Most of the escaped farmed salmon entering a river during a 5-year period were infected with one or more viruses.

Disease interactions between farmed and wild populations have been poorly documented for most aquaculture species, in part due to the complexities to study this. Here, we tested 567 farmed Atlantic salmon escapees, captured in a Norwegian river during 2014-2018, for five viral infections that are prevalent in global salmonid aquaculture. Over 90% of the escapees were infected with one or more viruses. Overall prevalences were: 75.7% for piscine orthoreovirus (PRV-1), 43.6% for salmonid alphavirus (SAV), 31.2% for piscine myocarditis virus (PMCV), 1.2% for infectious pancreatic necrosis virus (IPNV) and 0.4% for salmon anaemia virus (ISAV). A significantly higher prevalence of PMCV infection was observed in immature compared to mature individuals. The prevalence of both SAV and PMCV infections was higher in fish determined by fatty acid profiling to be 'recent' as opposed to 'early' escapees that had been in the wild for a longer period of time. This is the first study to establish a time-series of viral infection status of escapees entering a river with a native salmon population. Our results demonstrate that farmed escapees represent a continuous source of infectious agents which could potentially be transmitted to wild fish populations.

Oil droplet fouling on lesser sandeel (Ammodytes marinus) eggshells does not enhance the crude oil induced developmental toxicity.

The oil industry's expansion and increased operational activity at older installations, along with their demolition, contribute to rising cumulative pollution and a heightened risk of accidental oil spills. The lesser sandeel (Ammodytes marinus) is a keystone prey species in the North Sea and coastal systems. Their eggs adhere to the seabed substrate making them particularly vulnerable to oil exposure during embryonic development. We evaluated the sensitivity of sandeel embryos to crude oil in a laboratory by exposing them to dispersed oil at concentrations of 0, 15, 50, and 150 µg/L oil between 2 and 16 days post-fertilization. We assessed water and tissue concentrations of THC and tPAH, cyp1a expression, lipid distribution in the eyes, head and trunk, and morphological and functional deformities. Oil droplets accumulated on the eggshell in all oil treatment groups, to which the embryo responded by a dose-dependent rise in cyp1a expression. The oil exposure led to only minor sublethal deformities in the upper jaw and otic vesicle. The findings suggest that lesser sandeel embryos are resilient to crude oil exposure. The lowest observed effect level documented in this study was 36 µg THC/L and 3 µg tPAH/L. The inclusion of these species-specific data in risk assessment models will enhance the precision of risk evaluations for the North Atlantic ecosystems.

Discovery of polycyclic aromatic acid metabolites in fish exposed to the petroleum compounds 1-methylphenanthrene and 1,4-dimethylphenanthrene.

Most of the polycyclic aromatic hydrocarbons (PAHs) in petroleum are alkylated (alkyl PAHs), still the metabolism of these alkyl PAHs to the expected acid products (polycyclic aromatic acids; PAAs) has yet to be demonstrated in oil-exposed fish. Should these compounds be discovered in fish as they have in ragworm, rodents, and humans, they could present an indicative biomarker for assessing oil pollution. In this study, the ability to biotransform alkyl PAHs to PAAs was examined on Atlantic haddock (Melanogrammus aeglefinus). Exposure to phenanthrene, 1-methyphenanthrene or 1,4-dimethylphenanthrene was performed via intraperitoneal injection. An Ion Mobility Quadrupole Time-Of-Flight Mass Spectrometer (IMS-Q-TOF MS) was used in exploratory analysis of extracted bile samples. Acquisition of four-dimensional information by coupling liquid chromatography with the IMS-Q-TOF MS and in-silico prediction for feature prioritization in the data processing workflow allowed several tentative identifications with high degree of confidence. This work presents the first detection of PAAs in fish and suggests the importance of investigating alkyl PAHs in ecotoxicological studies of oil-polluted fish environments.

Mind the gap - Relevant design for laboratory oil exposure of fish as informed by a numerical impact assessment model.

