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.

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.

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.

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.

Growth and metabolism of adult polar cod (Boreogadus saida) in response to dietary crude oil.

The increasing human presence in the Arctic shelf seas, with the expansion of oil and gas industries and maritime shipping, poses a risk for Arctic marine organisms such as the key species polar cod (Boreogadus saida). The impact of dietary crude oil on growth and metabolism of polar cod was investigated in the early spring (March-April) when individuals are expected to be in a vulnerable physiological state with poor energy stores. Adult polar cod were exposed dietarily to three doses of Kobbe crude oil during an eight weeks period and followed by two weeks of depuration. Significant dose-responses in exposure biomarkers (hepatic ethoxyresorufine-O-deethylase [EROD] activity and 1-OH phenanthrene metabolites in bile) indicated that polycyclic aromatic hydrocarbons (PAHs) were bioavailable. Condition indices (i.e. Fulton's condition factor, hepatosomatic index), growth, whole body respiration, and total lipid content in the liver were monitored over the course of the experiment. The majority of females were immature, while a few had spawned during the season and showed low hepatic lipid content during the experiment. In contrast, males were all, except for one immature individual, in a post-spawning stage and had larger hepatic energy stores than females. Most specimens, independent of sex, showed a loss in weight, that was exacerbated by exposure to crude oil and low hepatic liver lipids. Furthermore, females exposed to crude oil showed a significant elevation of oxygen consumption compared to controls, although not dose-dependent. This study highlights the importance of the energy status of individuals for their response to a crude oil exposure.

Genomic architecture of haddock (Melanogrammus aeglefinus) shows expansions of innate immune genes and short tandem repeats.

BACKGROUND: Increased availability of genome assemblies for non-model organisms has resulted in invaluable biological and genomic insight into numerous vertebrates, including teleosts. Sequencing of the Atlantic cod (Gadus morhua) genome and the genomes of many of its relatives (Gadiformes) demonstrated a shared loss of the major histocompatibility complex (MHC) II genes 100 million years ago. An improved version of the Atlantic cod genome assembly shows an extreme density of tandem repeats compared to other vertebrate genome assemblies. Highly contiguous assemblies are therefore needed to further investigate the unusual immune system of the Gadiformes, and whether the high density of tandem repeats found in Atlantic cod is a shared trait in this group. RESULTS: Here, we have sequenced and assembled the genome of haddock (Melanogrammus aeglefinus) - a relative of Atlantic cod - using a combination of PacBio and Illumina reads. Comparative analyses reveal that the haddock genome contains an even higher density of tandem repeats outside and within protein coding sequences than Atlantic cod. Further, both species show an elevated number of tandem repeats in genes mainly involved in signal transduction compared to other teleosts. A characterization of the immune gene repertoire demonstrates a substantial expansion of MCHI in Atlantic cod compared to haddock. In contrast, the Toll-like receptors show a similar pattern of gene losses and expansions. For the NOD-like receptors (NLRs), another gene family associated with the innate immune system, we find a large expansion common to all teleosts, with possible lineage-specific expansions in zebrafish, stickleback and the codfishes. CONCLUSIONS: The generation of a highly contiguous genome assembly of haddock revealed that the high density of short tandem repeats as well as expanded immune gene families is not unique to Atlantic cod - but possibly a feature common to all, or most, codfishes. A shared expansion of NLR genes in teleosts suggests that the NLRs have a more substantial role in the innate immunity of teleosts than other vertebrates. Moreover, we find that high copy number genes combined with variable genome assembly qualities may impede complete characterization of these genes, i.e. the number of NLRs in different teleost species might be underestimates.

Vertebral column deformity in adult wild haddock (Melanogrammus aeglefinus).

