Dynamic Links between Lipid Storage, Toxicokinetics and Mortality in a Marine Copepod Exposed to Dimethylnaphthalene

Efficiently assessing and managing the risks of pollution in the marine environment requires mechanistic models for toxic effects. The General Unified Threshold model for Survival (GUTS) provides a framework for deriving toxicokinetic-toxicodynamic (TKTD) models for the end point survival. Two recurring questions in the application of GUTS concern the most appropriate death mechanism, and whether the total body residue is a proper dose metric for toxic effects. We address these questions with a case study for dimethylnaphthalene in the marine copepod <i>Calanus finmarchicus</i>. A detailed analysis revealed that body residues were best explained by representing copepods with two toxicokinetic compartments: separating structural biomass and lipid storage. Toxicity is most likely related to the concentration in structure, which led to identification of “stochastic death” as the most appropriate death mechanism. Interestingly, the parametrized model predicts that lipid content will have only minor influence on short-term toxicity. However, the toxicants stored in lipids may have more substantial impacts in situations not included in our experiments (e.g., during diapause and gonad maturation), and for contaminant transfer to eggs and copepod predators

Use of TLM derived models to estimate toxicity of weathered MC252 oil based on conventional chemical data and the potential impact of unresolved polar components

Target lipid model (TLM) and toxic unit (TU) approaches were applied to ecotoxicity and chemistry data from low-energy WAFs (LE-WAFs) of source and weathered crude oils originating from the Deepwater Horizon oil spill. The weathered oils included artificially weathered oils and naturally weathered samples collected in the Gulf of Mexico after the spill. Oil weathering greatly reduced the concentrations of identified LE-WAF components, however, the mass of uncharacterized polar material (UPC) in the LE-WAFs remained largely unchanged during the weathering process. While the TLM-derived calculations displayed a significant decrease in toxicity (TUs) for the heavily weathered oils, copepod toxicity, expressed as LC10-based TUs, were comparable between LE-WAFs of fresh and weathered oils. The discrepancy between observed and predicted toxicity for the LE-WAFs of artificially weathered oils may be related to limitations by the chemical analyses or increased toxicity due to generation of new unknown compounds during the weathering process.</p

Does Microbial Biodegradation of Water-Soluble Components of Oil Reduce the Toxicity to Early Life Stages of Fish?

Microbial degradation following oil spills results in metabolites from the original oil. Metabolites are expected to display lower bioaccumulation potential and acute toxicity to marine organisms due to microbial-facilitated incorporation of chemical functional groups and a general decrease in lipophilicity. The toxicity and characterization of metabolites are poorly studied. The purpose of the present work was to evaluate the toxicity of degraded (0–21 days) water-soluble oil components. Low-energy water accommodated fraction (LE-WAF) of a weathered crude oil was prepared with nutrient amended seawater at 5 °C, kept in the dark, and sampled at 0, 10, 14, and 21 days. Samples were extracted with dichloromethane and toxicity experiments were conducted with reconstituted extracts. Toxicity experiments were conducted for 4 days on developing cod (Gadus morhua) embryos during a critical period of their heart development. After exposure, embryos were kept in clean seawater and observed until 5 days post hatch. Survival, hatching, morphometric aberrations, and cardiac function was studied. The expected decrease in sublethal toxicity during the biodegradation period was not found, indicating that metabolites formed during biodegradation likely contributed to larvae toxicity

Maternal polycyclic aromatic hydrocarbon (PAH) transfer and effects on offspring of copepods exposed to dispersed oil with and without oil droplets

<p>Copepods of the genus <i>Calanus</i> have the potential for accumulating lipophilic oil components due to their high lipid content and found to filter and ingest oil droplets during exposure. As female copepods produce eggs at the expense of lipid storage, there is a concern for transfer of lipophilic contaminants to offspring. To assess the potential for maternal transfer of oil components, ovigerous female copepods (<i>Calanus finmarchicus</i>) were exposed to filtered and unfiltered oil dispersions for 4 days, collected and eggs maintained in clean seawater and hatching and gene expression examined in hatched nauplii. Oil droplet exposure contributed to polycyclic aromatic hydrocarbon (PAH) uptake in dispersion-treated adult copepods, as displayed through PAH body residue analyses and fluorescence microscopy. Applying the latter methodology, transfer of heavy PAH from copepod mothers to offspring were detected Subtle effects were observed in offspring as evidenced by a temporal reduction in hatching success appear to be occurring only when mothers were exposed to the unfiltered oil dispersions. Offspring reared in clean water through to late naupliar stages were collected for RNA extraction and preparation of libraries for high-throughput transcriptome sequencing. Differentially expressed genes were identified through pairwise comparisons between treatments. Among these, several expressed genes have known roles in responses to chemical stress including xenobiotic metabolism enzymes, antioxidants, chaperones, and components of the inflammatory response. While gene expression results suggest a transgenerational activation of stress responses, the increase in relatively small number of differentially expressed genes suggests a minor long-term effect on offspring following maternal exposure.</p

