Water-accommodated Fraction Research Articles

Physical Evidence of Oil Uptake and Toxicity Assessment of Amphiphilic Grafted Nanoparticles Used as Oil Dispersants

Herein, we report the toxicity evaluation of a new prototype dispersant system, silicon dioxide nanoparticles (NPs) functionalized with (3-glycidoxypropyl)triethoxysilane (GPS) and grafted poly(ε-caprolactone)-block-poly[oligo(ethylene glycol)methyl methacrylate mono-methyl ether] (NP-PCL-POEGMA). This serves as a follow up of our previous study where grafted silicon dioxide NPs functionalized with GPS and grafted hyperbranched poly(glycidol) (NP-HPG) were evaluated for reducing the toxicity in embryo, juvenile, and adult fish populations. In this study, the NP-HPG sample is used as a baseline to compare against the new NP-PCL-POEGMA samples. The relative size was established for three NP-PCL-POEGMA samples via cryogenic transmission electron microscopy. A quantitative mortality study determined that these NPs are non-toxic to embryo populations. An ethoxyresorufin-O-deethylase assay was performed on these NP-PCL-POEGMA samples to test for reduced cytochrome P450 1A after the embryos were exposed to the water-accommodated fraction of crude oil. Overall, these NP-PCL-POEGMA NPs better protected the embryo populations than the previous NP-HPG sample (using a protein activity end point), showing a trend in the right direction for prototype dispersants to replace the commercially utilized Corexit.

Legacy and dispersant influence microbial community dynamics in cold seawater contaminated by crude oil water accommodated fractions

Dispersants, used for combating oil spills, increase hydrocarbon bioavailability promoting their biodegradation. Oil weathering process introduces harmful soluble hydrocarbons, such as polycyclic aromatic hydrocarbons (PAHs), into the water column, resulting in water-accommodated fraction (WAF). The presence of dispersants can influence the weathering process by increasing PAHs solubility, toxicity and biodegradability. However, little is known on how dispersants affect microbial communities and their degradation capacities, especially in cold environment where low temperature decreases microbial activity and thus hydrocarbon degradation. Here, we investigated the microbial community dynamics in cold water contaminated by WAF prepared from crude oil with or without a commercial dispersant (Finasol OSR52).The WAFs, prepared with Naphthenic North Atlantic crude oil, were used to contaminate seawater from Norwegian cold sites, one oil-contaminated and the other pristine. The WAF-contaminated seawaters were maintained in microcosms at 4 °C for 21 days. The content of PAHs and microbial compositions (16S rRNA gene sequencing) were determined at days 0, 7, 14 and 21. In addition, the 96 h toxicity assay with adult Acartia tonsa revealed WAFs toxicity at days 0 and 21.The toxicity of WAF mixtures, with and without dispersant, against Acartia tonsa was reduced during the experiment, but PAHs removal was not increased. The water from the oil-contaminated site showed the highest PAHs removal revealing legacy effect (presence of microorganisms adapted to PAHs). Additionally, our results reveal: i) microbial community plasticity allowing the adaptation to the presence of PAHs and dispersant, ii) specific bacteria taxa probably involved in PAHs degradation, and iii) dispersants shape the microbial communities dynamics by stimulating potential dispersant-degrading taxa, such as Fusibacter. Thus, our results provide valuable insights on the role of microbial community in determining the fate of water-solubilized hydrocarbon in cold environment while questioning the role of dispersant used for fighting oil spill.