This study describes radiological malformations in the vertebral column of adult sexually mature wild haddock, Melanogrammus aeglefinus collected from Masfjorden, Western Norway, in 2014 and 2015. There were seven deformed fish (33% of sampled fish), five with mild deformities and two with severe deformities. The deformity types discovered were lordosis, vertebra body asymmetry, compression, fusion (ankylosis), remodelling, and haemal spine hyperostosis. The two severely deformed fish were as follows: (a) one specimen with 29 deformed vertebrae within the region vertebrae nos. 1-30, including two sites of lordosis; (b) one specimen with 15 deformed vertebrae within the region vertebrae nos. 31-52 (most caudal vertebrae). The results of this study show that wild haddock can survive severe vertebra deformities, both in the trunk and tail region, and reach adulthood and sexual maturity. Whether normal nondeformed wild conspecifics would mate and spawn with these individuals and thereby complete the life cycle of the deformed individuals is, however, unknown. A possible link between bone fracture and fish hyperostosis is discussed.

Developmental transcriptomics in Atlantic haddock: Illuminating pattern formation and organogenesis in non-model vertebrates.

Gadiforms such as Atlantic haddock comprise some of the world's most economically important fisheries. Understanding the early life history of these fish is a prerequisite for predicting effects of a changing environment and increased human activities. Robust assessment of the effects of environmental impacts on the embryos of non-model vertebrates is hampered by a lack of molecular resources and detailed knowledge regarding the regulation of genes and pathways in early development. Here we used mRNA sequencing to link transcriptional changes to developmental processes in haddock, specifically, pattern formation and organogenesis. Temporal expression of key developmental genes was tightly anchored to either the appearance of visible structures or cellular processes characterised in model organisms. These findings demonstrate the high potential of developmental transcriptomics as an analytical tool for improved understanding of pathophysiological mechanisms leading to abnormal development in any vertebrate.

Application of gas chromatography/tandem mass spectrometry to determine a wide range of petrogenic alkylated polycyclic aromatic hydrocarbons in biotic samples.

RATIONALE: Due to the inherent toxicity of these compounds, analysis of petrogenic alkylated polycyclic aromatic hydrocarbons (PAHs) and heteroaromatic compounds in environmental samples is crucial. However, most existing methods are not sensitive enough when the analyte concentrations are low. Gas chromatography/tandem mass spectrometry (GC/MS/MS) is well suited to tackle this challenge, but methods are yet to be fully developed. METHODS: A gas chromatograph coupled with a triple quadrupole mass spectrometer operated in multiple reaction monitoring (MRM) mode was applied for the analysis. MRM transitions and analytical conditions were optimized for the cluster analysis of 24 different compound classes of alkylated PAHs and heteroaromatics present in crude oils. Further, the method was verified and applied to analyze the body burden of these compounds in crude oil-exposed haddock eggs. RESULTS: A comprehensive and sensitive method for the determination of C1-C4 alkylated naphthalenes, benzothiophenes, fluorenes, phenanthrenes, dibenzothiophenes, chrysenes, pyrenes and fluoranthenes was developed. The method showed good precision (RSD <15%), and the obtained concentrations were similar to those obtained by traditional GC/MS analysis. CONCLUSIONS: We demonstrate the successful application of this method towards crude oil-exposed fish eggs, where we are able to precisely measure total PAH body burden below 0.5 µg/g. The demonstrated method facilitates further miniaturization in sample preparation. Copyright © 2016 John Wiley & Sons, Ltd.

Optimization and comparison of miniaturized extraction techniques for PAHs from crude oil exposed Atlantic cod and haddock eggs.

