Atlantic cod (Gadus morhua) larvae are attracted by low-frequency noise simulating that of operating offshore wind farms.

The number and size of offshore wind (OW) turbines is increasing rapidly. OW turbines produce continuous, low-frequency noise that could impact marine fish dispersing/migrating through the facilities. Any such impact would be relevant for larval stages, which have limited possibility to swim away from OW facilities. If directional movement of fish larvae at sea is impacted by low-frequency continuous sound is unknown. We observe the behavior of Atlantic cod larvae (N = 89) in response to low-frequency sound while they are drifting in a Norwegian fjord inside transparent drifting chambers. We transmit 100 Hz continuous sound in the fjord, in the intensity range of OW turbines' operational noise, and measure the sound pressure and 3-D particle motion. Half of the larvae (N = 45) are exposed to low-frequency (100 Hz) continuous sound, while the other half (N = 44) are observed under the same conditions but without the sound. Exposure does not affect the routine and maximum swimming speeds or the turning behavior of the larvae. Control larvae orient to the northwest. In contrast, exposed larvae orient towards the source of low-frequency sound and particle motion. This provides a basis to assess how OW might impact dispersal in this species.

Magnetic fields generated by submarine power cables have a negligible effect on the swimming behavior of Atlantic lumpfish (Cyclopterus lumpus) juveniles.

Submarine power cables carry electricity over long distances. Their geographic distribution, number, and areal coverage are increasing rapidly with the development of, for example, offshore wind facilities. The flow of current passing through these cables creates a magnetic field (MF) that can potentially affect marine organisms, particularly those that are magnetosensitive. The lumpfish (Cyclopterus lumpus) is a migratory species that is widely distributed in the North Atlantic Ocean and Barents Sea. It migrates between coastal spawning grounds and pelagic offshore feeding areas. We tested whether lumpfish respond to MFs of the same intensity as those emitted by high voltage direct current (HVDC) submarine power cables. Laboratory experiments were conducted by placing juvenile lumpfish in an artificial MF gradient generated by a Helmholtz coil system. The intensity of the artificial MF used (230 µT) corresponded to the field at 1 m from a high-power submarine cable. The fish were filmed for 30 min with the coil either on or off. Swimming speeds, and presence in the different parts of a raceway, were extracted from the videos and analyzed. Juvenile lumpfish activity, defined as the time that the fish spent swimming relative to stationary pauses (attached to the substrate), and the distance travelled, were unaffected by exposure to the artificial MF. The swimming speed of juvenile lumpfish was reduced (by 16%) when the coil was on indicating that the fish could either sense the MF or the induced electric field created by the movement of the fish through the magnetic field. However, it seems unlikely that a 16% decrease in swimming speed occurring within 1 m of HVDC cables would significantly affect Atlantic lumpfish migration or homing.

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.

Increasing temperature and prey availability affect the growth and swimming kinematics of Atlantic herring (Clupea harengus) larvae.

Climate change will increase the magnitude and duration of warming events and the variability in the phenology and abundance of available prey to the early life stages of fish. These factors influence physiological, behavioral and ecological processes, impacting growth, development and survival. Using a fully factorial design with two prey-availability treatments (1200 prey items L-1 (high prey abundance) or 40 prey items L-1 (low prey abundance)) under three temperature regimes (8, 10 and 12°C), the swimming kinematics of 6-week old spring-spawning Atlantic herring larvae were examined using silhouette video photography. Higher temperatures combined with food limitation significantly decreased the growth and swimming kinematics of larval herring, with the most negative effect observed in larvae reared at 12°C and exposed to low food abundances. Specifically, larvae displayed reduced locomotory behaviors and reduced vertical movements. By contrast, larvae reared at high prey abundance and at 12°C displayed more active swimming and exploratory behavior, as evidenced by an increase in both locomotory behavior and vertical and horizontal turn angles, suggesting increased motivation to search for food. This research highlights the importance of determining to what degree fish larvae are sensitive to changes in temperature and how these changes might be further influenced by food availability.

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.

Gene expression and epigenetic responses of the marine Cladoceran, Evadne nordmanni, and the copepod, Acartia clausi, to elevated CO2.