Chemical characterization of oil and water accommodated fraction (WAF) at different temperatures

In this work, the formation and the chemical composition of Water Accommodated Fractions (WAFs) of Intermediate Fuel Oil 180, Naphthenic North Atlantic, and Marine Gas-Oil oils were studied at different temperatures (between 5 and 25 °C) with and without a dispersant (Finasol OSR52). In terms of composition, both the raw oils and the WAFs showed high levels of lighter PAHs (naphthalene related compounds). The WAFs were prepared under slow stirring rate to avoid the formation of droplets. However, the use of dispersant increased the solubility of the PAHs around two orders of magnitude through the formation of small droplets. The kinetics of the WAF formation was followed by direct measurement on the water phase, and through the accumulation on PDMS sheets and commercial stir-bars. This study showed that the water phase achieved the steady state after 48 h of continuous stirring, while the PDMS sheet and Twister required slightly longer times. It was also observed that the solubility of the PAHs increased with the temperature, almost doubling the concentrations in the studied ranged but when the dispersant was used, only Naphthenic North Atlantic crude oil followed this pattern. Finally, the stability of WAF was monitored at room temperature and it was concluded that the concentrations halved after 24–30 h, regardless the use of dispersant. Therefore, for WAF-based toxicity testing, it is important to renew these solutions at least once or twice a day to assure the stable concentrations along the bioassay.

Changes in Chemical Composition and Copepod Toxicity during Petroleum Photo-oxidation.

Photoproducts can be formed rapidly in the initial phase of a marine oil spill. However, their toxicity is not well understood. In this study, oil was irradiated, chemically characterized, and tested for toxicity in three copepod species (Acartia tonsa, Temora longicornis, and Calanus finmarchicus). Irradiation led to a depletion of polycyclic aromatic hydrocarbons (PAHs) and n-alkanes in oil residues, along with an enrichment in aromatic and aliphatic oil photoproducts. Target lipid model-based calculations of PAH toxicity units predicted that PAH toxicities were lower in water-accommodated fractions (WAFs) of irradiated oil residues ("irradiated WAFs") than in WAFs of dark-control samples ("dark WAFs"). In contrast, biomimetic extraction (BE) measurements showed increased bioaccumulation potential of dissolved constituents of irradiated WAFs compared to dark WAFs, mainly driven by photoproducts present in irradiated oil. In line with the BE results, copepod mortality increased in irradiated WAFs compared to dark WAFs. However, low copepod toxicities were observed for WAFs produced with photo-oxidized oil slicks collected during the Deepwater Horizon oil spill. The results of this study suggest that while oil photoproducts have the potential to be a significant source of copepod toxicity, dilution and dispersion of these higher solubility products appear to help mitigate their toxicity at sea.

Physicochemical characteristics and toxicity studies of crude oil, dispersant and crude oil-- dispersant test media to marine organisms

In this study, the physicochemical characteristics of crude oil, dispersant and crude oil-dispersant test systems and their toxicities on representative marine organisms in a simulated marine ecosystem was assessed. Mechanically dispersed crude oil-in-water test system (MDO) and its water accommodated fraction (WAF) alone, and in combination with a third generation dispersant (D) (Ecobest®), which gave rise to chemically dispersed crude oil-in-water test system (CDO) and its water accommodated fraction (CEWAF), as well as a reference toxicant, sodium dodecyl sulphate (SDS) were used to evaluate toxicities on selected marine organisms (Fish – Tilapia guineensis, Crustacean – Palaeomontes africanus, and Bacteria – Total heterotrophic bacteria and hydrocarbon utilizing bacteria). Physicochemical characteristics were done using standard methods. The static with renewal bioassay option was employed for toxicity tests involving Fish and Crustacean, while the static without renewal option was used for microbial bioassays. Marine organisms were exposed to the following concentrations: 1000, 500, 250, 125, 62.5 and 0 mg/L of MDO, CDO, D, WF and CEWAF, respectively. The LC50 (lethal concentration that resulted in 50% mortality) at 96h and toxicity factors were determined. Results for physicochemical characteristics of the test systems showed that the pH, dissolved oxygen levels were sufficient for sustaining aquatic habitation. Lead metal was present in high amounts in the dispersant test system, but relatively low in chemically dispersed test systems. Cd and Ni were also present at relative amounts that could be toxic to aquatic lives. Toxicity data showed that the new dispersant, Ecobest® was non-toxic to the tested marine organisms relative to the reference toxicant (SDS), which was moderately toxic. The 96h LC50 of MDO, CDO, D, WAF, CEWAF and SDS were 89.8, 225.6, 1891.8, 683.9, 528.1 and 1.37 for T. guineensis, and 275.5, 137.7, 3800.9, 76.1, 168.3 and 9.99 for P. africanus, and 1658.5, 944.1, 17221.9, 228641.0, 1036319.3 and 3.84 for total heterotrophic bacteria. The reference toxicant, SDS, was the most toxic of the toxicants, but the dispersant reduced the toxicity y several folds. However, with the presence of some heavy metals, notably Pb, Cd and Ni, and some levels of polycyclic aromatic hydrocarbons in the test media water, there may be likelihood for bioaccumulation to occur in the tissues of marine organisms.

