Enhanced neurotoxic effect of PCB-153 when co-exposed with polystyrene nanoplastics in zebrafish larvae.

Nanoplastics (NPs) and persistent organic pollutants such as polychlorinated biphenyls (PCBs) are ubiquitous aquatic pollutants. The coexistence of these pollutants in the environment emphasises the need to study their combined toxicity. NPs can cross biological membranes and act as vectors for other pollutants, whereas PCBs are known for their ability to bioaccumulate and biomagnify. The present work aimed to study the combined toxicity of polystyrene NPs and PCB-153 using physiological (development, heart rate, respiration), behavioural (swimming behaviour) and molecular (transcriptome) endpoints in zebrafish larvae. The results show that exposure to NPs, PCB and their mixture significantly affected the development and respiration in zebrafish larvae. Larvae co-exposed to NPs and PCB exhibited significant hyperlocomotion, whereas no such effect was observed after exposure to NPs or PCB alone. The transcriptomic results revealed that NPs exposure significantly affected several pathways associated with DNA compaction and nucleosome assembly, whereas PCB exposure significantly affected critical neurogenic pathways. In contrast, co-exposure to NPs and PCB generated multi-faceted toxicity and suppressed neurobehavioural, immune-related and detoxification pathways. The study highlights the complex interplay between NPs and PCBs, and documents how the two toxicants in combination give a stronger effect than the single toxicants alone. Understanding the mixture toxicity of these two pollutants is important to assess the environmental risks and developing effective management strategies, ultimately safeguarding ecosystems and human health.

Mixture toxicity of 6PPD-quinone and polystyrene nanoplastics in zebrafish.

Plastic pollution, including micro- and nanoplastics, is a growing concern. Tyre-wear particles (TWPs) are the second largest source of microplastics in the ocean following abrasion of synthetic fibres. In addition to the particles themselves, TWPs contain many harmful chemicals, including 6PPD. This chemical reacts with atmospheric ozone and forms the toxic compound 6PPD-quinone (6PPDq), which poses a danger to aquatic life. There is a knowledge gap in understanding risks associated with the combined toxicity of nanoplastics (NPs) and 6PPDq. The present study aimed to investigate the toxicity of NPs and 6PPDq on adult zebrafish using phenotypic (behaviour, histology) and transcriptomic endpoints. Zebrafish were exposed to four treatments: control (contaminant-free), 50 µg/L 6PPDq, 3 mg/L polystyrene (PS)-NPs, and a combination of 50 µg/L 6PPDq and 3 mg/L PS-NPs. We did not observe locomotory dysregulation in zebrafish exposed to NPs. However, we found significant hyperlocomotion in zebrafish exposed to 6PPDq and this effect was even more substantial after co-exposure with PS-NPs. This study explores the molecular mechanisms behind these effects, identifying genes associated with neurotransmitters and fatty acid metabolism that were dysregulated by the co-exposure. Transcriptomic analysis further showed that both 6PPDq and PS-NPs impacted cellular processes associated with sterol biosynthesis, cholesterol metabolism, and muscle tissue development. The effects on these mechanisms were stronger in co-exposed zebrafish, indicating a heightened risk to cellular integrity and mitochondrial dysfunction. These results highlight the significance of mixture toxicity when studying the effects of NPs and associated chemicals like 6PPDq.

Gill epithelial cell line ASG-10 from Atlantic salmon as a new research tool for solving water quality challenges in aquaculture.

Here we evaluated the gill epithelial cell line ASG-10 from Atlantic salmon, as an in vitro model for research on known water quality challenges in aquaculture. Ammonia/ammonium (NH3/NH4+), a recognized challenge in water-intensive recirculating aquaculture systems (RAS), induced lysosomal vacuolization, reduced protein degradation and cell migration of the ASG-10 cells. Aluminium (Aln+), another challenge in freshwater aquaculture facilities had only minor effects. Next, we investigated the tolerance for direct water exposure of ASG-10. The cells tolerated water with osmolarity between 169 and 419 mOsmol/kg for 24 h. However, cells exposed for 3 h to water at 863 mOsmol/kg changed cellular morphology and induced gene expression related to stress (gpx1, casp3, hsp70), and after 24 h exposure cellular viability was severely reduced. Nevertheless, when the cells were grown in transwell inserts, they tolerated 863 mOsmol/kg for 3 h and induction of stress response associated genes was considerably reduced. Lastly, the ASG-10 cells were exposed to water samples, with no known quality issues, from different aquaculture facilities. The cells showed no differences in viability or morphology compared to their representative control. In conclusion, the ASG-10 cell line is a promising in vitro model to study water quality challenges and whole water samples.

