the Controller of HMSO and the King's Printer for Scotland.)

Atlantic salmon is an important species for Canadian culture and economy and its importance extends beyond Canada to Scandinavia and Western Europe. However, it is a vulnerable species facing decline due to habitat contamination and destruction. Existing and new Canadian pipeline projects pose a threat to salmonid habitat. The effects of diluted bitumen (dilbit), the main oil circulating in pipelines, are less studied than those of conventional oils, especially during the critical early embryonic developmental stage occurring in freshwater ecosystems. Therefore, this study aimed to compare the effects of water-accommodated fractions (WAF) of the Clearwater McMurray dilbit and the Lloydminster Heavy conventional oil on Atlantic salmon embryos exposed either from fertilization or from eyed stage. The dilbit contained the highest concentrations of low molecular weight (LMW) compounds (including BTEX and C 6 -C 10 ), while the conventional oil contained the highest concentrations of PAHs. The Clearwater dilbit caused a higher percentage of mortality and malformations than the conventional oil at similar WAF concentrations. In addition, the embryos exposed from fertilization suffered a higher mortality rate, more developmental delays, and malformations than embryos exposed from the eyed stage, suggesting that early development is the most sensitive developmental stage to oil exposure. Gene expression and enzymatic activity of the detoxification phase I and II enzymes (CYP1A and GST) were measured. Data showed increases in both cyp1a expression and GST activity with increasing WAF concentrations, while gst expression was not affected by the exposures. Also, gene expression of proteins involved in the biotransformation of vitamin A and DNA damage repair were modified by the oil exposures. Overall, this study indicates that Atlantic salmon is mostly affected by oil exposure at the beginning of its development, during which embryos accumulate deformities that may impact their survival at later life stages., Competing Interests: Declaration of Competing Interest The authors declare that they have no known competing financial interests or personal relationships that could have appeared to influence the work reported in this paper., (Copyright © 2022. Published by Elsevier Inc.)

The oil sands industry in Canada, produces heavy unconventional oils, diluted for transport and called diluted bitumen. However, despite advances in our knowledge of the ecotoxicological risk that these products represent, their effects on benthic organisms following a spill are still largely unknown. In order to fill these gaps, this study aims to determine the lethal and sublethal effects of two diluted bitumens (Bluesky and Cold Lake) and one conventional oil (Lloydminster) for two freshwater benthic invertebrates: Chironomus riparius and Hyalella azteca. The objective of this study is to assess the toxicity of dissolved hydrocarbons, resulting from the physical dispersion of oil, immediately after a spill on the benthic invertebrates. To this end, organisms were exposed for 7 days for chironomids and 14 days for amphipods to a fraction containing soluble hydrocarbons (WAF: water accommodated fraction; 10 g/L, 18 h of agitation, followed by 6 h of sedimentation) with natural or artificial sediment. After exposure, the effects of hydrocarbons were determined using size, mortality, and antioxidant capacities. Dissolved hydrocarbons induced mortality for both species, but these hydrocarbons disappeared very quickly from the water column, regardless of the oil type. The amphipods were sensitive to both types of oil while the chironomids were only sensitive to diluted bitumens. The presence of a natural sediment seems to provide a protective role against dissolved hydrocarbons. The antioxidant enzymes measured (CAT, SOD and GPx) do not appear to be relevant biomarkers for the exposure of these organisms to diluted bitumen., (Crown Copyright © 2022. Published by Elsevier Inc. All rights reserved.)

Canadian freshwater ecosystems are vulnerable to oil spills from pipelines, which contain mostly diluted bitumen. This study aimed to compare the toxicity of a dilbit and a conventional oil on developing rainbow trout. A total of five exposure scenarios were performed, from 10 to 43 days, using water-accommodated fraction (WAF) with an initial loading of 1:9 oil to water ratio (w/v) in a range of dilutions from 0.32 to 32% WAF, respectively, with TPAH and VOC concentrations from 2.41 to 17.5 μg/L and 7.94-660.99 μg/L, and with or without a recovery period. Following the five exposures, several endpoints were examined, including survivorship, morphometrics, gene expression, and enzymatic activity. Significant mortality rates were measured for the highest WAF concentration of the dilbit in all five exposures (60-100% mortality at 32% WAF). In comparison, the highest WAF concentration of the conventional oil induced significant mortality in three out of the five exposure (from 35 to 100% mortality at 32% WAF). Hatching delays were noted in embryos exposed to both oils. Developmental delays were observed in dilbit-exposed embryos and are suspected to be an indicator of reduced survivorship after hatching. The induced expression of cyp1a remained a reliable biomarker of exposure and of fish malformations, though it did not always predict mortality. Using CYP1A activity in combination with cyp1a may bring more insights in studies of oil risk assessment. This study demonstrates that dilbits are more toxic to early life stages compared to conventional oils and highlights the need to consider the most sensitive stage of development when performing risk assessment studies on oils., (Copyright © 2021 Elsevier Ltd. All rights reserved.)

Petroleum hydrocarbons are widely used and transported, increasing the risks of spills to the environment. Although conventional oils are the most commonly produced, the production of unconventional oils (i.e. diluted bitumen or dilbit) is increasing. In this study, we compared the effects of conventional oils (Arabian Light and Lloydminster) and dilbits (Bluesky and Clearwater) on early life stages of a salmonid. To this end, aqueous fractions (WAF: water accommodated fraction) of these oils were extracted using mountain spring water. Rainbow trout (Oncorhynchus mykiss) larvae were exposed to 10 and 50% dilutions of these WAFs from hatching (340 DD; degree days) until yolk sac resorption (541 DD). Exposure to WAFs increased skeletal malformations (both dilbits) and hemorrhage (both conventional oils and Bluesky) and decreased head growth (Arabian Light). In addition, increases in EROD activity and DNA damage were measured for all oils and an increase in cyp1a gene expression was measured for Arabian Light, Bluesky and Clearwater. The PAH and C 10 C 50 concentrations were positively correlated to total larval EROD activity, whereas concentrations of total hydrocarbons, VOCs, PAHs, and C 10 C 50 were positively correlated to cyp1a expression. Total hydrocarbon, VOC, and C 10 C 50 concentrations were also negatively correlated to larval growth. This study supports that petroleum hydrocarbons are toxic to early developmental stages of rainbow trout and show that their degree and spectrum of toxicity depends on their chemical composition., (Copyright © 2021. Published by Elsevier B.V.)