Effects of airgun discharges used in seismic surveys on development and mortality in nauplii of the copepod Acartia tonsa.

Seismic surveys are conducted worldwide to explore for oil and gas deposits and to map subsea formations. The airguns used in these surveys emit low-frequency sound waves. Studies on zooplankton responses to airguns report a range of effects, from none to substantial mortality. A field experiment was conducted to assess mortality and naupliar body length of the calanoid copepod Acartia tonsa when exposed to the discharge of two 40-inch airguns. Nauplii were placed in plastic bags and attached to a line at a depth of 6 m. For each treatment, three bags of nauplii were exposed to one of three treatments for 2.5 h: Airgun array discharge, a boat control, or a silent control. After exposure, nauplii were kept in filtered seawater in the laboratory without food. Immediate mortality in the nauplii was approximately 14% compared to less than 4% in the silent and boat control. Similarly, there was higher mortality in the airgun exposed nauplii up to six days after exposure compared to the control treatments. Nearly all of the airgun exposed nauplii were dead after four days, while >50% of the nauplii in the control treatments were alive at six days post-exposure. There was an interaction between treatment and time on naupliar body length, indicating lower growth in the nauplii exposed to the airgun discharge (growth rates after 4 days: 1.7, 5.4, and 6.1 µm d-1 in the airgun exposed, silent control, and boat control, respectively). These experiments indicate that the output of two small airguns affected mortality and growth of the naupliar stages of Acartia tonsa in close vicinity to the array.

Applying genetic technologies to combat infectious diseases in aquaculture.

Disease and parasitism cause major welfare, environmental and economic concerns for global aquaculture. In this review, we examine the status and potential of technologies that exploit genetic variation in host resistance to tackle this problem. We argue that there is an urgent need to improve understanding of the genetic mechanisms involved, leading to the development of tools that can be applied to boost host resistance and reduce the disease burden. We draw on two pressing global disease problems as case studies-sea lice infestations in salmonids and white spot syndrome in shrimp. We review how the latest genetic technologies can be capitalised upon to determine the mechanisms underlying inter- and intra-species variation in pathogen/parasite resistance, and how the derived knowledge could be applied to boost disease resistance using selective breeding, gene editing and/or with targeted feed treatments and vaccines. Gene editing brings novel opportunities, but also implementation and dissemination challenges, and necessitates new protocols to integrate the technology into aquaculture breeding programmes. There is also an ongoing need to minimise risks of disease agents evolving to overcome genetic improvements to host resistance, and insights from epidemiological and evolutionary models of pathogen infestation in wild and cultured host populations are explored. Ethical issues around the different approaches for achieving genetic resistance are discussed. Application of genetic technologies and approaches has potential to improve fundamental knowledge of mechanisms affecting genetic resistance and provide effective pathways for implementation that could lead to more resistant aquaculture stocks, transforming global aquaculture.

Changes in Chemical Composition and Copepod Toxicity during Petroleum Photo-oxidation.

Photoproducts can be formed rapidly in the initial phase of a marine oil spill. However, their toxicity is not well understood. In this study, oil was irradiated, chemically characterized, and tested for toxicity in three copepod species (Acartia tonsa, Temora longicornis, and Calanus finmarchicus). Irradiation led to a depletion of polycyclic aromatic hydrocarbons (PAHs) and n-alkanes in oil residues, along with an enrichment in aromatic and aliphatic oil photoproducts. Target lipid model-based calculations of PAH toxicity units predicted that PAH toxicities were lower in water-accommodated fractions (WAFs) of irradiated oil residues ("irradiated WAFs") than in WAFs of dark-control samples ("dark WAFs"). In contrast, biomimetic extraction (BE) measurements showed increased bioaccumulation potential of dissolved constituents of irradiated WAFs compared to dark WAFs, mainly driven by photoproducts present in irradiated oil. In line with the BE results, copepod mortality increased in irradiated WAFs compared to dark WAFs. However, low copepod toxicities were observed for WAFs produced with photo-oxidized oil slicks collected during the Deepwater Horizon oil spill. The results of this study suggest that while oil photoproducts have the potential to be a significant source of copepod toxicity, dilution and dispersion of these higher solubility products appear to help mitigate their toxicity at sea.

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.

American lobster postlarvae alter gene regulation in response to ocean warming and acidification.