A simple protocol for estimating the acute toxicity of unresolved polar compounds from field-weathered oils

Crude oil spilled at sea is chemically altered through environmental processes such as dissolution, biodegradation, and photodegradation. Transformation of hydrocarbons to oxygenated species increases water-solubility. Metabolites and oxidation products largely remain uncharacterized by common analytical methods but may be more bioavailable to aquatic organisms. Studies have shown that unresolved (i.e. unidentified) polar compounds (‘UPCs’) may constitute > 90% of the water-accommodated fraction (WAF) of heavily weathered crude oils, but still there is a paucity of information characterizing their toxicological significance in relation to other oil-derived toxicants. In this study, low-energy WAFs (no droplets) were generated from two field-weathered oils (collected during the 2010 Deepwater Horizon incident) and their polar fractions were isolated through fractionation. To allow establishment of thresholds for acute toxicity (LC50) of the dissolved and polar fraction of field collected oils, we concentrated both WAFs and polar fractions to beyond field-documented concentrations, and the acute toxicity of both to the marine copepod Acartia tonsa was measured and compared to the toxicity of the native WAF (non-concentrated). The difference in toxic units (TUs) between the total of the mixture and of identified compounds of known toxicity (polycyclic aromatic hydrocarbons [PAHs] and alkyl phenols) in both WAF and polar fractions was used to estimate the contribution of the UPC to overall toxicity. This approach identified that UPC had a similar contribution to toxicity as identified compounds within the WAFs of the field-weathered oils. This signifies the relative importance of polar compounds when assessing environmental impacts of spilled and weathered oil.</p

Acute and physical effects of water-based drilling mud in the marine copepod <i>Calanus finmarchicus</i>

<p>The aim of this study was to investigate impacts of fine particulate fraction of a commonly used barite-containing drilling mud on the pelagic filter feeding copepod <i>Calanus finmarchicus</i>. The results show that the tested drilling mud had a low acute toxicity on C. finmarchicus (LC50 > 320 mg/L) and that the observed toxicity was likely caused by dissolved constituents in the mud and not the particle phase containing the weighting agent barite. Further, animals were exposed to drilling mud at a concentration of 10 mg/L for 168 hr followed by a 100 hr recovery phase. A rapid uptake of drilling mud particles was observed, while the excretion was slow and incomplete even after 100 hr recovery in clean seawater. The uptake of drilling mud particles caused a significant increase in sinking velocity of copepods, indicating that uptake of drilling mud particles affected their buoyancy. Long-term exposure to low concentrations of drilling mud could therefore cause physical effects such as impacts on the animal's buoyancy which may affect the energy budget of the copepods.</p

Exposure of first-feeding cod larvae to dispersed crude oil results in similar transcriptional and metabolic responses as food deprivation

<p>Exposure of first-feeding cod larvae (<i>Gadus morhua</i>) to dispersed oil results in reduced feeding during an important transition period. First-feeding cod larvae were subjected to a 4-d treatment of food deprivation and sampled for microarray analyses. These microarray data were combined with data from cod larvae treated with mechanically and chemically dispersed oil in an attempt to understand to what extent starvation might explain some of the effects observed in first-feeding cod larvae during oil exposure. Transcriptional profiling of cod larvae suggested that the influence of oil exposure was almost as dramatic as being completely deprived of food. Protein and cellular degradation and loss of amino acids and glucose appear to be concomitant responses to both oil exposure and starvation. Fluorescence imaging of gut content indicated low uptake of food, and reduced growth (decrease in dry weight and in carbon and nitrogen content) was also noted in oil-exposed larvae, providing phenotypic anchoring of microarray data. The study displays the importance in combining use of high-throughput molecular tools with assessment of fitness-related endpoints in order to provide a greater understanding of toxicant-induced responses. This combined-approach investigation suggests that reduction of food uptake is an important process to be included when predicting effects of accidental oil spills. Finally, when comparing data from two oil treatments, exposure to chemically dispersed oil did not appear to result in greater toxicity than exposure to mechanically dispersed oil.</p

Mortality and transcriptional effects of inorganic mercury in the marine copepod <i>Calanus finmarchicus</i>