Impact of mixing and resting times on the droplet size distribution and the petroleum hydrocarbons’ concentration in diluted bitumen-based water-accommodated fractions (WAFs)

The preparation of Water-accommodated Fractions (WAFs) and chemically enhanced WAFs (CEWAFs) are essential for evaluating oil toxicity. The Chemical Response to Oil Spills: Ecological Research Forum (CROSERF) method was widely adopted, with variables (e.g., mixing time, oil loading, etc.) being continuously changed among research groups, which limits the cooperation in this area. Herein, we conducted WAF and CEWAF experiments using two loadings of diluted bitumen (Dilbit): 1 g/L and 10 g/L. For the CEWAF, the dispersant to oil ratio was 1:20. We investigated the impact of three mixing durations (18 h, 42 h, and 66 h) and two resting times (6 h and 24 h) on the droplet size distribution (DSD) and accommodated oil concentration. This would be highly beneficial for analyzing toxicity from oil spills, especially when considering the toxic effect of both suspended oil droplets and dissolved hydrocarbons. The DSD results and oil chemistry analysis showed that at a low oil loading concentration (1 g/L), both WAFs and CEWAFs had the same DSD, with an average d50 (volume median diameter) of 3.38 ± 0.70 μm and 3.85 ± 0.63 μm, respectively. At a high oil loading concentration (10 g/L), the WAFs had an average d50 of 3.69 ± 0.52 μm, showing no correlation with mixing and resting time. The DSD of CEWAFs increased significantly at 42 h mixing and 24 h resting time, with oil concentration reaching equilibrium after 42 h mixing. Therefore, WAFs appears to require only 18 h mixing and 6 h resting, while it is recommended to have 42 h mixing and 24 h resting for CEWAFs at high dilbit oil loading concentrations.

Dispersed Crude Oil Induces Dysbiosis in the Red Snapper Lutjanus campechanus External Microbiota.

ABSTRACTThe fish external microbiota competitively excludes primary pathogens and prevents the proliferation of opportunists. A shift from healthy microbiota composition, known as dysbiosis, may be triggered by environmental stressors and increases host susceptibility to disease. The Deepwater Horizon (DWH) oil spill was a significant stressor event in the Gulf of Mexico. Despite anecdotal reports of skin lesions on fishes following the oil spill, little information is available on the impact of dispersed oil on the fish external microbiota. In this study, juvenile red snapper (Lutjanus campechanus) were exposed to a chemically enhanced water-accommodated fraction (CEWAF) of Corexit 9500/DWH oil (CEWAF) and/or the bacterial pathogen Vibrio anguillarum in treatments designed to detect changes in and recovery of the external microbiota. In fish chronically exposed to CEWAF, immunoglobulin M (IgM) expression significantly decreased between 2 and 4 weeks of exposure, coinciding with elevated liver total polycyclic aromatic hydrocarbons (PAHs). Dysbiosis was detected on fish chronically exposed to CEWAF compared to seawater controls, and addition of a pathogen challenge altered the final microbiota composition. Dysbiosis was prevented by returning fish to clean seawater for 21 days after 1 week of CEWAF exposure. Four fish exhibited lesions during the trial, all of which were exposed to CEWAF but not all of which were exposed to V. anguillarum. This study indicates that month-long exposure to dispersed oil leads to dysbiosis in the external microbiota. As the microbiota is vital to host health, these effects should be considered when determining the total impacts of pollutants in aquatic ecosystems.IMPORTANCE Fish skin is an immunologically active tissue. It harbors a complex community of microorganisms vital to host homeostasis as, in healthy fish, they competitively exclude pathogens found in the surrounding aquatic environment. Crude oil exposure results in immunosuppression in marine animals, altering the relationship between the host and its microbial community. An alteration of the healthy microbiota, a condition known as dysbiosis, increases host susceptibility to pathogens. Despite reports of external lesions on fishes following the DWH oil spill and the importance of the external microbiota to fish health, there is little information on the effect of dispersed oil on the external microbiota of fishes. This research provides insight into the impact of a stressor event such as an oil spill on dysbiosis and enhances understanding of long-term sublethal effects of exposure to aid in regulatory decisions for protecting fish populations during recovery.