Laboratory experiments provide knowledge of species-specific effects thresholds that are used to parameterize impact assessment models of oil contamination on marine ecosystems. Such experiments typically place individuals of species and life stages in tanks with different contaminant concentrations. Exposure concentrations are usually fixed, and the individuals experience a shock treatment being moved from clean water directly into contaminated water and then back to clean water. In this study, we use a coupled numerical model that simulates ocean currents and state, oil dispersal and fate, and early life stages of fish to quantify oil exposure histories, specifically addressing oil spill scenarios of high rates and long durations. By including uptake modelling we also investigate the potential of buffering transient high peaks in exposure. Our simulation results are the basis for a recommendation on the design of laboratory experiments to improve impact assessment model development and parameterization. We recommend an exposure profile with three main phases: i) a gradual increase in concentration, ii) a transient peak that is well above the subsequent level, and iii) a plateau of fixed concentration lasting ∼3 days. In addition, a fourth phase with a slow decrease may be added.

Environmentally realistic concentrations of chlorinated, brominated, and fluorinated persistent organic pollutants induce the unfolded protein response as a shared stress pathway in the liver of Atlantic cod (Gadus morhua).

In the North Sea and North Atlantic coastal areas, fish experience relatively high background levels of persistent organic pollutants. This study aimed to compare the mode of action of environmentally relevant concentrations of mixtures of halogenated compounds in Atlantic cod. Juvenile male cod with mean weight of 840 g were exposed by gavage to dietary mixtures of chlorinated (PCBs, DDT analogs, chlordane, lindane, and toxaphene), brominated (PBDEs), and fluorinated (PFOS) compounds for 4 weeks. One group received a combined mixture of all three compound groups. The results showed that the accumulated levels of chemicals in cod liver after 4 weeks of exposure reflected concentrations found in wild fish in this region. Pathway analysis revealed that the treatment effects by each of the three groups of chemicals (chlorinated, brominated, and fluorinated) converged on activation of the unfolded protein response (UPR). Upstream regulator analysis predicted that almost all the key transcription factors (XBP1, ERN1, ATF4, EIF2AK3, and NFE2L2) regulating the UPR were significantly activated. No additive effect was observed in cod co-treated with all three compound groups. In conclusion, the genome-wide transcriptomic study suggests that the UPR pathway is a sensitive common target of halogenated organic environmental pollutants in fish.

Co-Exposure of Phenanthrene and the cyp-Inducer 3-Methylchrysene Leads to Altered Biotransformation and Increased Toxicity in Fish Egg and Larvae.

Polycyclic aromatic hydrocarbons (PAHs) have frequently been suspected of governing crude oil toxicity because of similar morphological defects in fish. However, PAH concentrations are often not high enough to explain the observed crude oil toxicity. We hypothesize that one PAH can enhance the metabolism and toxicity of another PAH when administered as a mixture. Early life stage Atlantic haddock (Melanogrammus aeglefinus) were in this study exposed to phenanthrene in the presence and absence of 3-methylchrysene that is known to induce the metabolic enzyme cytochrome P450 1A via cyp1a gene expression. Uptake, metabolism, and multiple toxicity endpoints were then measured in a time-course study up to 3 days post-hatching. Passive dosing provided aqueous concentrations ≈180 µg/L for phenanthrene and ≈0.6 µg/L for 3-methylchrysene, which resulted in tissue concentrations ≈60 µg/g ww for phenanthrene and ≈0.15 µg/g ww for 3-methylchrysene. The low concentration of 3-methylchrysene led to the elevated expression of cyp1a but no toxicity. Levels of phenanthrene metabolites were 5-fold higher, and morphological defects and cardiotoxicity were consistently greater when co-exposed to both compounds relative to phenanthrene alone. This work highlights the metabolic activation of PAH toxicity by a co-occurring PAH, which can lead to excess toxicity, synergistic effects, and the overproportional contribution of PAHs to crude oil toxicity.

Cardiac toxicity of phenanthrene depends on developmental stage in Atlantic cod (Gadus morhua).