Ecotoxicological assessment of Cu-rich acid mine drainage of Sulitjelma mine using zebrafish larvae as an animal model.

Acid mine drainage (AMD) is widely acknowledged as a substantial threat to the biodiversity of aquatic ecosystems. The present study aimed to study the toxicological effects of Cu-rich AMD from the Sulitjelma mine in zebrafish larvae. The AMD from this mine was found to contain elevated levels of dissolved metals including Mg (46.7 mg/L), Al (20.2 mg/L), Cu (18.3 mg/L), Fe (19.8 mg/L) and Zn (10.6 mg/L). To investigate the toxicological effects, the study commenced by exposing zebrafish embryos to various concentrations of AMD (ranging from 0.75% to 9%) to determine the median lethal concentration (LC50). Results showed that 96 h LC50 for zebrafish larvae following AMD exposure was 2.86% (95% CI: 2.32-3.52%). Based on acute toxicity results, zebrafish embryos (<2 hpf) were exposed to 0.1% AMD (Cu: 21.7 µg/L) and 0.45% AMD (Cu: 85.7 µg/L) for 96 h to assess development, swimming behaviour, heart rate, respiration and transcriptional responses at 116 hpf. Light microscopy results showed that both 0.1% and 0.45% AMD reduced the body length, eye size and swim bladder area of zebrafish larvae and caused phenotypic abnormalities. Swimming behaviour results showed that 0.45% AMD significantly decreased the locomotion of zebrafish larvae. Heart rate was not affected by AMD exposure. Furthermore, exposure caused a significant increase in oxygen consumption indicating vascular stress in developing larvae. Taken altogether, the study shows that even heavily diluted AMD with environmentally relevant levels of Cu caused toxicity in zebrafish larvae.

Lufenuron treatment temporarily represses gene expression and affects the SUMO pathway in liver of Atlantic salmon.

Lufenuron is a benzoylurea insecticide currently in use to combat sea lice infestation in salmon aquaculture in Chile. With pending approval in Norway, the aim of this work was to study the uptake and toxicity of lufenuron in liver tissue of Atlantic salmon. Juvenile salmon weighing 40 g were given a standard 7-day oral dose, and bioaccumulation and transcriptional responses in the liver were examined 1 day after the end-of-treatment (day 8) and after 1 week of elimination (day 14). Bioaccumulation levels of lufenuron were 29 ± 3 mg/kg at day 8 and 14 ± 1 mg/kg at day 14, indicating relatively rapid clearance. However, residues of lufenuron were still present in the liver after 513 days of depuration. The exposure gave a transient inhibition of transcription in the liver at day 8 (2437 significant DEGs, p-adj < .05), followed by a weaker compensatory response at day 14 (169 significant DEGs). Pathways associated with RNA metabolism such as the sumoylation pathway were most strongly affected at day 8, while the apelin pathway was most profoundly affected at day 14. In conclusion, this study shows that lufenuron easily bioaccumulates and that a standard 7-day oral dose induces a transient inhibition of transcription in liver of salmon.

Effects of gadolinium (Gd) and a Gd-based contrast agent (GBCA) on early life stages of zebrafish (Danio rerio).

Gadolinium (Gd) is one of the rare earth elements (REY) and is widely used in magnetic resonance imaging (MRI) contrast agents. Anthropogenic Gd enrichment has frequently been found in wastewater treatment plant effluents in industrialised countries, rising concerns regarding effects on aquatic biota. This study investigates the acute toxicity and sublethal effects of Gd in two forms, as inorganic salt (GdCl3) and as Gd-based contrast agent (GBCA), on early life stages of zebrafish (Danio rerio). Nominal exposure concentrations ranged from 3 to 3000 µg L-1, with an exposure duration of 96 h. None of the two tested compounds were acutely toxic to embryos and larvae. Similarly, we did not observe any effects on larval development and locomotive behaviour. However, we found significant changes in the brain activity of larvae exposed to the highest concentrations of GdCl3 and the GBCA. Our findings show that Gd can have sublethal effects on developing fish at lower concentrations than reported previously, highlighting the necessity of investigating the long-term fate and effects of GBCAs released into the aquatic environment.

Effects of mine tailing exposure on the development of early life stages of the marine copepod Calanus finmarchicus.