Characterizing the capacity of marine organisms to adapt to climate change related drivers (e.g., pCO2 and temperature), and the possible rate of this adaptation, is required to assess their resilience (or lack thereof) to these drivers. Several studies have hypothesized that epigenetic markers such as DNA methylation, histone modifications and noncoding RNAs, act as drivers of adaptation in marine organisms, especially corals. However, this hypothesis has not been tested in zooplankton, a keystone organism in marine food webs. The objective of this study is to test the hypothesis that acute ocean acidification (OA) exposure alters DNA methylation in two zooplanktonic species-copepods (Acartia clausii) and cladocerans (Evadne nordmanii). We exposed these two species to near-future OA conditions (400 and 900 ppm pCO2) for 24 h and assessed transcriptional and DNA methylation patterns using RNA sequencing and Reduced Representation Bisulfite Sequencing (RRBS). OA exposure caused differential expression of genes associated with energy metabolism, cytoskeletal and extracellular matrix functions, hypoxia and one-carbon metabolism. Similarly, OA exposure also caused altered DNA methylation patterns in both species but the effect of these changes on gene expression and physiological effects remains to be determined. The results from this study form the basis for studies investigating the potential role of epigenetic mechanisms in OA induced phenotypic plasticity and/or adaptive responses in zooplanktonic organisms.

Effects of Exposure to Low Concentrations of Oil on the Expression of Cytochrome P4501a and Routine Swimming Speed of Atlantic Haddock (Melanogrammus aeglefinus) Larvae In Situ.

Exposure to environmentally relevant concentrations of oil could impact survival of fish larvae in situ through subtle effects on larval behavior. During the larval period, Atlantic haddock (Melanogrammus aeglefinus) are transported toward nursery grounds by ocean currents and active swimming, which can modify their drift route. Haddock larvae are sensitive to dispersed oil; however, whether exposure to oil during development impacts the ability of haddock larvae to swim in situ is unknown. Here, we exposed Atlantic haddock embryos to 10 and 80 µg oil/L (0.1 and 0.8 µg ∑PAH/L) of crude oil for 8 days and used a novel approach to measure its effect on the larval swimming behavior in situ. We assessed the swimming behavior of 138 haddock larvae in situ, in the North Sea, using a transparent drifting chamber. Expression of cytochrome P4501a (cyp1a) was also measured. Exposure to 10 and 80 µg oil/L significantly reduced the average in situ routine swimming speed by 30-40% compared to the controls. Expression of cyp1a was significantly higher in both exposed groups. This study reports key information for improving oil spill risk assessment models and presents a novel approach to study sublethal effects of pollutants on fish larvae in situ.

Age of European silver eels during a period of declining abundance in Norway.

The European eel (Anguilla anguilla) is critically endangered throughout its range. Knowledge about age distribution of future spawners (silver eels) is essential to monitor the status and contribute to the recovery of this species. Determination of age in anguillid eels is challenging, especially in eels from the northern part of the distribution area where growth is slow and age at maturation can be up to 30 years or more. Eels from the river Imsa in Norway have been monitored since 1975, and this reference time series has been used to assess the stock at the European level. Population dynamics in this catchment were analyzed during the late 1980s by estimating ages on whole cleared otoliths. However, techniques for revealing annual increments on otoliths have evolved over the years sometimes yielding significant differences in age estimates. In this study, the historical otolith data were reanalyzed using a grinding and polishing method rather than reading the whole otolith. The new age estimates were considerably higher than the previous ones, sometimes by up to 29 years. Since the 1980s, mean age of silver eels only slightly increased (from 19 to 21 years in the 2010s). This was mainly due to the disappearance of younger silver eels (<15 years) in the 2010s. The new age estimates agreed with the steep decline in recruitment which occurred in the late 1980s in the Imsa catchment. Mean growth (30 mm/year, min-max: 16-64 mm/year) has not changed since the 1980s, although density in the catchment has decreased. Revealing and reading age of slow-growing eels remain a challenge but adding a measure of otolith reading uncertainty may improve age data collection and contribute to recovery measures for this species.

The relationship between the moon cycle and the orientation of glass eels (Anguilla anguilla) at sea.

Links between the lunar cycle and the life cycle (migration patterns, locomotor activity, pulses in recruitment) of the European eel (Anguilla anguilla) are well documented. In this study, we hypothesized that the orientation of glass eels at sea is related to the lunar cycle. The European eel hatches in the Sargasso Sea and migrates across the Atlantic Ocean towards Europe. Upon reaching the continental shelf, the larvae metamorphose into glass eels and migrate up the estuaries, where some individuals colonize freshwater habitats. How glass eels navigate pelagic waters is still an open question. We tested the orientation of 203 glass eels in a transparent circular arena that was drifting in situ during the daytime, in the coastal Norwegian North Sea, during different lunar phases. The glass eels swimming at sea oriented towards the azimuth of the moon at new moon, when the moon rose above the horizon and was invisible but not during the other moon phases. These results suggest that glass eels could use the moon position for orientation at sea and that the detection mechanism involved is not visual. We hypothesize a possible detection mechanism based on global-scale lunar disturbances in electrical fields and discuss the implications of lunar-related orientation for the recruitment of glass eels to estuaries. This behaviour could help glass eels to reach the European coasts during their marine migration.