Complex mixtures like crude oil, and single components such as Phenanthrene (Phe), induce cardiotoxicity by interfering with excitation-contraction coupling. However, recent work has demonstrated that the timing of pollutant exposure during embryogenesis greatly impacts the degree of cardiac dysfunction caused. Here, we aimed to clarify the temporal dependence of Phe toxicity and the downstream effects of cardiac dysfunction using Atlantic cod (Gadus morhua). Phe (nominal concentration, 1.12 µmol/L), or the L-type­calcium channel blocker Nicardipine (Nic) (nominal concentration, 2 and 4 µmol/L), were individually applied to cod embryos either during cardiogenesis (early) or after the onset of cardiac function (late). Phe toxicity was highly dependent on the timing of exposure. Exposure after the onset of cardiac function (i.e. late) caused more severe cardiac and extracardiac abnormalities at 3 days post hatching (dph) than early exposure. Late Phe exposure resulted in a smaller ventricle, eliminated ventricular contraction, and reduced atrial contraction. In contrast, early Phe exposure did not have an effect on cardiac development and function. This temporal difference was not as evident in the Nic treatment. Early Nic exposure created similar morphological phenotypes to the late Phe exposure. The two treatments (early Nic and late Phe) also shared a cardiofunctional phenotype, comprised of eliminated ventricular, and reduced atrial, contraction. These data suggest that extracardiac abnormalities, such as the craniofacial deformities seen after late embryonic exposure to cardiotoxic oil components and mixtures, are mostly downstream effects of cardiac dysfunction.

Crude oil exposure of early life stages of Atlantic haddock suggests threshold levels for developmental toxicity as low as 0.1 µg total polyaromatic hydrocarbon (TPAH)/L.

Atlantic haddock (Melanogrammus aeglefinus) embryos bind dispersed crude oil droplets to the eggshell and are consequently highly susceptible to toxicity from spilled oil. We established thresholds for developmental toxicity and identified any potential long-term or latent adverse effects that could impair the growth and survival of individuals. Embryos were exposed to oil for eight days (10, 80 and 300 µg oil/L, equivalent to 0.1, 0.8 and 3.0 µg TPAH/L). Acute and delayed mortality were observed at embryonic, larval, and juvenile stages with IC50 = 2.2, 0.39, and 0.27 µg TPAH/L, respectively. Exposure to 0.1 µg TPAH/L had no negative effect on growth or survival. However, yolk sac larvae showed significant reduction in the outgrowth (ballooning) of the cardiac ventricle in the absence of other extracardiac morphological defects. Due to this propensity for latent sublethal developmental toxicity, we recommend an effect threshold of 0.1 µg TPAH/L for risk assessment models.

Alkyl-phenanthrenes in early life stage fish: differential toxicity in Atlantic haddock (Melanogrammus aeglefinus) embryos.

Tricyclic polycyclic aromatic hydrocarbons (PAHs) are believed to be the primary toxic components of crude oil. Such compounds including phenanthrene are known to have direct effects on cardiac tissue, which lead to malformations during organogenesis in early life stage fish. We tested a suite of 13 alkyl-phenanthrenes to compare uptake and developmental toxicity in early life stage haddock (Melanogrammus aeglefinus) embryos during gastrulation/organogenesis beginning at 2 days post fertilization via passive dosing. The alkyl-phenanthrenes were tested at their solubility limits, and three of them also at lower concentrations. Measured body burdens were linearly related to measured water concentrations. All compounds elicited one or more significant morphological defects or functional impairment, such as decreased length, smaller eye area, shorter jaw length, and increased incidence of body axis deformities and eye deformities. The profile of developmental toxicities appeared unrelated to the position of alkyl substitution, and gene expression of cytochrome 1 a (cyp1a) was low regardless of alkylation. Mortality and sublethal effects were observed below the expected range for baseline toxicity, thus indicating excess toxicity. Additionally, PAH concentrations that resulted in toxic effects here were far greater than when measured in whole crude oil exposures that cause toxicity. This work demonstrates that, while these phenanthrenes are toxic to early life stage fish, they cannot individually account for most of the developmental toxicity of crude oil, and that other compounds and/or mixture effects should be given more consideration.

