A review of the toxicology of oil in vertebrates: what we have learned following the Deepwater Horizon oil spill.

In the wake of the Deepwater Horizon (DWH) oil spill, a number of government agencies, academic institutions, consultants, and nonprofit organizations conducted lab- and field-based research to understand the toxic effects of the oil. Lab testing was performed with a variety of fish, birds, turtles, and vertebrate cell lines (as well as invertebrates); field biologists conducted observations on fish, birds, turtles, and marine mammals; and epidemiologists carried out observational studies in humans. Eight years after the spill, scientists and resource managers held a workshop to summarize the similarities and differences in the effects of DWH oil on vertebrate taxa and to identify remaining gaps in our understanding of oil toxicity in wildlife and humans, building upon the cross-taxonomic synthesis initiated during the Natural Resource Damage Assessment. Across the studies, consistency was found in the types of toxic response observed in the different organisms. Impairment of stress responses and adrenal gland function, cardiotoxicity, immune system dysfunction, disruption of blood cells and their function, effects on locomotion, and oxidative damage were observed across taxa. This consistency suggests conservation in the mechanisms of action and disease pathogenesis. From a toxicological perspective, a logical progression of impacts was noted: from molecular and cellular effects that manifest as organ dysfunction, to systemic effects that compromise fitness, growth, reproductive potential, and survival. From a clinical perspective, adverse health effects from DWH oil spill exposure formed a suite of signs/symptomatic responses that at the highest doses/concentrations resulted in multi-organ system failure.

Sublethal effects of oil-contaminated sediment to early life stages of the Eastern oyster, Crassostrea virginica.

The explosion of the Deepwater Horizon (DWH) oil drilling rig resulted in the release of crude oil into the Gulf of Mexico. This event coincided with the spawning season of the Eastern oyster, Crassostrea virginica. Although oil bound to sediments constitutes an important source of polycyclic aromatic hydrocarbon (PAH) exposure to benthic organisms, toxicity of sediment-associated DWH oil has not been investigated in any bivalve species. Here, we evaluated the sublethal effects of acute exposure of gametes, embryos and veliger larvae of the Eastern oyster to different concentrations of unfiltered elutriates of sediment contaminated with DWH oil. Our results suggest that gametes, embryos and veliger larvae are harmed by exposure to unfiltered elutriates of contaminated sediment. Effective concentrations for fertilization inhibition were 40.6 µg tPAH50 L-1 and 173.2 µg tPAH50 L-1 for EC201h and EC501h values, respectively. Embryo exposure resulted in dose-dependent abnormalities (EC20 and EC50 values were 77.7 µg tPAH50 L-1 and 151 µg tPAH50 L-1, respectively) and reduction in shell growth (EC2024h value of 1180 µg tPAH50 L-1). Development and growth of veliger larvae were less sensitive to sediment-associated PAHs compared to embryos. Fertilization success and abnormality of larvae exposed as embryos were the most sensitive endpoints for assessing the toxicity of oil-contaminated sediment. Bulk of measured polycyclic aromatic hydrocarbons were sediment-bound and caused toxic effects at lower tPAH50 concentrations than high energy water accommodated fractions (HEWAF) preparations from the same DWH oil. This study suggests risk assessments would benefit from further study of suspended contaminated sediment.

Crude oil cardiotoxicity to red drum embryos is independent of oil dispersion energy.

The potential bioavailability of toxic chemicals from oil spills to water column organisms such as fish embryos may be influenced by physical dispersion along an energy gradient. For example, a surface slick with minimal wave action (low energy) could potentially produce different toxic effects from high energy situations such as pressurized discharge from a blown wellhead. Here we directly compared the toxicity of water accommodated fractions (WAFs) of oil prepared with low and high mixing energy (LEWAFs and HEWAFs, respectively) using surface oil samples collected during the 2010 Deepwater Horizon spill, and embryos of a representative nearshore species, red drum (Sciaenops ocellatus). Biological effects of each WAF type was quantified with several functional and morphological indices of developmental cardiotoxicity, providing additional insight into species-specific responses to oil exposure. Although the two WAF preparations yielded different profiles of polycyclic aromatic hydrocarbons (PAHs), cardiotoxic phenotypes were essentially identical. Based on benchmark thresholds for both morphological and functional cardiotoxicity, in general LEWAFs had lower thresholds for these phenotypes than HEWAFs based on total PAH measures. However, HEWAF and LEWAF toxicity thresholds were more similar when calculated based on estimates of dissolved PAHs only. Differences in thresholds were attributable to the weathering state of the oil samples.