The planktonic stages of the salmon louse (Lepeophtheirus salmonis) are tolerant of end-of-century pCO2 concentrations.

The copepod Lepeophtheirus salmonis is an obligate ectoparasite of salmonids. Salmon lice are major pests in salmon aquaculture and due to its economic impact Lepeophtheirus salmonis is one of the most well studied species of marine parasite. However, there is limited understanding of how increased concentration of pCO2 associated with ocean acidification will impact host-parasite relationships. We investigated the effects of increased pCO2 on growth and metabolic rates in the planktonic stages, rearing L. salmonis from eggs to 12 days post hatch copepodids under three treatment levels: Control (416 µatm), Mid (747 µatm), and High (942 µatm). The pCO2 treatment had a significant effect on oxygen consumption rate with the High treatment animals exhibiting the greatest respiration. The treatments did not have a significant effect on the other biological endpoints measured (carbon, nitrogen, lipid volume, and fatty acid content). The results indicate that L. salmonis have mechanisms to compensate for increased concentration of pCO2and that populations will be tolerant of projected future ocean acidification scenarios. The work reported here also describes catabolism during the lecithotrophic development of L. salmonis, information that is not currently available to parameterize models of dispersal and viability of the planktonic free-living stages.

Glass eels (Anguilla anguilla) imprint the magnetic direction of tidal currents from their juvenile estuaries.

The European eel (Anguilla anguilla) hatches in the Sargasso Sea and migrates to European and North African freshwater. As glass eels, they reach estuaries where they become pigmented. Glass eels use a tidal phase-dependent magnetic compass for orientation, but whether their magnetic direction is innate or imprinted during migration is unknown. We tested the hypothesis that glass eels imprint their tidal-dependent magnetic compass direction at the estuaries where they recruit. We collected 222 glass eels from estuaries flowing in different cardinal directions in Austevoll, Norway. We observed the orientation of the glass eels in a magnetic laboratory where the magnetic North was rotated. Glass eels oriented towards the magnetic direction of the prevailing tidal current occurring at their recruitment estuary. Glass eels use their magnetic compass to memorize the magnetic direction of tidal flows. This mechanism could help them to maintain their position in an estuary and to migrate upstream.

Low Abundance of Circulating Tumor DNA in Localized Prostate Cancer.

PURPOSE: Despite decreased screening-based detection of clinically insignificant tumors, most diagnosed prostate cancers are still indolent, indicating a need for better strategies for detection of clinically significant disease before treatment. We hypothesized that patients with detectable circulating tumor DNA (ctDNA) were more likely to harbor aggressive disease. METHODS: We applied ultra-low-pass whole-genome sequencing to profile cell-free DNA from 112 patients diagnosed with localized prostate cancer and performed targeted resequencing of plasma DNA for somatic mutations previously identified in matched solid tumor in nine cases. We also performed similar analyses of data from patients with metastatic prostate cancer. RESULTS: In all cases of localized prostate cancer, even in clinically high-risk patients who subsequently had recurrent disease, ultra-low-pass whole-genome sequencing and targeted resequencing did not detect ctDNA in plasma acquired before surgery or before recurrence. In contrast, using both approaches, ctDNA was detected in patients with metastatic prostate cancer. CONCLUSION: Our findings demonstrate clear differences between localized and advanced prostate cancer with respect to the dissemination and detectability of ctDNA. Because allele-specific alterations in ctDNA are below the threshold for detection in localized prostate cancer, other approaches to identify cell-free nucleic acids of tumor origin may demonstrate better specificity for aggressive disease.

Atlantic Haddock (Melanogrammus aeglefinus) Larvae Have a Magnetic Compass that Guides Their Orientation.

Atlantic haddock (Melanogrammus aeglefinus) is a commercially important species of gadoid fish. In the North Sea, their main spawning areas are located close to the northern continental slope. Eggs and larvae drift with the current across the North Sea. However, fish larvae of many taxa can orient at sea using multiple external cues, including the Earth's magnetic field. In this work, we investigated whether haddock larvae passively drift or orient using the Earth's magnetic field. We observed the behavior of 59 and 102 haddock larvae swimming in a behavioral chamber deployed in the Norwegian North Sea and in a magnetic laboratory, respectively. In both in situ and laboratory settings, where the magnetic field direction was modified, haddock larvae significantly oriented toward the northwest. We conclude that haddock larvae orientation at sea is guided by a magnetic compass mechanism. These results have implications for retention and dispersal of pelagic haddock larvae.

