Severe Toxic Effects on Pelagic Copepods from Maritime Exhaust Gas Scrubber Effluents.

To reduce sulfur emission from global shipping, exhaust gas cleaning systems are increasingly being installed on board commercial ships. These so-called scrubbers extract SOX by spraying water into the exhaust gas. An effluent is created which is either released directly to the sea (open-loop system) or treated to remove harmful substances before release (closed-loop system). We found severe toxic effects in the ubiquitous planktonic copepod Calanus helgolandicus of exposure to effluents from two closed-loop systems and one open-loop system on North Sea ships. The effluents contained high concentrations of heavy metals and polycyclic aromatic hydrocarbons (PAHs), including alkylated PAHs. We observed significantly elevated mortality rates and impaired molting already in the lowest tested concentrations of each effluent: 0.04 and 0.1% closed-loop effluents and 1% open-loop effluent. These concentrations correspond to total hydrocarbon concentrations of 2.8, 2.0, and 3.8 µg L-1, respectively, and compared to previous studies on oil toxicity in copepods, scrubber effluents appear more toxic than, for example, crude oil. None of the individual PAHs or heavy metals analyzed in the effluents occurred in concentrations which could explain the high toxicity. The effluents showed unexpected alkylated PAH profiles, and we hypothesize that scrubbers act as witch's cauldrons where undesired toxic compounds form so that the high toxicity stems from compounds we know very little about.

Contrasting physiological responses to future ocean acidification among Arctic copepod populations.

Widespread ocean acidification (OA) is modifying the chemistry of the global ocean, and the Arctic is recognized as the region where the changes will progress at the fastest rate. Moreover, Arctic species show lower capacity for cellular homeostasis and acid-base regulation rendering them particularly vulnerable to OA. In the present study, we found physiological differences in OA response across geographically separated populations of the keystone Arctic copepod Calanus glacialis. In copepodites stage CIV, measured reaction norms of ingestion rate and metabolic rate showed severe reductions in ingestion and increased metabolic expenses in two populations from Svalbard (Kongsfjord and Billefjord) whereas no effects were observed in a population from the Disko Bay, West Greenland. At pHT 7.87, which has been predicted for the Svalbard west coast by year 2100, these changes resulted in reductions in scope for growth of 19% in the Kongsfjord and a staggering 50% in the Billefjord. Interestingly, these effects were not observed in stage CV copepodites from any of the three locations. It seems that CVs may be more tolerant to OA perhaps due to a general physiological reorganization to meet low intracellular pH during hibernation. Needless to say, the observed changes in the CIV stage will have serious implications for the C. glacialis population health status and growth around Svalbard. However, OA tolerant populations such as the one in the Disko Bay could help to alleviate severe effects in C. glacialis as a species.

Transgenerational effects alleviate severe fecundity loss during ocean acidification in a ubiquitous planktonic copepod.

Ocean acidification (OA) caused by anthropogenic CO2 emission is projected for thousands of years to come, and significant effects are predicted for many marine organisms. While significant evolutionary responses are expected during such persistent environmental change, most studies consider only short-term effects. Little is known about the transgenerational effects of parental environments or natural selection on the capacity of populations to counter detrimental OA effects. In this study, six laboratory populations of the calanoid copepod Pseudocalanus acuspes were established at three different CO2 partial pressures (pCO2 of 400, 900 and 1550 µatm) and grown for two generations at these conditions. Our results show evidence of alleviation of OA effects as a result of transgenerational effects in P. acuspes. Second generation adults showed a 29% decrease in fecundity at 900 µatm CO2 compared to 400 µatm CO2 . This was accompanied by a 10% increase in metabolic rate indicative of metabolic stress. Reciprocal transplant tests demonstrated that this effect was reversible and the expression of phenotypic plasticity. Furthermore, these tests showed that at a pCO2 exceeding the natural range experienced by P. acuspes (1550 µatm), fecundity would have decreased by as much as 67% compared to at 400 µatm CO2 as a result of this plasticity. However, transgenerational effects partly reduced OA effects so that the loss of fecundity remained at a level comparable to that at 900 µatm CO2 . This also relieved the copepods from metabolic stress, and respiration rates were lower than at 900 µatm CO2 . These results highlight the importance of tests for transgenerational effects to avoid overestimation of the effects of OA.

Ocean acidification causes fundamental changes in the cellular metabolism of the Arctic copepod Calanus glacialis as detected by metabolomic analysis.