Co-exposure to UV radiation and crude oil increases acute embryotoxicity and sublethal malformations in the early life stages of Atlantic haddock (Melanogrammus aeglefinus).

Crude oil causes severe abnormalities in developing fish. Photomodification of constituents in crude oil increases its toxicity several fold. We report on the effect of crude oil, in combination with ultraviolet (UV) radiation, on Atlantic haddock (Melanogrammus aeglefinus) embryos. Accumulation of crude oil on the eggshell makes haddock embryos particularly susceptible to exposure. At high latitudes, they can be exposed to UV radiation many hours a day. Haddock embryos were exposed to crude oil (5-300 µg oil/L nominal loading concentrations) for three days in the presence and absence of UV radiation (290-400 nm). UV radiation partly degraded the eggs' outer membrane resulting in less accumulation of oil droplets in the treatment with highest oil concentration (300 µg oil/L). The co-exposure treatments resulted in acute toxicity, manifested by massive tissue necrosis and subsequent mortality, reducing LC50 at hatching stage by 60 % to 0.24 µg totPAH/L compared to 0.62 µg totPAH/L in crude oil only. In the treatment with nominal low oil concentrations (5-30 µg oil/L), only co-exposure to UV led to sublethal morphological heart defects. Including phototoxicity as a parameter in risk assessments of accidental oil spills is recommended.

Monitoring ocean water quality by deployment of lumpfish (Cyclopterus lumpus) eggs: In situ bioaccumulation and toxicity in embryos.

Fish embryos can bioaccumulate and are particularly sensitive to a wide range of contaminants, which makes them suitable sentinels for environmental biomonitoring. However, fish embryos are very rarely utilized in environmental monitoring surveys, possibly due to their fragility and seasonality. In the present work, we assessed the applicability of caged lumpfish (Cyclopterus lumpus) eggs for in situ biomonitoring of exposure and effects of organic contaminants focusing on polyaromatic hydrocarbons and phenolic compounds. Fertilized eggs (1 dpf) were transplanted for 17-19 days at different locations that differed in terms of contaminant load, depths and weather conditions, namely at three stations close to the city of Trondheim (two harbour areas and a one in the Fjord) and three stations at a coastal aquaculture facility. High survival upon retrieval after deployment showed that lumpfish eggs are relatively robust and survive encaging in different environments. Bioaccumulation of organic contaminants (PAHs and phenolic compounds) was measured and potential effects on hatching, development, survival and larvae morphometry were determined. Chemical analyses showed that especially PAHs were effectively accumulated in eggs in contaminated sites, with concentrations of Æ©PAHs being 15 - 25 times higher in harbour areas compared to those at the aquaculture facility. A higher incidence of embryonic deformations was observed in the most polluted deployment location, but larvae morphometry revealed no evidence of toxicity related to pollutant body burden. In conclusion, the in-situ exposure method was proven to work well, making it attractive for implementations in environmental monitoring programs.

Photo-enhanced toxicity of crude oil on early developmental stages of Atlantic cod (Gadus morhua).

Photo-enhanced toxicity of crude oil is produced by exposure to ultraviolet (UV) radiation. Atlantic cod (Gadus morhua) embryos were exposed to crude oil with and without UV radiation (290-400 nm) from 3 days post fertilization (dpf) until 6 dpf. Embryos from the co-exposure experiment were continually exposed to UV radiation until hatching at 11 dpf. Differences in body burden levels and cyp1a expression in cod embryos were observed between the exposure regimes. High doses of crude oil produced increased mortality in cod co-exposed embryos, as well as craniofacial malformations and heart deformities in larvae from both experiments. A higher number of differentially expressed genes (DEGs) and pathways were revealed in the co-exposure experiment, indicating a photo-enhanced effect of crude oil toxicity. Our results provide mechanistic insights into crude oil and photo-enhanced crude oil toxicity, suggesting that UV radiation increases the toxicity of crude oil in early life stages of Atlantic cod.

Cardiac dysfunction affects eye development and vision by reducing supply of lipids in fish.