<p>Inorganic mercury (Hg) is highly toxic to organisms including crustaceans and displays multiple toxic modes of action (MoA). The main aim of this investigation was to assess the acute and sublethal toxicity mediated by mercury chloride (HgCl₂) in the marine copepod <i>Calanus finmarchicus</i>. A combination of short-term static studies to determine acute toxicity and a transcriptional investigation to characterize the sublethal MoA of HgCl₂ were conducted with an in-house continuous culture of <i>C. finmarchicus</i>. Transcriptional changes were determined by a custom 6.6 k <i>C. finmarchicus</i> Agilent oligonucleotide microarray and quantitative RT-PCR analysis. Data demonstrate that HgCl₂ produced a concentration- and time-dependent reduction in survival (NOEC_{48 h} = 6.9 μg/L [Hg²⁺] and LC₅₀ of 279, 73, 48, and 34 µg/L [Hg²⁺] after 24, 48, 72, and 96 h, respectively) and that exposure to sublethal concentrations of HgCl₂ (5 μg/L [Hg2⁺]) induced differential expression of 98 features (probes) on the microarray. Gene ontology (GO) and toxicological pathway analyses suggested that the main MOA were (1) uncoupling of mitochondrial oxidative phosphorylation (OXPHOS) and ATP production, (2) oxidative stress and macromolecular damage, (3) inactivation of cellular enzymes, (4) induction of cellular apoptosis and autophagocytosis, (5) over-excitation of glutamate receptors (neurotoxicity), (6) disruption of calcium homeostasis and signaling, and (7) modulation of nuclear receptor activity involved in vitamin D receptor signaling. Quantitative RT-PCR analysis verified that oligoarray performed reliably in terms of specificity and response, thus demonstrating that Hg²⁺ exerts multiple potential MoA in <i>C. finmarchicus</i>.</p

Toxicokinetics of Crude Oil Components in Arctic Copepods

The risk of accidental oil spills in the Arctic is on the rise due to increased shipping and oil exploration activities, making it essential to calibrate parameters for risk assessment of oil spills to Arctic conditions. The toxicokinetics of crude oil components were assessed by exposing one lipid-poor (CIII) and one lipid-rich (CV) stage of the Arctic copepod <i>Calanus hyperboreus</i> to crude oil WSF (water-soluble fraction). Water concentrations and total body residues (BR), as well as lipid volume fractions, were measured at regular intervals during exposure and recovery. Bioconcentration factors (BCFs) and elimination rates (<i>k</i>_e) for 26 petrogenic oil components were estimated from one-compartment models fitted to the BR data. Our parameters were compared to estimations made by the OMEGA bioaccumulation model, which uses the octanol–water partitioning coefficient (<i>K</i>_OW) in QSAR (quantitative structure–activity relationship) predictions. Our parameters for the lipid-poor CIIIs generally agreed with the OMEGA predictions, while neither the BCFs nor the <i>k</i>_es for the lipid-rich CVs fitted within the realistic range of the OMEGA parameters. Both the uptake and elimination rates for the CVs were in general half an order of magnitude lower than the OMEGA predictions, showing an overestimation of these parameters by the OMEGA model

Individual and molecular level effects of produced water contaminants on nauplii and adult females of <i>Calanus finmarchicus</i>

<p>In the Barents Sea region new petroleum fields are discovered yearly and extraction of petroleum products is expected to increase in the upcoming years. Despite enhanced technology and stricter governmental legislation, establishment of the petroleum industry in the Barents Sea may potentially introduce a new source of contamination to the area, as some discharges of produced water will be allowed. Whether the presence of produced water poses a risk to the Arctic marine life remains to be investigated. The aim of this study was to examine effects of exposure to several compounds found in produced water—a mixture of selected organic compounds (APW), radium-226 (²²⁶Ra), barium (Ba), and a scale inhibitor—on the copepod species <i>Calanus finmarchicus</i>. Experiments were performed using exposure concentrations at realistic levels based on those detected in the vicinity of known discharge points. The influence of lethal and sublethal effects on early life stages was determined and significantly lower survival in the APW exposure groups was found. In the Ba treatment the life stage development did not proceed to the same advanced stages as observed in the control (filtered sea water). The scale inhibitor and ²²⁶Ra treatments showed no significant difference from control. In addition, adult females were exposed to APW, ²²⁶Ra, and a mixture of the two. Both individual-level effects (egg production and feeding) and molecular-level effects (gene expression) were assessed. On the individual level endpoints, only treatments including APW produced an effect compared to control. However, on the molecular level the possibility that also ²²⁶Ra induced toxicologically relevant effects cannot be ruled out.</p