Anthropogenic carbon emissions released into the atmosphere is driving rapid, concurrent increases in temperature and acidity across the world's oceans. Disentangling the interactive effects of warming and acidification on vulnerable life stages is important to our understanding of responses of marine species to climate change. This study evaluates the interactive effects of these stressors on the acute response of gene expression of postlarval American lobster (Homarus americanus), a species whose geographic range is warming and acidifying faster than most of the world's oceans. In the context of our experiment, we found two especially noteworthy results: First, although physiological end points have consistently been shown to be more responsive to warming in similar experimental designs, our study found gene regulation to be considerably more responsive to elevated pCO2. Furthermore, the combined effect of both stressors on gene regulation was significantly greater than either stressor alone. Using a full factorial experimental design, lobsters were raised in control and elevated pCO2 concentrations (400 ppm and 1,200 ppm) and temperatures (16°C and 19°C). A transcriptome was assembled from an identified 414,517 unique transcripts. Overall, 1,108 transcripts were differentially expressed across treatments, several of which were related to stress response and shell formation. When temperature alone was elevated (19°C), larvae downregulated genes related to cuticle development; when pCO2 alone was elevated (1,200 ppm), larvae upregulated chitinase as well as genes related to stress response and immune function. The joint effects of end-century stressors (19°C, 1,200 ppm) resulted in the upregulation of those same genes, as well as cellulase, the downregulation of calcified cuticle proteins, and a greater upregulation of genes related to immune response and function. These results indicate that changes in gene expression in larval lobster provide a mechanism to respond to stressors resulting from a rapidly changing environment.

Response of the copepod Acartia tonsa to the hydrodynamic cues of small-scale, dissipative eddies in turbulence.

This study quantifies the behavioral response of a marine copepod (Acartia tonsa) to individual, small-scale, dissipative vortices that are ubiquitous in turbulence. Vortex structures were created in the laboratory using a physical model of a Burgers vortex with characteristics corresponding to typical dissipative vortices that copepods are likely to encounter in the turbulent cascade. To examine the directional response of copepods, vortices were generated with the vortex axis aligned in either the horizontal or vertical direction. Tomographic particle image velocimetry was used to measure the volumetric velocity field of the vortex. Three-dimensional copepod trajectories were digitally reconstructed and overlaid on the vortex flow field to quantify A. tonsa's swimming kinematics relative to the velocity field and to provide insight into the copepod behavioral response to hydrodynamic cues. The data show significant changes in swimming kinematics and an increase in relative swimming velocity and hop frequency with increasing vortex strength. Furthermore, in moderate-to-strong vortices, A. tonsa moved at elevated speed in the same direction as the swirling flow and followed spiral trajectories around the vortex, which would retain the copepod within the feature and increase encounter rates with other similarly behaving Acartia While changes in swimming kinematics depended on vortex intensity, orientation of the vortex axis showed minimal significant effect. Hop and escape jump densities were largest in the vortex core, which is spatially coincident with the peak in vorticity, suggesting that vorticity is the hydrodynamic cue that evokes these behaviors.

Separating Thermal and Viscous Effects of Temperature on Copepod Respiration and Energy Budget.

AbstractChanges in temperature alter the viscosity of fluids, which impacts the force needed to move and the diffusion rates of gases. This is particularly salient for organisms that operate at mid to low Reynolds numbers. In this study, we investigated the independent effects of changes in temperature and viscosity on oxygen consumption rates of two coastal copepods (Acartia tonsa and Parvocalanus crassirostris) and used bioenergetic models to predict how these patterns could influence copepods in the natural environment. We found that only temperature influenced copepod oxygen consumption rates, indicating that copepods were not impacted by reduced oxygen diffusivity or increased energetics of movement resulting from higher seawater viscosity. We developed energy budgets based on novel respiration experiments in conjunction with data from the literature and found that cold temperatures do not result in higher scope for growth, because decreased metabolic costs are offset by reduced feeding capability. Our energy budgets imply that observed copepod temperature ranges in natural waters match theoretical ranges of optimal net carbon assimilation. At cold temperatures, feeding on motile prey yielded higher net carbon assimilation compared to feeding on non-motile prey, implying that motile prey are more favorable and may be actively selected for at cold temperatures. Finally, our models predicted that A. tonsa had a higher maximum net carbon assimilation as a percentage of body mass, indicating that copepods that use a similar sink-and-wait feeding strategy may be better able to exploit ephemeral food sources compared to continuous-swimming copepods such as P. crassirostris.

Accumulation and effects of microplastic fibers in American lobster larvae (Homarus americanus).