Chemical and toxicological characterisation of residues from offshore in-situ burning of spilled fuel oils

Burn residues collected after large scale experimental in situ burns performed in the North Sea were characterised with emphasis on chemistry and acute toxicity. Low-energy water accommodated fractions (WAFs) of three marine fuels (Ultra Low Sulphur Fuel Oil (ULSFO), bunker oil (IFO180), and marine gas oil (MGO) and their field-generated residues from in situ burning (ISB). were prepared to evaluate the potential impact of ISB residue to the environment. The toxicity effects on primary consumers were assessed by testing on early life stage (nauplii) of the marine copepod Calanus finmarchicus. Toxicity studies showed that ISB decreased the acute toxicity of the WAFs compared to the initial oils. WAF of MGO had highest toxicity, and ISB residue of MGO seems to be more toxic than WAFs of fresh ULSFO and IFO180. Additive toxicity expressed as toxic unit (TU) based on the chemical composition also indicated that the toxicity of WAFs from ISB residues were lower than for the initial oils. The 2−3 ring PAHs seem to contribute most to the TU. Overall, the three offshore burns reduced the total mass of PAHs in the water accommodated fractions by >90 % compared to the released unburned oils and caused a reduction of the acute toxicity to copepod nauplii.

Oil-mediated oxidative-stress responses in a keystone zooplanktonic species, Calanus finmarchicus

The copepod Calanus finmarchicus is an ecologically important species in the North Atlantic, Norwegian and Barents seas. Accidental or continuous petroleum pollution from oil and gas production in these seas may pose a significant threat to this low trophic level keystone species.Responses related to oxidative stress, protein damage and lipid peroxidation were investigated in C. finmarchicus exposed to a water-accommodated fraction (WAF) of a naphthenic North Atlantic crude oil. The exposure concentration corresponded to 50% of the 96 h LC50, and samples were obtained at 0, 24, 48, 72 and 96 h after exposure initiation. Gene expressions (superoxide dismutase, catalase, glutathione S-transferase, glutathione synthetase, heat shock protein 70 and 90, ubiquitin and cytochrome P-450 330A1), enzyme activities (superoxide dismutase, catalase, glutathione S-transferase) and concentrations of total glutathione and malondialdehyde were analyzed. Gene expression analyses showed no differences between controls and the exposed animals, however significantly higher glutathione S-transferase activity and malondialdehyde concentrations were found in the exposed group, suggests lipid peroxidation as main toxic effect.

Comparative toxicity of conventional and unconventional oils during rainbow trout (Oncorhynchus mykiss) embryonic development: From molecular to health consequences

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.