The demand for mineral resources is increasing mining activities worldwide. In Norway, marine tailing disposal (MTD) is practiced, introducing mineral particles into fjord ecosystems. We investigated the effects of two concentrations (high and low) of fine tailings from a CaCO3 processing plant on early life stages of the marine copepod Calanus finmarchicus. Results show that the exposure did not significantly impact hatching success or development in non- and early feeding life stages. However, feeding stage nauplii ingested tailings, which caused a significantly slower development in later nauplii stages in high exposure groups, with most individuals being two stages behind the control group. Further, high mortality occurred in late nauplii and early copepodite stages in low exposure groups, which could be caused by insufficient energy accumulation and depleted energy reserves during development. Individuals exposed to high exposure concentrations seemed to survive by arresting development and potentially by reduced activity, thereby conserving energy reserves. In nature, slower development could affect lipid storage buildup and reproduction.

Microbial oil, alone or paired with ß-glucans, can control hypercholesterolemia in a zebrafish model.

Dyslipidemia is often associated with unhealthy dietary habits, and many mammalian studies have explored the mode of action of certain bioactive compounds such as ß-glucans and n-3 PUFAs to understand their potential to normalize the lipid metabolism. There are only a few investigations that adopted omic approaches to unveil their combined effect on hypercholesterolemia. Zebrafish (Danio rerio) was used as a model organism to reveal the efficacy of Schizochytrium oil and ß-glucans (from Euglena gracilis and Phaeodactylum tricornutum) against cholesterol-rich diet induced dyslipidemia. One of the folowing four diets was fed to a particular group of fish: a control high-cholesterol diet, a Schizochytrium oil diet or one of the two diets containing the oil and ß-glucan. The plasma HDL, expression of hepatic genes linked to, among others, ferric ion binding and plasma phosphatidylcholines were higher and plasma cholesterol esters and triacylglycerols were lower in the microbial oil-fed fish compared to the fish fed high cholesterol diet. While the fish fed a mix of microbial oil and Euglena ß-glucan had lower plasma triacylglycerols and expression of hepatic genes linked to PPAR signaling pathway and enriched biosynthesis of plasma unsaturated fatty acids, the fish fed microbial oil-Phaeodactylum ß-glucan combination had lower abundance of triacylglycerols rich in saturated and mono-unsaturated fatty acids and cholesterol esters in the plasma.

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.

Protective mechanisms of a microbial oil against hypercholesterolemia: evidence from a zebrafish model.

A Western diet elevates the circulating lipoprotein and triglyceride levels which are the major risk factors in cardiovascular disease (CVD) development. Consumption of long-chain omega-3 fatty acids can stall the disease progression. Although these fatty acids can significantly impact the intestine under a hypercholesterolemic condition, the associated changes have not been studied in detail. Therefore, we investigated the alterations in the intestinal transcriptome along with the deviations in the plasma lipids and liver histomorphology of zebrafish offered DHA- and EPA-rich oil. Fish were allocated to 4 dietary treatments: a control group, a high cholesterol group and microbial oil groups with low (3.3%) and high (6.6%) inclusion levels. We quantified the total cholesterol, lipoprotein and triglyceride levels in the plasma. In addition, we assessed the liver histology, intestinal transcriptome and plasma lipidomic profiles of the study groups. The results suggested that higher levels of dietary microbial oil could control the CVD risk factor indices in zebrafish plasma. Furthermore, microbial oil-fed fish had fewer liver vacuoles and higher mRNA levels of genes involved in ß-oxidation and HDL maturation. Analyses of the intestine transcriptome revealed that microbial oil supplementation could influence the expression of genes altered by a hypercholesterolemic diet. The plasma lipidomic profiles revealed that the higher level of microbial oil tested could elevate the long-chain poly-unsaturated fatty acid content of triglyceride species and lower the concentration of several lysophosphatidylcholine and diacylglycerol molecules. Our study provides insights into the effectiveness of microbial oil against dyslipidemia in zebrafish.

Polystyrene nanoplastics enhance the toxicological effects of DDE in zebrafish (Danio rerio) larvae.