Developing organisms are especially vulnerable to environmental stressors. Crude oil exposure in early life stages of fish result in multiple functional and developmental defects, including cardiac dysfunction and abnormal and smaller eyes. Phenanthrene (Phe) has a reversible impact on cardiac function, and under exposure Phe reduces cardiac contractility. Exposure to a known L-type channel blocker, nicardipine hydrochloride (Nic) also disrupts cardiac function and creates eye deformities. We aimed to investigate whether cardiac dysfunction was the major underlying mechanism of crude oil-, Phe- and Nic-induced eye malformations. We exposed Atlantic haddock (Melanogrammus aeglefinus) early embryos to Nic and crude oil (Oil) and late embryos/early larvae to Phe exposure. All three exposures resulted in cardiac abnormalities and lead to severe, eye, jaw and spinal deformities at early larval stages. At 3 days post hatching, larvae from the exposures and corresponding controls were dissected. Eyes, trunk, head and yolk sac were subjected to lipid profiling, and eyes were also subjected to transcriptomic profiling. Among most enriched pathways in the eye transcriptomes were fatty acid metabolism, calcium signaling and phototransduction. Changes in lipid profiles and the transcriptome suggested that the dysfunctional and abnormal eyes in our exposures were due to both disruption of signaling pathways and insufficient supply of essential fatty acids and other nutrients form the yolk.

Differential developmental toxicity of crude oil in early life stages of Atlantic halibut (Hippoglossus hippoglossus).

To further understand the complexity of developmental toxicity of dispersed oil and importance of exposure timing on fish early life stages, Atlantic halibut (Hippoglossus hippoglossus) were exposed to environmentally relevant concentrations through two embryonic developmental windows: the first period occurred during the epiboly process (named as "early embryonic exposure") and the second period overlapped the ontogenesis and cardiogenesis processes (named as "late embryonic exposure"). Following 72 hour oil exposure, embryos were transferred to clean seawater and a toxicity screening was performed in the yolk-sac larvae until first-feeding stages (56 days). The current study demonstrated that the exposure timing is essential for the development of toxic effects of crude oil in Atlantic halibut. Neither embryonic exposures (early or late) showed notable acute toxicity during exposure, yet both showed global latent teratogenic effects during yolk sac stages. Fish exposed during organogenesis (late) displayed stronger and more severe toxic effects than fish exposed during epiboly process (early), including reduced condition, severe craniofacial deformities and cardiovascular disruptions. The uptake level of polycyclic aromatic hydrocarbons into larval tissue and metabolic activity were greater following the late embryonic exposure and remained high during the depuration period at the highest exposure concentration. Overall, the long yolk sac stage development timing of Atlantic halibut makes this species a good candidate for evaluation of embryonic crude oil toxicity and its mechanisms.

Investigation by mass spectrometry and 32P post-labelling of DNA adducts formation from 1,2-naphthoquinone, an oxydated metabolite of naphthalene.

Naphthalene is the simplest representative of polycyclic aromatic hydrocarbons (PAHs). It is detected as major pollutant in the different compartments of the environment. This compound is considered by the international agency for research on cancer (IARC), the specialized cancer agency of the World Health Organisation (WHO), as a possible carcinogenic (group 2B) since 2002, mainly based on studies on chronic inhalation in rodent by the national toxicology program of the U.S. department of health and human services. In humans, its main metabolites correspond to derivatives substituted in position and 1 and 2 as 1,2-naphthoquinone (1,2-NphQ). Based on previous studies, 1,2-NphQ is supposed to react with DNA to form mostly depurinating adducts, a possible initiating step of carcinogenicity. To confirm this potentiality, adducts were synthetized by the reaction of 1,2-NphQ with 2'-deoxyguanosine (2'-dG) in N,N-dimethylformamide (DMF), water and calf thymus DNA. 2'-dG adducts were analyzed by 32P post-labelling, HPLC with ultra-violet detection and ultra-performance liquid chromatography coupled to tandem mass spectrometry (UHPLC-MS/MS). We found stable DNA adducts detected in DNA. We proposed a formation mechanism by a 1,4-Michael addition with 2'-dG. Adducts with 2'-deoxyxanthosine are formed after a spontaneous deamination of 2'-dG. These adducts are good candidates as biomarkers allowing evaluation of exposure to naphthalene and its derivatives in the development of pathologies such as cancer.