Multilevel responses of adult zebrafish to crude and chemically dispersed oil exposure

BackgroundThe application of chemical dispersants is a common remediation strategy when accidental oil spills occur in aquatic environments. Breaking down the oil slick into small droplets, dispersants facilitate the increase of particulate and dissolved oil compounds, enhancing the bioavailability of toxic oil constituents. The aim of the present work was to explore the effects of water accommodated fractions (WAF) of a naphthenic North Sea crude oil produced with and without the addition of the chemical dispersant FINASOL OSR 52 to adult zebrafish exposed for 3 and 21 d. Fish were exposed to environmentally relevant concentrations of 5% and 25% WAFOIL (1:200) and to 5% WAFOIL+D (dispersant–oil ratio 1:10) in a semi-static exposure setup.ResultsThe chemically dispersed WAF presented a 20-fold increase of target polycyclic aromatic hydrocarbons (PAHs) in the water phase compared to the corresponding treatment without dispersant and was the only treatment resulting in markedly bioaccumulation of PAHs in carcass after 21 d compared to the control. Furthermore, only 5% WAFOIL+D caused fish mortality. In general, the undispersed oil treatments did not lead to significant effects compared to control, while the dispersed oil induced significant alterations at gene transcription and enzyme activity levels. Significant up-regulation of biotransformation and oxidative stress response genes (cyp1a, gstp1, sod1 and gpx1a) was recorded in the livers. For the same group, a significant increment in EROD activity was detected in liver along with significant increased GST and CAT activities in gills. The addition of the chemical dispersant also reduced brain AChE activity and showed a potential genotoxic effect as indicated by the increased frequency of micronuclei in erythrocytes after 21 d of exposure.ConclusionsThe results demonstrate that the addition of chemical dispersants accentuates the effect of toxic compounds present in oil as it increases PAH bioavailability resulting in diverse alterations on different levels of biological organization in zebrafish. Furthermore, the study emphasizes the importance to combine multilevel endpoints for a reliable risk assessment due to high variable biomarker responses. The present results of dispersant impact on oil toxicity can support decision making for oil spill response strategies.

Marine Algal Sensitivity to Source and Weathered Oils.

After the Deepwater Horizon oil spill in 2010, toxicity tests were conducted using 4 microalgae (Dunaliella tertiolecta, Skeletonema costatum, Isochrysis galbana, and Thalassiosira pseudonana) and one macroalga (Ectocarpus siliculosus) to study potential impacts on phytoplankton and other primary producers in the Gulf of Mexico and characterize species sensitivity. Tests were performed with Corexit 9500 and fresh source oil and weathered oil samples collected from the field during the Deepwater Horizon oil spill. Because crude oils are mixtures of poorly water-soluble hydrocarbons, dosing was performed using water-accommodated fractions (WAFs) and chemically enhanced (CE) WAFs with the addition of dispersant at a 1:20 dispersant:oil ratio using standard toxicity testing protocols. Exposure media were analyzed for volatile organic compounds, parent and alkylated polycyclic aromatic hydrocarbons, and saturated hydrocarbon compounds. Toxicity was reported as no-observable effect concentration and median effect concentration (EC50) values for average specific growth rate based on nominal percent dilution of stock solution WAFs and sum of dissolved oil toxic units for WAF/CEWAF tests. The macroalga and green alga D. tertiolecta were largely unaffected by any WAF or CEWAFs tested. Isochrysis galbana was found to be the most sensitive species overall with significant growth rate inhibitions for dispersant and all the WAFs/CEWAFs tested. Physically dispersed source oils were generally more toxic than weathered oils. The protectiveness of the chronic toxic units was effective at identifying observed algal growth rate inhibitions across algal species and oil types despite the impact of dispersants. Environ Toxicol Chem 2021;40:2742-2754. © 2021 SETAC.

Cellular accumulation of crude oil compounds reduces the competitive fitness of the coral symbiont Symbiodinium glynnii