Two miniaturized extraction methods for a wide range of 2-6 ring polycyclic aromatic hydrocarbons (PAHs) and their alkylated homologues in small lipid-rich biota samples (≤100 mg) have been developed. Both methods utilize liquid extraction (LE) prior to a clean-up step using either normal phase solid phase extraction (SPE) or mixed-phase dispersive SPE (dSPE). Optimization of the methods was achieved by comparing the type and amount of sorbents, drying agents, and solvents used. In order to improve the limits of detection (LOD) of target PAHs under high sensitivity gas chromatography-tandem mass spectrometry analysis, specific emphasis was given to minimizing lipid co-extraction. The optimized LE-SPE method comprised extraction with dichloromethane/n-hexane (1:1, v/v) and clean-up by silica SPE, whereas the optimized LE-dSPE method comprised extraction with acetonitrile and clean-up with PSA and C18 sorbents. The methods were validated and directly compared through the analysis of Atlantic cod (Gadus morhua) and haddock (Melanogrammus aeglefinus) eggs exposed to oil. The LE-SPE method resulted in lower levels of co-extracted lipids (14.1 ± 1.7 ng/µL) than the LE-dSPE method (60 ± 14 ng/µL). Achieved PAH LODs for the LE-SPE method were typically an order of magnitude lower (<5 ng/g) than for the LE-dSPE method (<125 ng/g). The LE-SPE method offers the possibility for PAH analysis of small samples of fish eggs (~100 mg) exposed to small quantities of crude oil (~1-10 µg/L total PAHs).

Dispersants have limited effects on exposure rates of oil spills on fish eggs and larvae in shelf seas.

Early life stages of fish are particularly vulnerable to oil spills. Simulations of overlap of fish eggs and larvae with oil from different oil-spill scenarios, both without and with the dispersant Corexit 9500, enable quantitative comparisons of dispersants as a mitigation alternative. We have used model simulations of a blow out of 4500 m(3) of crude oil per day (Statfjord light crude) for 30 days at three locations along the Norwegian coast. Eggs were released from nine different known spawning grounds, in the period from March 1st until the end of April, and all spawning products were followed for 90 days from the spill start at April first independent of time for spawning. We have modeled overlap between spawning products and oil concentrations giving a total polycyclic hydrocarbon (TPAH) concentration of more than 1.0 or 0.1 ppb (µg/l). At these orders of magnitude, we expect acute mortality or sublethal effects, respectively. In general, adding dispersants results in higher concentrations of TPAHs in a reduced volume of water compared to not adding dispersants. Also, the TPAHs are displaced deeper in the water column. Model simulations of the spill scenarios showed that addition of chemical dispersant in general moderately decreased the fraction of eggs and larvae that were exposed above the selected threshold values.

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.

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.

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.

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.

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.

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.

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.

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.

Transcriptional effects of dietary exposure of oil-contaminated Calanus finmarchicus in Atlantic herring (Clupea harengus).

Suppression subtractive hybridization (SSH) cDNA library construction and characterization was used to identify differentially regulated transcripts from oil exposure in liver of male Atlantic herring (Clupea harengus) fed a diet containing 900 mg crude oil/kg for 2 mo. In total, 439 expressed sequence tags (EST) were sequenced, 223 from the forward subtracted library (enriched for genes putatively upregulated by oil exposure) and 216 from the reverse subtracted library (enriched for genes putatively downregulated by oil exposure). Follow-up reverse-transcription (RT) quantitative polymerase chain reaction (qPCR) analyses of gene transcription were conducted on additional herring exposed to food containing 9 (low), 90 (medium), and 900 (high) mg crude oil/kg feed for 2 mo. Chronic exposure of Atlantic herring to an oil-contaminated diet mediated upregulation of transcripts encoding antifreeze proteins, proteins in the classical complement pathway (innate immunity), and iron-metabolism proteins. Gene ontology (GO) analysis showed that "cellular response to stress," "regulation to biological quality," "response to abiotic stimuli," and "temperature homeostasis" were the most affected go at the biological processes level, and "carbohydrate binding," "water binding," and "ion binding" at the molecular function level. Of the genes examined with RT-qPCR, CYP1A, antifreeze protein, retinol binding protein 1, deleted in malignant brain tumor 1, and ovary-specific C1q-like factor demonstrated a significant upregulation. Myeloid protein 1, microfibrillar-associated protein 4, WAP65, and pentraxin were downregulated in liver of fish from the high exposure group. In conclusion, this study suggests that 2 mo of oil exposure affected genes encoding proteins involved in temperature homeostasis and possible membrane stability in addition to immune-responsive proteins in Atlantic herring.