The effects of microplastic fibers (MPF) on the survival, molting and oxygen consumption rates of larval (I-III) and post-larval (IV) stages of the American lobster, Homarus americanus, were quantified as a function of MPF concentration and food availability. Only the highest MPF concentration decreased early larval survival. MPF did not affect the timing or rate of molting across MPF treatments. While all larval and post-larval stages accumulated MPF under the cephalothorax carapace, stage II larvae and stage IV post-larvae showed the highest and lowest accumulation, respectively. MPF ingestion increased with larval stage and with MPF concentration; under starvation conditions, stage I larvae only ingested them at low MPF concentrations. Oxygen consumption rates were lower only in later larval stages when exposed to high MPF concentrations. Combined, our results indicate that MPF interactions and effects on American lobster larvae are dependent on larval stage, MPF concentration, and presence of food.

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.

Physiological responses of Oxyrrhis marina to a diet of virally infected Emiliania huxleyi.

The coccolithophore Emiliania huxleyi forms some of the largest phytoplankton blooms in the ocean. The rapid demise of these blooms has been linked to viral infections. E. huxleyi abundance, distribution, and nutritional status make them an important food source for the heterotrophic protists which are classified as microzooplankton in marine food webs. In this study we investigated the fate of E. huxleyi (CCMP 374) infected with virus strain EhV-86 in a simple predator-prey interaction. The ingestion rates of Oxyrrhis marina were significantly lower (between 26.9 and 50.4%) when fed virus-infected E. huxleyi cells compared to non-infected cells. Despite the lower ingestion rates, O. marina showed significantly higher growth rates (between 30 and 91.3%) when fed infected E. huxleyi cells, suggesting higher nutritional value and/or greater assimilation of infected E. huxleyi cells. No significant differences were found in O. marina cell volumes or fatty acids profiles. These results show that virally infected E. huxleyi support higher growth rates of single celled heterotrophs and in addition to the "viral shunt" hypothesis, viral infections may also divert more carbon to mesozooplankton grazers.

Silencing of ionotropic receptor 25a decreases chemosensory activity in the salmon louse Lepeophtheirus salmonis during the infective stage.

Chemoreception is critical for marine ectoparasites - such as salmon lice (Lepeophtheirus salmonis) - to identify and locate salmonid hosts. The molecular receptors that parasites employ to detect host-specific chemical stimuli from hosts (kairomones) have not been well characterised. In the present study, transcription of the sea louse Ionotropic receptor 25a (IR25a) was blocked to evaluate whether it functions as a chemical-perception related gene for a specific chemical cue from the Atlantic salmon host. Double-strand RNA interference (dsRNA) oligonucleotides were applied to salmon lice by in vitro transcription and then exposing salmon lice nauplii to dsRNA by soaking overnight. Silencing of the IR25a gene was confirmed by qPCR in experimental groups of knock-down copepodids (dsIR25a). Behavioural responses associated with host recognition were evaluated in dsIR25a sea lice after exposure to a peptide produced by the salmon host (Cath-2). The dsIR25a group decreased expression levels of IR25a by >7-fold with respect to the control group. This group was also 26% slower than the control group (control swimming speed was 69 mm/s, while the treated group was 51 mm/s). Since the swimming activity of salmon lice copepodids is associated with the activation of the chemosensory system, these results indicate that the L. salmonis chemosensory perception system was not fully activated due to gene silencing. The results of this study demonstrate the role of ionotropic receptor 25a during host recognition by sea lice.

Microplastic fiber uptake, ingestion, and egestion rates in the blue mussel (Mytilus edulis).

Microplastic fibers (MPF) are a ubiquitous marine contaminant, making up to 90% of global microplastic concentrations. Imaging flow cytometry was used to measure uptake and ingestion rates of MPF by blue mussels (Mytilus edulis). Mussels were fed a diet of Rhodomonas salina and MPF concentrations up to 30 MPF mL-1, or 0.374% of available seston. Filtration rates were greatly reduced in mussels exposed to MPF. Uptake of MPF followed a Holling's Type II functional response with 95% of the maximum rate (5227 MPF h-1) occurring at 13 MPF mL-1. An average of 39 MPF (SE ±â€¯15, n = 4) was found in feces (maximum of 70 MPF). Most MPF (71%) were quickly rejected as pseudofeces, with approximately 9% ingested and <1% excreted in feces. Mussels may act as microplastic sinks in Gulf of Maine coastal waters, where MPF concentrations are near the order of magnitude as the experimental treatments herein.

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.