The Atlantic salmon (Salmo salar) antimicrobial peptide cathelicidin-2 is a molecular host-associated cue for the salmon louse (Lepeophtheirus salmonis).

Chemical signals are a key element of host-parasite interactions. In marine ecosystems, obligate ectoparasites, such as sea lice, use chemical cues and other sensory signals to increase the probability of encountering a host and to identify appropriate hosts on which they depend to complete their life cycle. The chemical compounds that underlie host identification by the sea lice are not fully described or characterized. Here, we report a novel compound - the Atlantic salmon (Salmo salar) antimicrobial peptide cathelicidin-2 (Cath-2) - that acts as an activation cue for the marine parasitic copepod Lepeophtheirus salmonis. L. salmonis were exposed to 0, 7, 70 and 700 ppb of Cath-2 and neural activity, swimming behaviour and gene expression profiles of animals in response to the peptide were evaluated. The neurophysiological, behavioural and transcriptomic results were consistent: L. salmonis detects Cath-2 as a water-soluble peptide released from the skin of salmon, triggering chemosensory neural activity associated with altered swimming behaviour of copepodids exposed to the peptide, and chemosensory-related genes were up-regulated in copepodids exposed to the peptide. L. salmonis are activated by Cath-2, indicating a tight link between this peptide and the salmon louse chemosensory system.

HOXA9 methylation and blood vessel invasion in FFPE tissues for prognostic stratification of stage I lung adenocarcinoma patients.

OBJECTIVES: Surgery with curative intent is the standard treatment for stage I lung adenocarcinoma. However, disease recurrence occurs in a third of patients. Prognostic biomarkers are needed to improve postoperative management. Here, we evaluate the utility of Homeobox A9 (HOXA9) promoter methylation, alone or in combination with Blood Vessel Invasion (BVI) assessment, for prognostic stratification of stage I lung adenocarcinoma patients. MATERIALS AND METHODS: We developed a Droplet Digital PCR (ddPCR) assay to measure HOXA9 promoter methylation in formalin-fixed paraffin-embedded (FFPE) biospecimens generated during routine pathology. The prognostic value of HOXA9 promoter methylation and BVI, alone and in combination, was evaluated by Kaplan-Meier survival and Cox regression analyses in a cohort of 177 stage I lung adenocarcinoma patients from the NCI-MD study. RESULTS: The ddPCR assay showed linearity, sensitivity and specificity for measuring HOXA9 promoter methylation down to 0.1% methylated DNA input. The HOXA9 promoter was methylated de novo in FFPE tumors (P < 0.0001). High methylation was independently associated with worse cancer-specific survival (Hazard Ratio [HR], 3.37; P = 0.0002) and identified high-risk stage IA and IB patients. Addition of this molecular marker improved a risk model comprised of clinical and pathologic parameters (age, gender, race, stage, and smoking history; nested likelihood ratio test; P = 0.0004) and increased the C-index from 0.60 (95% CI 0.51-0.69) to 0.68 (0.60-0.76). High methylation tumors displayed high frequency of TP53 mutations and other molecular characteristics associated with aggressiveness. BVI was independently associated with poor outcome (HR, 2.62; P = 0.054). A score that combined BVI with HOXA9 promoter methylation further stratified high-risk patients (trend P = 0.0001 comparing 0, 1 or 2 positive markers). CONCLUSIONS: ddPCR can be used to quantify HOXA9 promoter methylation in FFPE samples. Alone or combined with BVI in a prognostic classifier, HOXA9 promoter methylation could potentially inform the clinical management of patients with early-stage lung adenocarcinoma.

Genomic perturbations reveal distinct regulatory networks in intrahepatic cholangiocarcinoma.