Untangling mechanisms of crude oil toxicity: Linking gene expression, morphology and PAHs at two developmental stages in a cold-water fish.

Early life stages of fish are highly sensitive to crude oil exposure and thus, short term exposures during critical developmental periods could have detrimental consequences for juvenile survival. Here we administered crude oil to Atlantic haddock (Melanogrammus aeglefinus) in short term (3-day) exposures at two developmental time periods: before first heartbeat, from gastrulation to cardiac cone stage (early), and from first heartbeat to one day before hatching (late). A frequent sampling regime enabled us to determine immediate PAH uptake, metabolite formation and gene expression changes. In general, the embryotoxic consequences of an oil exposure were more severe in the early exposure animals. Oil droplets on the eggshell resulted in severe cardiac and craniofacial abnormalities in the highest treatments. Gene expression changes of Cytochrome 1 a, b, c and d (cyp1a, b, c, d), Bone morphogenetic protein 10 (bmp10), ABC transporter b1 (abcb1) and Rh-associated G-protein (rhag) were linked to PAH uptake, occurrence of metabolites of phenanthrene and developmental and functional abnormalities. We detected circulation-independent, oil-induced gene expression changes and separated phenotypes linked to proliferation, growth and disruption of formation events at early and late developmental stages. Changes in bmp10 expression suggest a direct oil-induced effect on calcium homeostasis. Localized expression of rhag propose an impact on osmoregulation. Severe eye abnormalities were linked to possible inappropriate overexpression of cyp1b in the eyes. This study gives an increased knowledge about developmentally dependent effects of crude oil toxicity. Thus, our findings provide more knowledge and detail to new and several existing adverse outcome pathways of crude oil toxicity.

DNA damage and health effects in juvenile haddock (Melanogrammus aeglefinus) exposed to PAHs associated with oil-polluted sediment or produced water.

The research objective was to study the presence of DNA damages in haddock exposed to petrogenic or pyrogenic polyaromatic hydrocarbons (PAHs) from different sources: 1) extracts of oil produced water (PW), dominated by 2-ring PAHs; 2) distillation fractions of crude oil (representing oil-based drilling mud), dominated by 3-ring PAHs; 3) heavy pyrogenic PAHs, mixture of 4/5/6-ring PAHs. The biological effect of the different PAH sources was studied by feeding juvenile haddock with low doses of PAHs (0.3-0.7 mg PAH/kg fish/day) for two months, followed by a two-months recovery. In addition to the oral exposure, a group of fish was exposed to 12 single compounds of PAHs (4/5/6-ring) via intraperitoneal injection. The main endpoint was the analysis of hepatic and intestinal DNA adducts. In addition, PAH burden in liver, bile metabolites, gene and protein expression of CYP1A, GST activity, lipid peroxidation, skeletal deformities and histopathology of livers were evaluated. Juvenile haddock responded quickly to both intraperitoneal injection and oral exposure of 4/5/6-ring PAHs. High levels of DNA adducts were detected in livers three days after the dose of the single compound exposure. Fish had also high levels of DNA adducts in liver after being fed with extracts dominated by 2-ring PAHs (a PW exposure scenario) and 3-ring PAHs (simulating an oil exposure scenario). Elevated levels of DNA adducts were observed in the liver of all exposed groups after the 2 months of recovery. High levels of DNA adduct were found also in the intestines of individuals exposed to oil or heavy PAHs, but not in the PW or control groups. This suggests that the intestinal barrier is very important for detoxification of orally exposures of PAHs.

Undesirables in Mesopelagic Species and Implications for Food and Feed Safety-Insights from Norwegian Fjords.