Combined effects of hypoxia or elevated temperature and Deepwater Horizon crude oil exposure on juvenile mahi-mahi swimming performance.

This study examined potential interactive effects of co-exposure to Deepwater Horizon (DWH) crude oil (∼30 µg L-1 ΣPAHs) for 24 h and either hypoxia (2.5 mg O2 L-1; 40% O2 saturation) or elevated temperature (30 °C) on the swimming performance of juvenile mahi-mahi (Coryphaena hippurus). Additionally, effects of shorter duration exposures to equal or higher doses of oil alone either prior to swimming or during the actual swim trial itself were examined. Only exposure to hypoxia alone or combined with crude oil elicited significant decreases in critical swimming speed (Ucrit) and to a similar extent (∼20%). In contrast, results indicate that elevated temperature might ameliorate some effects of oil exposure on swimming performance and that effects of shorter duration exposures are either reduced or delayed.

Estimating incident ultraviolet radiation exposure in the northern Gulf of Mexico during the Deepwater Horizon oil spill.

Millions of barrels of oil were released into the Gulf of Mexico following the 2010 explosion of the Deepwater Horizon oil rig. Polycyclic aromatic hydrocarbons (PAHs) are toxic components of crude oil, which may become more toxic in the presence of ultraviolet (UV) radiation, a phenomenon known as photo-induced toxicity. The Deepwater Horizon spill impacted offshore and estuarine sites, where biota may be co-exposed to UV and PAHs. Penetration of UV into the water column is affected by site-specific factors. Therefore, measurements and/or estimations of UV are necessary when one is assessing the risk to biota posed by photo-induced toxicity. We describe how estimates of incident UV were determined for the area impacted by the Deepwater Horizon oil spill, using monitoring data from radiometers near the spill, in conjunction with reference spectra characterizing the composition of solar radiation. Furthermore, we provide UV attenuation coefficients for both near- and offshore sites in the Gulf of Mexico. These estimates are specific to the time and location of the spill, and fall within the range of intensities utilized during photo-induced toxicity tests performed in support of the Deepwater Horizon Natural Resource Damage Assessment (NRDA). These data further validate the methodologies and findings of phototoxicity tests included in the Deepwater Horizon NRDA, while underscoring the importance of considering UV exposure when assessing possible risks following oil spills. Environ Toxicol Chem 2018;37:1679-1687. © 2018 SETAC.

Photo-induced toxicity in early life stage fiddler crab (Uca longisignalis) following exposure to Deepwater Horizon oil.

The 2010 explosion of the Deepwater Horizon (DWH) oil rig led to the release of millions of barrels of oil in the Gulf of Mexico. Oil in aquatic ecosystems exerts toxicity through multiple mechanisms, including photo-induced toxicity following co-exposure with UV radiation. The timing and location of the spill coincided with both fiddler crab reproduction and peak yearly UV intensities, putting early life stage fiddler crabs at risk of injury due to photo-induced toxicity. The present study assessed sensitivity of fiddler crab larvae to photo-induced toxicity during co-exposure to a range of environmentally relevant dilutions of high-energy water accommodated fractions of DWH oil, and either <10, 50, or 100% ambient sunlight, achieved with filters that allowed for variable UV penetration. Solar exposures (duration: 7-h per day) were conducted for two consecutive days, with a dark recovery period (duration: 17-h) in between. Survival was significantly decreased in treatments the presence of >10% UV and relatively low concentrations of oil. Results of the present study indicate fiddler crab larvae are sensitive to photo-induced toxicity in the presence of DWH oil. These results are of concern, as fiddler crabs play an important role as ecosystem engineers, modulating sediment biogeochemical processes via burrowing action. Furthermore, they occupy an important place in the food web in the Gulf of Mexico.

Photo-induced toxicity following exposure to crude oil and ultraviolet radiation in 2 Australian fishes.

Some polycyclic aromatic hydrocarbons (PAHs), components of crude oil, are known to cause increased toxicity when organisms are co-exposed with ultraviolet radiation, resulting in photo-induced toxicity. The photodynamic characteristics of some PAHs are of particular concern to places like Australia with high ultraviolet radiation levels. The objective of the present study was to characterize the photo-induced toxicity of an Australian North West Shelf oil to early life stage yellowtail kingfish (Seriola lalandi) and black bream (Acanthopagrus butcheri). The fish were exposed to high-energy water accommodated fractions for 24 to 36 h. During the exposure, the fish were either co-exposed to full-intensity or filtered natural sunlight and then transferred to clean water. At 48 h, survival, cardiac effects, and spinal deformities were assessed. Yellowtail kingfish embryos co-exposed to oil and full-spectrum sunlight exhibited decreased hatching success and a higher incidence of cardiac arrhythmias, compared with filtered sunlight. A significant increase in the incidence of pericardial edema occurred in black bream embryos co-exposed to full-spectrum sunlight. These results highlight the need for more studies investigating the effects of PAHs and photo-induced toxicity under environmental conditions relevant to Australia. Environ Toxicol Chem 2018;37:1359-1366. © 2018 SETAC.