Oil spill events in the marine environment can have a deleterious impact on the affected ecosystems, such as coral reefs, with direct consequences for their socioeconomic value. The mutualistic relationship between tropical corals and their dinoflagellate symbionts (Symbiodiniaceae) provide structural and nutritional basis for a high local biodiversity in oligotrophic waters. Here, we investigated effects of crude oil water-accommodated fraction on the competitive fitness of the model zooxanthellae species Symbiodinium glynnii. Results of laboratory essays demonstrate that crude oil carbon is incorporated into the cellular biomass with a concomitant change of δ13C isotopic value. Carcinogenic/mutagenic polycyclic aromatic hydrocarbons were identified in the culture media and were responsible for a linear reduction in population growth of S. glynnii, presumably related to energy relocation for DNA repair. Additionally, the experiments revealed that physiological effects induced by crude oil compounds are genetically inherited by the following generations under non-contaminated growth conditions, and induce a reduction in the competitive fitness to cope with other environmental parameters, such as low salinity. We suggest that the effects of crude oil contamination represent an imparing factor for S. glynnii coping with anthropogenic drivers (e.g. warming and acidification) and interfere with the delicate symbiont-host relationship of tropical corals. This is especially relevant in the coastal areas of northeastern Brazil where an oil spill event deposited crude oil on shallow water sediments with the potential to be resuspended to the water column by physical and/or biological activity, enhancing the risk of future coral bleaching events.

Biodegradation of water-accommodated aromatic oil compounds in Arctic seawater at 0 °C

Oil spills in Arctic marine environments are expected to increase concurrently with the expansion of shipping routes and petroleum exploitation into previously inaccessible ice-dominated regions. Most research on oil biodegradation focusses on the bulk oil, but the fate of the water-accommodated fraction (WAF), mainly composed of toxic aromatic compounds, is largely underexplored. To evaluate the bacterial degradation capacity of such dissolved aromatics in Greenlandic seawater, microcosms consisting of 0 °C seawater polluted with WAF were investigated over a 3-month period. With a half-life (t1/2) of 26 days, m-xylene was the fastest degraded compound, as measured by gas chromatography - mass spectrometry. Substantial slower degradation was observed for ethylbenzene, naphthalenes, phenanthrene, acenaphthylene, acenaphthene and fluorenes with t1/2 of 40–105 days. Colwellia, identified by 16S rRNA gene sequencing, was the main potential degrader of m-xylene. This genus occupied up to 47 % of the bacterial community until day 10 in the microcosms. Cycloclasticus and Zhongshania aliphaticivorans, potentially utilizing one-to three-ringed aromatics, replaced Colwellia between day 10 and 96 and occupied up to 6 % and 23 % of the community, respectively. Although most of the WAF can ultimately be eliminated in microcosms, our results suggest that the restoration of an oil-impacted Arctic environment may be slow as most analysed compounds had t1/2 of over 2–3 months and the detrimental effects of a spill towards the marine ecosystem likely persist during this time.

Aquatic Toxicity Calculation of Mixtures: A Chemical Activity Approach Incorporating a Bioavailability Reduction Concept.

A calculation estimating the effect concentration (EL/LL50) of a water-accommodated fraction (WAF) for mixture toxicity is proposed. The method is based on chemical activity where the activity of a molecule is its effective concentration taking into account intermolecular interactions. First, the thermodynamic influence of each constituent on the solubility of the others within the mixture (i.e. the concentration of each constituent in the "loading rate") is determined. Then, the non-bioavailable fraction is determined and removed to calculate the true concentration of each constituent exerting toxicity. Finally, the loading rate is adjusted until the sum of activities of the bioavailable fractions is equal to the fraction-weighted average of toxic activity of each constituent. This process is a mechanistic interpretation of experimental WAF tests. The methodology has been validated comparing toxic loading rates of 13 reliable experimental WAF studies on fish, daphnids, and algae. The predictions were all within a factor of 2 of the study outcomes and can be considered as accurate as the laboratory studies. This is in contrast to the standard additivity method which consistently overestimates the toxicity of these mixtures by at least a factor of 2 up to over an order of magnitude or even more.

Lethal and sublethal effects of diluted bitumen and conventional oil on fathead minnow (Pimephales promelas) larvae exposed during their early development.