Anthropogenic releases of plastics, persistent organic pollutants (POPs), and heavy metals can impact the environment, including aquatic ecosystems. Nanoplastics (NPs) have recently emerged as pervasive environmental pollutants that have the ability to adsorb POPs and can cause stress in organisms. Among POPs, DDT and its metabolites are ubiquitous environmental pollutants due to their long persistence. Despite the discontinued use of DDT in Europe, DDT and its metabolites (primarily p,p'-DDE) are still found at detectable levels in fish feed used in salmon aquaculture. Our study aimed to look at the individual and combined toxicity of NPs (50 mg/L polystyrene) and DDE (100 µg/L) using zebrafish larvae as a model. We found no significant morphological, cardiac, respiratory, or behavioural changes in zebrafish larvae exposed to NPs alone. Conversely, morphological, cardiac and respiratory alterations were observed in zebrafish larvae exposed to DDE and NPs + DDE. Interestingly, behavioural changes were only observed in zebrafish larvae exposed to NPs + DDE. These findings were supported by RNA-seq results, which showed that some cardiac, vascular, and immunogenic pathways were downregulated only in zebrafish larvae exposed to NPs + DDE. In summary, we found an enhanced toxicological impact of DDE when combined with NPs.

Developmental defects and behavioral changes in a diet-induced inflammation model of zebrafish.

Soybean meal evokes diet-induced intestinal inflammation in certain fishes. Although the molecular aspects of soybean-induced intestinal inflammation in zebrafish are known, the impact of the inflammatory diet on fish behavior remain largely underexplored. We fed zebrafish larvae with three diets - control, soybean meal and soybean meal with ß-glucan to gain deeper insight into the behavioral changes associated with the soybean meal-induced inflammation model. We assessed the effect of the diets on the locomotor behavior, morphological development, oxygen consumption and larval transcriptome. Our study revealed that dietary soybean meal can reduce the locomotor activity, induce developmental defects and increase the oxygen demand in zebrafish larvae. Transcriptomic analysis pointed to the suppression of genes linked to visual perception, organ development, phototransduction pathway and activation of genes linked to the steroid biosynthesis pathway. On the contrary, ß-glucan, an anti-inflammatory feed additive, counteracted the behavioral and phenotypic changes linked to dietary soybean. Although we did not identify any differentially expressed genes from the soybean meal alone fed group vs soybean meal + ß-glucan-fed group comparison, the unique genes from the comparisons of the two groups with the control likely indicate reduction in inflammatory cytokine signaling, inhibition of proteolysis and induction of epigenetic modifications by the dietary glucan. Furthermore, we found that feeding an inflammatory diet at the larval stage can lead to long-lasting developmental defects. In conclusion, our study reveals the extra-intestinal manifestations associated with soybean meal-induced inflammation model.

Dietary chlorpyrifos-methyl exposure impair transcription of immune-, detoxification- and redox signaling genes in leukocytes isolated from cod (Gadus morhua).

Inclusion of new environmental toxicants increase with the amount of plant ingredients substituting marine proteins and oils in feed for farmed Atlantic salmon (Salma salar). Agricultural pesticides like chlorpyrifos-methyl, present in commercial salmon feeds, may affect salmon immune and detoxification responses. Atlantic cod (Gadus morhua), surrounding the net pens, grazing on feces and uneaten pellets may be affected accordingly. The aim of this study was to analyze transcription responses in Atlantic cod head kidney tissue and isolated leukocytes following dietary chlorpyrifos-methyl inclusions and possible interactions with proinflammatory signals. Head kidney tissues and leukocytes were isolated from cod fed diets contaminated with chlorpyrifos-methyl (0.5 mg/kg, 2.4 mg/kg, 23.2 mg/kg) for 30 days. The isolated leukocytes were further challenged with bacteria (lipopolysaccharide (LPS), virus (polyinosinic acid:polycytidylic acid (PIC) mimic and l-arginine, an immuno-modulating amino acid, in vitro. The LPS-induced transcription of the interleukin genes il-1ß, il-6, il-8 increased in leukocytes isolated from cod fed chlorpyrifos-methyl 23.2 mg/kg, compared to cod fed the control diet, indicating increased inflammation. Transcriptional levels of carnitine palmitoyl transferase (cpt1a), aryl hydrogen receptor (ahr) and catalase (cat) were all reduced by dietary inclusions of chlorpyrifos-methyl in the leukocytes. The findings suggests that dietary chlorpyrifos-methyl exposure impair inflammation, detoxification and redox signaling in cod leukocytes.

Distinct polymer-dependent sorption of persistent pollutants associated with Atlantic salmon farming to microplastics.