The increase in the global population demands more biomass from the ocean as future food and feed, and the mesopelagic species might contribute significantly. In the present study, we evaluated the food and feed safety of six of the most abundant mesopelagic species in Norwegian fjords. Trace elements (i.e., arsenic, cadmium, mercury, and lead), organic pollutants (i.e., dioxins, furans, dioxin-like polychlorinated biphenyls, and polybrominated flame-retardants), and potentially problematic lipid compounds (i.e., wax esters and erucic acid) were analyzed and compared to existing food and feed maximum levels and intake recommendations. Furthermore, contaminant loads in processed mesopelagic biomass (protein, oil, and fish meal) was estimated using worst-case scenarios to identify possible food and feed safety issues. While most undesirables were low considering European food legislation, we identified a few potential food safety issues regarding high levels of fluoride in Northern krill, wax esters in glacier lanternfish, and long-chain monounsaturated fatty acids in silvery lightfish. Our estimates in processed biomass indicated high levels of undesirable trace elements in the protein fraction, frequently exceeding the maximum levels for feed ingredients. However, in fish meal, almost no exceedances were seen. In the oil fraction, dioxins and furans were above the maximum levels, given for food and feed ingredients. The present study is crucial to enable an evaluation of the value of these species; however, more data is needed before proceeding with large-scale harvesting of mesopelagic biomass.

Expression and localization of the aryl hydrocarbon receptors and cytochrome P450 1A during early development of Atlantic cod (Gadus morhua).

The aryl hydrocarbon receptor (Ahr) is a ligand-activated transcription factor that mediates the toxicity of dioxins and dioxin-like compounds (DLCs) in vertebrates. Two clades of the Ahr family exist in teleosts (Ahr1 and Ahr2), and it has been demonstrated that Ahr2 is the main protein involved in mediating the toxicity of dioxins and DLCs in most teleost species. Recently, we characterized the Atlantic cod (Gadus morhua) Ahr1a and Ahr2a receptors. To further explore a possible subfunction partitioning of Ahr1a and Ahr2a in Atlantic cod we have mapped the expression and localization of ahr1a and ahr2a in early developmental stages. Atlantic cod embryos were continuously exposed in a passive-dosing exposure system to the Ahr agonist, benzo[a]pyrene (B[a]P), from five days post fertilization (dpf) until three days post hatching (dph). Expression of ahr1a, ahr2a, and the Ahr-target genes, cyp1a and ahrrb, was assessed in embryos (8 dpf and 10 dpf) and larvae (3 dph) with quantitative real-time PCR analyses (qPCR), while in situ hybridization was used to assess the localization of expression of ahr1a, ahr2a and cyp1a. Quantitative measurements showed an increased cyp1a expression in B[a]P-exposed samples at all sampling points, and for ahr2a at 10 dpf, confirming the activation of the Ahr-signalling pathway. Furthermore, B[a]P strongly induced ahr2a and cyp1a expression in the cardiovascular system and skin, respectively, of embryos and larvae. Induced expression of both ahr2a and cyp1a was also revealed in the liver of B[a]P-exposed larvae. Our results suggest that Ahr2a is the major subtype involved in mediating responses to B[a]P in early developmental stages of Atlantic cod, which involves transcriptional regulation of biotransformation genes, such as cyp1a. The focused expression of ahr1a in the eye of embryos and larvae, and the presence of ahr2a transcripts in the jaws and fin nodes, further indicate evolved specialized roles of the two Ahrs in ontogenesis.

Embryonic Crude Oil Exposure Impairs Growth and Lipid Allocation in a Keystone Arctic Forage Fish.

As Arctic ice recedes, future oil spills pose increasing risk to keystone species and the ecosystems they support. We show that Polar cod (Boreogadus saida), an energy-rich forage fish for marine mammals, seabirds, and other fish, are highly sensitive to developmental impacts of crude oil. Transient oil exposures ≥300 µg/L during mid-organogenesis disrupted the normal patterning of the jaw as well as the formation and function of the heart, in a manner expected to be lethal to post-hatch larvae. More importantly, we found that exposure to lower levels of oil caused a dysregulation of lipid metabolism and growth that persisted in morphologically normal juveniles. As lipid content is critical for overwinter survival and recruitment, we anticipate Polar cod losses following Arctic oil spills as a consequence of both near-term and delayed mortality. These losses will likely influence energy flow within Arctic food webs in ways that are as-yet poorly understood.