The increasing extraction of bitumen from the oil sands region in Canada is creating a need for transport. Spills from current and projected pipelines represent a significant environmental risk, especially for freshwater ecosystems. The toxicity of diluted bitumen (dilbit) on freshwater fish is largely unknown. This study assessed the toxicity of two dilbits (Clearwater McMurray and Bluesky) and compared their toxicity to a conventional oil (Lloydminster Heavy) on fathead minnow (Pimephales promelas) larvae. Larvae were exposed to various concentrations of the water-accommodated fraction (WAF) of the oils during 7 days from hatching. In the WAF treatments, the concentrations of volatile organic compounds (VOCs), including benzene, toluene, ethylbenzene, xylene (BTEX), hydrocarbons containing 6 to 10 carbon atoms (C6-10), and polycyclic aromatic hydrocarbons (PAHs) and their alkylated forms were measured. Both dilbits contained higher concentrations of light components, while the conventional oil contained the highest concentrations of PAHs and alkylated PAHs. The Clearwater McMurray dilbit induced a higher mortality, with a maximum of 65.3%, while the other oils induced a similar mortality up to 16.5% and 18.6% for Lloydminster and for Bluesky, respectively. All three oils induced an increase in gene expression of the phase I detoxification enzyme (cyp1a) with increasing total hydrocarbon concentrations. All three exposures induced a similar increase in glutathione S-transferase (GST) activity, but no change in gst gene expression. For the Bluesky and Lloydminster exposures, an increase in malondialdehyde concentration was also observed, suggesting a rate limiting capacity of GST and phase II enzymes to perform the biotransformation of the PAH metabolites. Overall, this study brings new insights on the toxicity of dilbits in comparison to conventional oils on early life stages of North American freshwater fish and demonstrated that dilbits can be more toxic than conventional oils, depending on their composition and diluent proportions.

A Comparison of Short-Term and Continuous Exposures in Toxicity Tests of Produced Waters, Condensate, and Crude Oil to Marine Invertebrates and Fish.

Petroleum hydrocarbons can be discharged into the marine environment during offshore oil and gas production or as a result of oil spills, with potential impacts on marine organisms. Ecotoxicological assay durations (typically 24–96 h) used to characterize risks to exposed organisms may not always reflect realistic environmental exposure durations in a high‐energy offshore environment where hydrocarbons are mixed and diluted rapidly in the water column. To investigate this, we adapted 3 sensitive toxicity tests to incorporate a short‐term pulse exposure to 3 petroleum‐based products: a produced water, the water‐accommodated fraction (WAF) of a condensate, and a crude oil WAF. We measured 48‐h mobility of the copepod Acartia sinjiensis, 72‐h larval development of the sea urchin Heliocidaris tuberculata, and 48‐h embryo survival and deformities of yellowtail kingfish Seriola lalandi, after exposure to a dilution series of each of the 3 products for 2, 4 to 12, and 24 h and for the standard duration of each toxicity test (continuous exposure). Effects on copepod survival and sea urchin larval development were significantly reduced in short‐term exposures to produced water and WAFs compared to continuous exposures. Fish embryos, however, showed an increased frequency of deformities at elevated concentrations regardless of exposure duration, although there was a trend toward increased severity of deformities with continuous exposure. The results demonstrate how exposure duration alters toxic response and how incorporating relevant exposure duration to contaminants into toxicity testing may aid interpretation of more realistic effects (and hence an additional line of evidence in risk assessment) in the receiving environment. Environ Toxicol Chem 2021;40:2587–2600. © 2021 CSIRO. Environmental Toxicology and Chemistry published by Wiley Periodicals LLC on behalf of SETAC.

Environmental modulators of diluted bitumen effects in juvenile pink salmon (Oncorhynchus gorbuscha)