Interactions of microplastics and persistent organic pollutants (POPs) associated with Atlantic salmon farming were studied to assess the potential role of microplastics in relation to the environmental impact of aquaculture. HDPE, PP, PET and PVC microplastics placed for 3 months near fish farms sorbed POPs from aquafeeds. PET and PVC sorbed significantly higher levels of dioxins and PCBs compared to HDPE, while the levels sorbed to PP were intermediate and did not differ statistically from PET, PVC or HDPE. In addition, the composition of dioxins accumulated in caged blue mussels did not reflect the patterns observed on the microplastics, probably due to polymer-specific affinity of POPs. In conclusion, the results of this study show that microplastics occurring near fish farms can sorb aquafeed-associated POPs and, therefore, microplastics could potentially be vectors of such chemicals in the marine environment and increase the environmental impact of fish farming.

Toxicological effects of 6PPD and 6PPD quinone in zebrafish larvae.

N-(1,3-dimethylbutyl)-N'-phenyl-p-phenylenediamine (6PPD) is the most widely used antioxidant in automobile tyres and many rubber products. We investigated the impact of 6PPD and 6PPD quinone on acute toxicity, morphology, swimming behaviour, heart rate, and oxygen consumption in zebrafish larvae. Zebrafish embryos were exposed to 6PPD and 6PPD quinone at concentrations of 1, 10, and 25 µg/L during the development period of 1-96 hpf. In the present study, 6PPD quinone was found to be toxic to zebrafish larvae with a 24 h LC50 of 308.67 µg/L. No significant mortality was observed at any of the tested concentrations. A dose-dependent reduction in swimming performance was observed in the exposed larvae at 116 hpf for both toxicants. Overall, our study shows that exposure of zebrafish embryos to 6PPD and 6PPD quinone at environmentally relevant concentrations (1 µg/L) does not affect its behaviour. However, exposure to higher but still sublethal concentrations of 6PPD and 6PPD quinone (10 and 25 µg/L) can affect behavioural endpoints. These findings reveal the toxicity of 6PPD and 6PPD quinone to early life stages of fish.

Exposure to low environmental copper concentrations does not affect survival and development in Atlantic cod (Gadus morhua) early life stages.

In this study we investigated potential impacts of Cu exposure at low, environmentally relevant, concentrations on early live stages of Atlantic cod (Gadus morhua). Cod embryos and larvae were exposed to 0.5 µg/L (low), 2 µg/L (medium), and 6 µg/L (high) Cu from 4 to 17 days post fertilisation (dpf). Hatching success, mortality, oxygen consumption, biometric traits, and malformations were determined. A dynamic energy budget (DEB) model was applied to identify potential impacts on bioenergetics. A positive correlation was found between Cu exposure concentrations and Cu body burden in eggs, but not in larvae. The tested concentrations did not increase mortality in neither embryos nor larvae, or larvae deformations. Further, the DEB model did not indicate effects of the tested Cu concentrations.

Sources, distribution and effects of rare earth elements in the marine environment: Current knowledge and research gaps.

Rare earth elements and yttrium (REY) are critical elements for a wide range of applications and consumer products. Their growing extraction and use can potentially lead to REY and anthropogenic-REY chemical complexes (ACC-REY) being released in the marine environment, causing concern regarding their potential effects on organisms and ecosystems. Here, we critically review the scientific knowledge on REY sources (geogenic and anthropogenic), factors affecting REY distribution and transfer in the marine environment, as well as accumulation in- and effects on marine biota. Further, we aim to draw the attention to research gaps that warrant further scientific attention to assess the potential risk posed by anthropogenic REY release. Geochemical processes affecting REY mobilisation from natural sources and factors affecting their distribution and transfer across marine compartments are well established, featuring a high variability dependent on local conditions. There is, however, a research gap with respect to evaluating the environmental distribution and fate of REY from anthropogenic sources, particularly regarding ACC-REY, which can have a high persistence in seawater. In addition, data on organismal uptake, accumulation, organ distribution and effects are scarce and at best fragmentary. Particularly, the effects of ACC-REY at organismal and community levels are, so far, not sufficiently studied. To assess the potential risks caused by anthropogenic REY release there is an urgent need to i) harmonise data reporting to promote comparability across studies and environmental matrices, ii) conduct research on transport, fate and behaviour of ACC-REY vs geogenic REY iii) deepen the knowledge on bioavailability, accumulation and effects of ACC-REY and REY mixtures at organismal and community level, which is essential for risk assessment of anthropogenic REY in marine ecosystems.

Effects of marine mine tailing exposure on the development, growth, and lipid accumulation in Calanus finmarchicus.