Exposure to Deepwater Horizon oil and Corexit 9500 at low concentrations induces transcriptional changes and alters immune transcriptional pathways in sheepshead minnows.

The 2010 Deepwater Horizon (DWH) oil spill caused the release of 4.9 million barrels of crude oil into the Gulf of Mexico, followed by the application of 2.9 million L of the dispersant, Corexit™ to mitigate the spread of oil. The spill resulted in substantial shoreline oiling, potentially exposing coastal organisms to polyaromatic hydrocarbon (PAH) and dispersant contaminants. To investigate molecular effects in fish following exposure to environmentally relevant concentrations of DWH oil and dispersants, we exposed adult sheepshead minnows (Cyprinodon variegatus) to two concentrations of high-energy water-accommodated fraction (HEWAF), chemically enhanced water-accommodated fraction (CEWAF) or Corexit 9500™ for 7 and 14days. Resulting changes in hepatic gene expression were measured using 8×15K microarrays. Analytical chemistry confirmed PAH concentrations in HEWAF and CEWAF treatments were low (ranging from 0.26 to 5.98µg/L), and likely representative of post-spill environmental concentrations. We observed significant changes to gene expression in all treatments (relative to controls), with Corexit and CEWAF having a greater effect on expression patterns in the liver than HEWAF treatments. Sub-network enrichment analysis of biological pathways revealed that the greatest number of altered pathways in high dose HEWAF and CEWAF treatments occurred following a 7-day exposure. Pathways related to immunity comprised the majority of pathways affected in each treatment, followed by pathways related to blood and circulation processes. Our results indicate that oil composition, concentration, and exposure duration all affect molecular responses in exposed fish, and suggest that low-concentration exposures may result in sub-lethal adverse effects.

Characterization of dissolved and particulate phases of water accommodated fractions used to conduct aquatic toxicity testing in support of the Deepwater Horizon natural resource damage assessment.

In response to the Deepwater Horizon oil spill, the Natural Resource Trustees implemented a toxicity testing program that included 4 different Deepwater Horizon oils that ranged from fresh to weathered, and 3 different oil-in-water preparation methods (including one that used the chemical dispersant Corexit 9500) to prepare a total of 12 chemically unique water accommodated fractions (WAFs). We determined how the different WAF preparation methods, WAF concentrations, and oil types influenced the chemical composition and concentration of polycyclic aromatic hydrocarbons (PAHs) in the dissolved and particulate phases over time periods used in standard toxicity tests. In WAFs prepared with the same starting oil and oil-to-water ratio, the composition and concentration of the dissolved fractions were similar across all preparation methods. However, these similarities diverged when dilutions of the 3 WAF methods were compared. In WAFs containing oil droplets, we found that the dissolved phase was a small fraction of the total PAH concentration for the high-concentration stock WAFs; however, the dissolved phase became the dominant fraction when it was diluted to lower concentrations. Furthermore, decreases in concentration over time were mainly related to surfacing of the larger oil droplets. The initial mean diameters of the droplets were approximately 5 to 10 µm, with a few droplets larger than 30 µm. After 96 h, the mean droplet size decreased to 3 to 5 µm, with generally all droplets larger than 10 µm resurfacing. These data provide a detailed assessment of the concentration and form (dissolved vs particulate) of the PAHs in our WAF exposures, measurements that are important for determining the effects of oil on aquatic species. Environ Toxicol Chem 2017;36:1460-1472. © 2017 SETAC.

Assessment of early life stage mahi-mahi windows of sensitivity during acute exposures to Deepwater Horizon crude oil.