Intrahepatic cholangiocarcinoma remains a highly heterogeneous malignancy that has eluded effective patient stratification to date. The extent to which such heterogeneity can be influenced by individual driver mutations remains to be evaluated. Here, we analyzed genomic (whole-exome sequencing, targeted exome sequencing) and epigenomic data from 496 patients and used the three most recurrently mutated genes to stratify patients (IDH, KRAS, TP53, "undetermined"). Using this molecular dissection approach, each subgroup was determined to possess unique mutational signature preferences, comutation profiles, and enriched pathways. High-throughput drug repositioning in seven patient-matched cell lines, chosen to reflect the genetic alterations specific for each patient group, confirmed in silico predictions of subgroup-specific vulnerabilities linked to enriched pathways. Intriguingly, patients lacking all three mutations ("undetermined") harbored the most extensive structural alterations, while isocitrate dehydrogenase mutant tumors displayed the most extensive DNA methylome dysregulation, consistent with previous findings. CONCLUSION: Stratification of intrahepatic cholangiocarcinoma patients based on occurrence of mutations in three classifier genes (IDH, KRAS, TP53) revealed unique oncogenic programs (mutational, structural, epimutational) that influence pharmacologic response in drug repositioning protocols; this genome dissection approach highlights the potential of individual mutations to induce extensive molecular heterogeneity and could facilitate advancement of therapeutic response in this dismal disease. (Hepatology 2018).

Regulation of gene expression is associated with tolerance of the Arctic copepod Calanus glacialis to CO2-acidified sea water.

Ocean acidification is the increase in seawater pCO 2 due to the uptake of atmospheric anthropogenic CO 2, with the largest changes predicted to occur in the Arctic seas. For some marine organisms, this change in pCO 2, and associated decrease in pH, represents a climate change-related stressor. In this study, we investigated the gene expression patterns of nauplii of the Arctic copepod Calanus glacialis cultured at low pH levels. We have previously shown that organismal-level performance (development, growth, respiration) of C. glacialis nauplii is unaffected by low pH. Here, we investigated the molecular-level response to lowered pH in order to elucidate the physiological processes involved in this tolerance. Nauplii from wild-caught C. glacialis were cultured at four pH levels (8.05, 7.9, 7.7, 7.5). At stage N6, mRNA was extracted and sequenced using RNA-seq. The physiological functionality of the proteins identified was categorized using Gene Ontology and KEGG pathways. We found that the expression of 151 contigs varied significantly with pH on a continuous scale (93% downregulated with decreasing pH). Gene set enrichment analysis revealed that, of the processes downregulated, many were components of the universal cellular stress response, including DNA repair, redox regulation, protein folding, and proteolysis. Sodium:proton antiporters were among the processes significantly upregulated, indicating that these ion pumps were involved in maintaining cellular pH homeostasis. C. glacialis significantly alters its gene expression at low pH, although they maintain normal larval development. Understanding what confers tolerance to some species will support our ability to predict the effects of future ocean acidification on marine organisms.

Glass eels (Anguilla anguilla) have a magnetic compass linked to the tidal cycle.

The European eel (Anguilla anguilla) has one of the longest migrations in the animal kingdom. It crosses the Atlantic Ocean twice during its life history, migrating between the spawning area in the Sargasso Sea and Europe, where it is widely distributed. The leptocephalus larvae drift with the Gulf Stream and other currents for more than a year and metamorphose into glass eels when they arrive on the continental shelf and move toward coastal areas. The mechanisms underlying glass eel orientation toward the coast and into freshwater systems are poorly known. However, anguillid eels, including the glass eel life stage, have a geomagnetic sense, suggesting the possibility that they use Earth's magnetic field to orient toward the coast. To test this hypothesis, we used a unique combination of laboratory tests and in situ behavioral observations conducted in a drifting circular arena. Most (98%) of the glass eels tested in the sea exhibited a preferred orientation that was related to the tidal cycle. Seventy-one percent of the same eels showed the same orientation during ebb tide when tested in the laboratory under a manipulated simulated magnetic field in the absence of any other cue. These results demonstrate that glass eels use a magnetic compass for orientation and suggest that this magnetic orientation system is linked to a circatidal rhythm.

Light primes the escape response of the calanoid copepod, Calanus finmarchicus.

The timing and magnitude of an escape reaction is often the determining factor governing a copepod's success at avoiding predation. Copepods initiate rapid and directed escapes in response to fluid signals created by predators; however little is known about how copepods modulate their behavior in response to additional sensory input. This study investigates the effect of light level on the escape behavior of Calanus finmarchicus. A siphon flow was used to generate a consistent fluid signal and the behavioral threshold and magnitude of the escape response was quantified in the dark and in the light. The results show that C. finmarchicus initiated their escape reaction further from the siphon and traveled with greater speed in the light than in the dark. However, no difference was found in the escape distance. These results suggest that copepods use information derived from multiple sensory inputs to modulate the sensitivity and strength of the escape in response to an increase risk of predation. Population and IBM models that predict optimal vertical distributions of copepods in response to visual predators need to consider changes in the copepod's behavioral thresholds when predicting predation risk within the water column.