Marine tailing disposal (MTD) is sometimes practiced as an alternative to traditional mine tailing deposition on land. Environmental challenges connected to MTD include spreading of fine particulate matter in the water column and the potential release of metals and processing chemicals. This study investigated if tailing exposure affects the marine copepod Calanus finmarchicus, and whether effects are related to exposure to mineral particles or the presence of metals and/or processing chemicals in the tailings. We investigated the impacts of three different tailing compositions: calcium carbonate particles with and without processing chemicals and fine-grained tailings from a copper ore. Early life stages of C. finmarchicus were exposed over several developmental stages to low and high suspension concentrations for 15 days, and their development, oxygen consumption and biometry determined. The data was fitted in a dynamic energy budget (DEB) model to determine mechanisms underlying responses and to understand the primary modes of action related to mine tailing exposure. Results show that copepods exposed to tailings generally exhibited slower growth and accumulated less lipids. The presence of metals and processing chemicals did not influence these responses, suggesting that uptake of mineral particles was responsible for the observed effects. This was further supported by the applied DEB model, confirming that ingestion of tailing particles while feeding can result in less energy being available for growth and development.

Polycyclic aromatic hydrocarbons modulate the activity of Atlantic cod (Gadus morhua) vitamin D receptor paralogs in vitro.

Vitamin D receptor (VDR) mediates the biological function of the steroid hormone calcitriol, which is the metabolically active version of vitamin D. Calcitriol is important for a wide array of physiological functions, including calcium and phosphate homeostasis. In contrast to mammals, which harbor one VDR encoding gene, teleosts possess two orthologous vdr genes encoding Vdr alpha (Vdra) and Vdr beta (Vdrb). Genome mining identified the vdra and vdrb paralogs in the Atlantic cod (Gadus morhua) genome, which were further characterized regarding their phylogeny, tissue-specific expression, and transactivational properties induced by calcitriol. In addition, a selected set of polycyclic aromatic hydrocarbons (PAHs), including naphthalene, phenanthrene, fluorene, pyrene, chrysene, benzo[a]pyrene (BaP), and 7-methylbenzo[a]pyrene, were assessed for their ability to modulate the transcriptional activity of gmVdra and gmVdrb in vitro. Both gmVdra and gmVdrb were activated by calcitriol with similar potencies, but gmVdra produced significantly higher maximal fold activation. Notably, none of the tested PAHs showed agonistic properties towards the Atlantic cod Vdrs. However, binary exposures of calcitriol together with phenanthrene, fluorene, or pyrene, antagonized the activation of gmVdra, while chrysene and BaP significantly potentiated the calcitriol-mediated activity of both receptors. Homology modeling, solvent mapping, and docking analyses complemented the experimental data, and revealed a putative secondary binding site in addition to the canonical ligand-binding pocket (LBP). Calcitriol was predicted to interact with both binding sites, whereas PAHs docked primarily to the LBP. Importantly, our in vitro data suggest that PAHs can interact with the paralogous gmVdrs and interfere with their transcriptional activities, and thus potentially modulate the vitamin D signaling pathway and contribute to adverse effects of crude oil and PAH exposures on cardiac development and bone deformities in fish.

Effects of mine tailing exposure on early life stages of cod (Gadus morhua) and haddock (Melanogrammus aeglefinus).

Mining and processing of minerals produce large quantities of tailings as waste. Some countries, including Norway, allow disposal of mine tailings in the sea. In this study we investigated the impacts of tailings from a calcium carbonate (CaCO3) processing plant on early live stages of haddock (Melanogrammus aeglefinus) and Atlantic cod (Gadus morhua). Fish eggs (3 days post fertilisation; dpf) were exposed for 48 h to three concentrations of tailings, nominally 1 mg L-1 (low, L); 10 mg L-1 (medium, M) and 100 mg L-1 (high, H); with L and M representing concentrations occurring at tailing release points. Results show that tailings rapidly adhered to eggs of both species, causing negative buoyancy (sinking of eggs) in M and H exposures. While tailings remained on egg surfaces in both species also after exposure termination, adhesion seemed more pronounced in cod, leading to larger impacts on buoyancy even after exposure. Tailing exposure further induced early hatching and significantly reduced survival in M and H exposed embryos in both fish species, and in cod from the L exposure group. Moreover, tailing exposure caused reduced survival and malformations in larvae, potentially related to premature hatching. This study shows that mineral particles adhere to haddock and cod eggs, affecting egg buoyancy, survival and development.