Windows of exposure to a weathered Deepwater Horizon oil sample (slick A) were examined for early life stage mahi-mahi (Coryphaena hippurus) to determine whether there are developmental periods of enhanced sensitivity during the course of a standard 96-h bioassay. Survival was assessed at 96 h following oil exposures ranging from 2 h to 96 h and targeting 3 general periods of development, namely the prehatch phase, the period surrounding hatch, and the posthatch phase. In addition, 3 different oil preparations were used: high- and low-energy water accommodated fractions of oil and very thin surface slicks of oil (∼1 µm). The latter 2 were used to distinguish between effects due to direct contact with the slick itself and the water underlying the slick. Considering the data from all 3 exposure regimes, it was determined that the period near or including hatch was likely the most sensitive. Furthermore, toxicity was not enhanced by direct contact with slick oil. These findings are environmentally relevant given that the concentrations of polycyclic aromatic hydrocarbons eliciting mortality from exposures during the sensitive periods of development were below or near concentrations measured during the active spill phase. Environ Toxicol Chem 2017;36:1887-1895. © 2016 SETAC.

Responses of juvenile southern flounder exposed to Deepwater Horizon oil-contaminated sediments.

The Deepwater Horizon oil spill released millions of barrels of crude oil into the northern Gulf of Mexico, much of which remains associated with sediments and can have continuing impacts on biota. Juvenile southern flounder (Paralichthys lethostigma) were exposed for 28 d in the laboratory under controlled conditions to reference and Deepwater Horizon oil-contaminated sediments collected from coastal Louisiana to assess the impacts on an ecologically and commercially important benthic fish. The measured polycyclic aromatic hydrocarbon (PAH) concentrations in the sediments ranged from 0.25 mg/kg to 3940 mg/kg suite of 50 PAH analytes (tPAH50). Mortality increased with both concentration and duration of exposure. Exposed flounder length and weight was lower compared to controls after 28 d of exposure to the sediments with the highest PAH concentration, but condition factor was significantly higher in these fish compared with all other treatments. Histopathological analyses showed increased occurrence of gill abnormalities, including telangiectasis, epithelial proliferation, and fused lamellae in flounder exposed to sediments with the highest tPAH50 concentrations. In addition, hepatic vascular congestion and macrovesicular vacuolation were observed in flounder exposed to the more contaminated sediments. These data suggest that chronic exposure to field collected oil-contaminated sediments results in a variety of sublethal impacts to a benthic fish, with implications for long-term recovery from oil spills. Environ Toxicol Chem 2017;36:1067-1076. © 2016 SETAC.

Characterization of oil and water accommodated fractions used to conduct aquatic toxicity testing in support of the Deepwater Horizon oil spill natural resource damage assessment.

The Deepwater Horizon blowout resulted in the release of millions of barrels of crude oil. As part of the Trustees' Natural Resource Damage Assessment, a testing program was implemented to evaluate the toxicity of Deepwater Horizon oil and oil/dispersant mixtures to aquatic organisms from the Gulf of Mexico. Because of the variety of exposures that likely occurred, the program included 4 Deepwater Horizon oils, which encompassed a range of weathering states, and 3 different oil-in-water mixing methods, for a total of 12 unique water accommodated fractions (WAFs). The present study reports on the chemical characteristics of these 4 Deepwater Horizon oils and 12 WAFs. In addition, to better understand exposure chemistry, an examination was conducted of the effects of WAF preparation parameters-including mixing energy, starting oil composition, and oil-to-water mixing ratios-on the chemical profiles and final concentrations of these 12 WAFs. The results showed that the more weathered the starting oil, the lower the concentrations of the oil constituents in the WAF, with a shift in composition to the less soluble compounds. In addition, higher mixing energies increased the presence of insoluble oil constituents. Finally, at low to mid oil-to-water mixing ratios, the concentration and composition of the WAFs changed with changing mixing ratios; this change was not observed at higher mixing ratios (i.e., >1 g oil/L). Ultimately, the present study provides a basic characterization of the oils and WAFs used in the testing program, which helps to support interpretation of the more than 500 Deepwater Horizon Natural Resource Damage Assessment toxicity testing results and to enable a comparison of these results with different tests and with the field. Environ Toxicol Chem 2017;36:1450-1459. © 2016 SETAC.

Effects of sediment amended with Deepwater Horizon incident slick oil on the infaunal amphipod Leptocheirus plumulosus.

Crude oil released from the Deepwater Horizon disaster into the Gulf of Mexico posed potential impacts to infaunal invertebrates inhabiting near shore habitats. The effects of sediment-associated weathered slick oil on the amphipod Leptocheirus plumulosus was assessed using 28-d exposures to total PAH sediment concentrations ranging from 0.3 to 24mg/kg (sum of 50 PAHs or tPAH50). Survival and growth rate were significantly decreased in the 2.6, 11.4 and 24.2mg/kg treatments, but only growth in 5.5mg/kg. Offspring production was dramatically decreased but was variable and significantly different only for 24.2mg/kg. The concentrations associated with 20% decreases relative to reference were 1.05 (95% CI=0-2.89) mg/kg tPAH50 for growth rate and 0.632 (95% CI=0.11-2.15) mg/kg tPAH50 for offspring production. The concentrations of PAHs affecting amphipods are within the range of concentrations measured in marsh areas reportedly impacted by DWH oil after its release.