Coccolith dissolution within copepod guts affects fecal pellet density and sinking rate.

The most common biomineral produced in the contemporary ocean is calcium carbonate, including the polymorph calcite produced by coccolithophores. The surface waters of the ocean are supersaturated with respect to calcium carbonate. As a result, particulate inorganic carbon (PIC), such as calcite coccoliths, is not expected thermodynamically to dissolve in waters above the lysocline (~4500-6000 m). However, observations indicate that up to 60-80% of calcium carbonate is lost in the upper 500-1000 m of the ocean. This is hypothesized to occur in microenvironments with reduced saturation states, such as zooplankton guts. Using a new application of the highly precise 14C microdiffusion technique, we show that following a period of starvation, up to 38% of ingested calcite dissolves in copepod guts. After continued feeding, our data show the gut becomes increasingly buffered, which limits further dissolution; this has been termed the Tums hypothesis (after the drugstore remedy for stomach acid). As less calcite dissolves in the gut and is instead egested in fecal pellets, the fecal pellet sinking rates double, with corresponding increases in pellet density. Our results empirically demonstrate that zooplankton guts can facilitate calcite dissolution above the chemical lysocline, and that carbon export through fecal pellet production is variable, based on the feeding history of the copepod.

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.

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.

Grazing rates of Calanus finmarchicus on Thalassiosira weissflogii cultured under different levels of ultraviolet radiation.

UVB alters photosynthetic rate, fatty acid profiles and morphological characteristics of phytoplankton. Copepods, important grazers of primary production, select algal cells based upon their size, morphological traits, nutritional status, and motility. We investigated the grazing rates of the copepod Calanus finmarchicus on the diatom Thalassiosira weissflogii cultured under 3 levels of ultraviolet radiation (UVR): photosynthetically active radiation (PAR) only (4 kJ-m(-2)/day), and PAR supplemented with UVR radiation at two intensities (24 kJ-m(-2)/day and 48 kJ-m(-2)/day). There was no significant difference in grazing rates between the PAR only treatment and the lower UVR treatment. However, grazing rates were significantly (∼66%) higher for copepods feeding on cells treated with the higher level of UVR. These results suggest that a short-term increase in UVR exposure results in a significant increase in the grazing rate of copepods and, thereby, potentially alters the flow rate of organic matter through this component of the ecosystem.

The three-dimensional prey field of the northern krill, Meganyctiphanes norvegica, and the escape responses of their copepod prey.

In the north Atlantic, Meganyctiphanes norvegica feeds predominantly on copepods, including Calanus spp. To quantify its perceptual field for prey, and the sensory systems underlying prey detection, the responses of tethered krill to free-swimming Calanus spp. were observed in 3D using silhouette video imaging. An attack-which occurred despite the krill's being tethered-was characterized by a pronounced movement of the krill's antennae towards the target, followed by a propulsion and opening of the feeding basket. Frequency distributions of prey detection distances were significantly different in the light vs. the dark, with median values of 26.5 mm and 19.5 mm, respectively. There were no significant differences in the angles at which prey were detected by krill (relative to the predator's longitudinal body axis) in the light vs. the dark. Prey detections were symmetrically distributed on either side of the predator, in both light and dark. However, significant asymmetry was found in the dorsal-ventral direction with 80% of the prey detections located below the midline of the krill's body axis and, given the placement and orientation of the compound eyes, presumably outside its visual field of view. This indicates that, at least under these conditions, vision was not the main sensory modality involved in the detection of active prey by M. norvegica. However, under some circumstances, vision may provide supplemental information. Avoidance responses of copepod prey were nearly twice the velocity of their nominal background swimming speed (153 ± 48 and 85 ± 75 mm s-1, respectively), on average taking them 43 ± 16 mm away from the predator. This is far beyond the krill's perceptual range, suggesting that the escape reaction provides an effective deterrent to predation (although perhaps less so for free-swimming krill). This information can be used to parameterize models that assess the role of krill as predators in marine ecosystems.

Rapid firing rates from mechanosensory neurons in copepod antennules.

Small detection distances coupled with rapid movements require copepods to respond to stimuli with behavioral latencies on the order of milliseconds. Receiving adequate sensory information in such a short time necessitates extremely rapid firing rates of the efferent receptors. Here we show that copepod mechanoreceptors can fire at frequencies up to 5 kHz in response to fluid mechanical stimuli. Neural activity at these frequencies enables these animals to code for a range of fluid velocities thus providing important information regarding the nature of different fluid disturbances.