Recent and potential expansions in the transportation of diluted bitumen (dilbit) through marine terminals in coastal regions of British Columbia require the examination of potential risks to estuarine species such as Pacific salmon. The estuarine habitat of out-migrated pink salmon (Oncorhynchus gorbuscha) exhibits dynamic temperature and salinity regimes, possibly modifying dilbit exposure, bioavailability and/or its effects. To examine dilbit toxicity and its modification by environmental stressors, juvenile pinks were subchronically exposed for 3 months to the water-accommodated fraction (WAF) of Cold Lake Blend dilbit (winter) in seawater at three salinities (7, 14, and 28‰ [temperature 12.5 °C]) and three temperatures (8.5, 12.5, and 16.5 °C [salinity of 28‰]). Temperature and salinity alone did not affect any measured endpoints in control fish. Dilbit exposure induced higher mortality at high (16.5 °C) and low temperatures (8.5 °C) as well as at higher salinity (28‰) in fish exposed to the highest dilution of WAF [total polycyclic aromatic compounds (TPAC) = 128.9 μg/L]. A concentration-dependent reduction of growth was evident in fish exposed to the medium (TPAC = 97.3 μg/L) and high dilution of WAF at higher temperatures (12.5 and 16.5 °C) and high salinity (28‰). At 28‰, swimming performance (Uburst) was decreased in fish exposed to the highest concentration of dilbit at all 3 temperatures. Gill Na+-K+-ATPase activity, white muscle lactate, glycogen, and triglyceride concentrations were altered by dilbit exposure and modified by temperature and salinity. In addition, gene expression associated with phase I biotransformation, energy metabolism, mitochondrial activity, and inflammation showed significant upregulation with exposure and temperature stress. Dilbit exposure at PAC concentrations in the ppb range, affected pink salmon at the molecular, biochemical, and whole organism level; effects that were exacerbated by environmental temperature and salinity.

Application of a biological multilevel response approach in the copepod Acartia tonsa for toxicity testing of three oil Water Accommodated Fractions

Copepods play a critical role in the marine food webs, being a food source for marine organisms. In this study, we investigated the toxic effects of Water Accommodated Fractions (WAFs) from three types of oil: Naphthenic North Sea crude oil (NNS), Intermediate Fuel Oil (IFO 180) and a commercial Marine Gas Oil (MGO). The WAFs were prepared at 10 °C and 30 PSU (practical salinity unit), and tested on the marine copepod Acartia tonsa at different endpoints and at different levels of biological organization. We determined the median lethal concentrations after 96 h (LC50) and reproduction capabilities were calculated in adult females following seven days of exposure to sublethal WAF doses. The total lipid content was measured in reproductive females using Nile red lipophilic dye after 96 h of WAF exposure. We also measured the transcription levels of genes involved in antioxidant response and xenobiotic biotransformation after short exposure for 48 h. High doses (7% WAF) of MGO affected survival, percentage of fecund females, egg hatching success, and total lipid content. The IFO 180 WAF affected, at medium (20%) and high (40%) doses, the number of fecund females, mortality and produced significant effects on gene expression levels. In conclusion, toxicity assays showed that the WAFs prepared from refined oils were more toxic than crude oil WAF to Acartia tonsa.

Differential Gene Expression Induced by Acute Exposure to Water Accommodated Fraction (WAF) and Chemically Enhanced WAF (CEWAF) of Light Crude Oil and Nokomis 3-F4 in Limulus polyphemus Larvae.

In 2018 we evaluated at 48h and 96h, the gene expression profile of larvae of Limulus polyphemus exposed to 10% and 100% of water-accommodated fraction (WAF) of light crude oil (API 35), and 10% and 100% of a chemically enhanced WAF (CEWAF) with the dispersant Nokomis 3-F4® in a static-acute (96h) bioassay. Alkanes and PAHs concentrations were higher in CEWAF than in WAF stock solutions. Under the proved conditions, the expression profile of genes associated to detoxification processes (glutathione S-transferase and glutathione peroxidase), stress (heat shock protein), innate immunity (tumor necrosis factor receptor-associated factor 4 traf4), cell death (apoptosis inhibitor 5) and DNA repairing (E3 ubiquitin protein ligase), showed a deregulation at 48h followed by an upregulation at 96h, with exception of glutathione peroxidase, heat shock protein and innate immunity that remained low in CEWAF. In conclusion, by using genes that have been proposed as biomarkers to pollutants exposure, L. polyphemus larvae showed an early activation of genes related to the immune system, antioxidant, heat shock and NER.