The effects of weathering and chemical dispersion on Deepwater Horizon crude oil toxicity to mahi-mahi (Coryphaena hippurus) early life stages.

To better understand the impact of the Deepwater Horizon (DWH) incident on commercially and ecologically important pelagic fish species, a mahi-mahi spawning program was developed to assess the effect of embryonic exposure to DWH crude oil with particular emphasis on the effects of weathering and dispersant on the magnitude of toxicity. Acute lethality (96 h LC50) ranged from 45.8 (28.4-63.1) µg l(-1) ΣPAH for wellhead (source) oil to 8.8 (7.4-10.3) µg l(-1) ΣPAH for samples collected from the surface slick, reinforcing previous work that weathered oil is more toxic on a ΣPAH basis. Differences in toxicity appear related to the amount of dissolved 3 ringed PAHs. The dispersant Corexit 9500 did not influence acute lethality of oil preparations. Embryonic oil exposure resulted in cardiotoxicity after 48 h, as evident from pericardial edema and reduced atrial contractility. Whereas pericardial edema appeared to correlate well with acute lethality at 96 h, atrial contractility did not. However, sub-lethal cardiotoxicity may impact long-term performance and survival. Dispersant did not affect the occurrence of pericardial edema; however, there was an apparent reduction in atrial contractility at 48 h of exposure. Pericardial edema at 48 h and lethality at 96 h were equally sensitive endpoints in mahi-mahi.

Enhanced biodegradation of hydrocarbon-contaminated sediments using microbial fuel cells.

A sediment microbial fuel cell (MFC) was tested to determine if electron transfer from the anaerobic zone of contaminated sediments to the overlying aerobic water could facilitate an enhanced and aerobic equivalent degradation of total petroleum hydrocarbons (TPH). Results indicate that voltages as high as 190 mV (2162 mW/m(3)) were achieved in a sediment MFC with an anode buried in sediments containing TPH concentrations at approximately 16,000 mg kg(-1). Additionally, after approximately 66 days, the TPH degradation rates were 2% and 24% in the open-circuit control sediment MFC and active sediment MFC, respectively. Therefore, it appears that applying MFC technology to contaminated sediments enhances natural biodegradation by nearly 12 fold. Additionally, a novel sediment MFC was designed to provide a cost-effective method of passive oxidation or indirect aerobic degradation of contaminants in an otherwise anaerobic environment. In addition, the use of a wicking air cathode in this study maintained dissolved oxygen concentrations 1-2 mg l(-1) higher than submerged cathodes, demonstrating that this technology can be applied to environments with either aerobic or anaerobic overlying water and an anaerobic matrix, such as shallow lagoon, ponds, and marshes, and groundwater.

Feasibility of using microbial fuel cell technology for bioremediation of hydrocarbons in groundwater.

A single-cell microbial fuel cell (MFC) design was used to study anaerobic microbes that utilize petroleum contaminants as a sole substrate to produce power during remediation. Additionally, we tested various proton bridge designs to physically separate the anode and cathode chambers of a two-cell MFC by approximately 9 m (approximately 30 ft.). This separation enables the potential use of MFC technology for in situ bioremediation of petroleum hydrocarbons in the groundwater, in which oxygen is usually depleted and oxygen availability only exists at or near the surface. Sustained power generation (as high as 120 mW/m(2) cathode) was recorded for approximately 6 d in a single-cell MFC utilizing a mixture of refinery waste (containing various concentrations of hydrocarbon contaminants) and cell growth media. MFC cell potential (1KOmega external resistance) decreased by approximately 55% over the length of the 9 m proton bridge with a 6.9% decrease in potential per m of bridge. This preliminary data indicates that using MFC technology (with our modifications) may enhance bioremediation of petroleum contaminants in groundwater under anaerobic conditions. Because oxygen is eventually used as the terminal electron acceptor for anaerobic biodegradation inside an MFC, this technology may be a cost-effective innovation to enhanced biodegradation in groundwater, by substituting or eliminating conventional in situ aeration. To our knowledge, this is the first report on power generation from MFCs utilizing mixed hydrocarbon substrates. In addition, this study is the first to show the applicability of using extended proton bridges for the physical separation of anode and cathode chambers over extended distances that may be encountered in the field.