Using a targeted metabolomic approach we investigated the effects of low seawater pH on energy metabolism in two late copepodite stages (CIV and CV) of the keystone Arctic copepod species Calanus glacialis. Exposure to decreasing seawater pH (from 8.0 to 7.0) caused increased ATP, ADP and NAD+ and decreased AMP concentrations in stage CIV, and increased ATP and phospho-L-arginine and decreased AMP concentrations in stage CV. Metabolic pathway enrichment analysis showed enrichment of the TCA cycle and a range of amino acid metabolic pathways in both stages. Concentrations of lactate, malate, fumarate and alpha-ketoglutarate (all involved in the TCA cycle) increased in stage CIV, whereas only alpha-ketoglutarate increased in stage CV. Based on the pattern of concentration changes in glucose, pyruvate, TCA cycle metabolites, and free amino acids, we hypothesise that ocean acidification will lead to a shift in energy production from carbohydrate metabolism in the glycolysis toward amino acid metabolism in the TCA cycle and oxidative phosphorylation in stage CIV. In stage CV, concentrations of most of the analysed free fatty acids increased, suggesting in particular that ocean acidification increases the metabolism of stored wax esters in this stage. Moreover, aminoacyl-tRNA biosynthesis was enriched in both stages indicating increased enzyme production to handle low pH stress.

No maternal or direct effects of ocean acidification on egg hatching in the Arctic copepod Calanus glacialis.

Widespread ocean acidification (OA) is transforming the chemistry of the global ocean and the Arctic is recognised as the region where this transformation will occur at the fastest rate. Moreover, many Arctic species are considered less capable of tolerating OA due to their lower capacity for acid-base regulation. This inability may put severe restraints on many fundamental functions, such as growth and reproductive investments, which ultimately may result in reduced fitness. However, maternal effects may alleviate severe effects on the offspring rendering them more tolerant to OA. In a highly replicated experiment we studied maternal and direct effects of OA predicted for the Arctic shelf seas on egg hatching time and success in the keystone copepod species Calanus glacialis. We incubated females at present day conditions (pHT 8.0) and year 2100 extreme conditions (pHT 7.5) during oogenesis and subsequently reciprocally transplanted laid eggs between these two conditions. Statistical tests showed no effects of maternal or direct exposure to OA at this level. We hypothesise that C. glacialis may be physiologically adapted to egg production at low pH since oogenesis can also take place at conditions of potentially low haemolymph pH of the mother during hibernation in the deep.

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.

Ocean acidification effects on mesozooplankton community development: Results from a long-term mesocosm experiment.

Ocean acidification may affect zooplankton directly by decreasing in pH, as well as indirectly via trophic pathways, where changes in carbon availability or pH effects on primary producers may cascade up the food web thereby altering ecosystem functioning and community composition. Here, we present results from a mesocosm experiment carried out during 113 days in the Gullmar Fjord, Skagerrak coast of Sweden, studying plankton responses to predicted end-of-century pCO2 levels. We did not observe any pCO2 effect on the diversity of the mesozooplankton community, but a positive pCO2 effect on the total mesozooplankton abundance. Furthermore, we observed species-specific sensitivities to pCO2 in the two major groups in this experiment, copepods and hydromedusae. Also stage-specific pCO2 sensitivities were detected in copepods, with copepodites being the most responsive stage. Focusing on the most abundant species, Pseudocalanus acuspes, we observed that copepodites were significantly more abundant in the high-pCO2 treatment during most of the experiment, probably fuelled by phytoplankton community responses to high-pCO2 conditions. Physiological and reproductive output was analysed on P. acuspes females through two additional laboratory experiments, showing no pCO2 effect on females' condition nor on egg hatching. Overall, our results suggest that the Gullmar Fjord mesozooplankton community structure is not expected to change much under realistic end-of-century OA scenarios as used here. However, the positive pCO2 effect detected on mesozooplankton abundance could potentially affect biomass transfer to higher trophic levels in the future.

Copepods induce paralytic shellfish toxin production in marine dinoflagellates.

Among the thousands of unicellular phytoplankton species described in the sea, some frequently occurring and bloom-forming marine dinoflagellates are known to produce the potent neurotoxins causing paralytic shellfish poisoning. The natural function of these toxins is not clear, although they have been hypothesized to act as a chemical defence towards grazers. Here, we show that waterborne cues from the copepod Acartia tonsa induce paralytic shellfish toxin (PST) production in the harmful algal bloom-forming dinoflagellate Alexandrium minutum. Induced A. minutum contained up to 2.5 times more toxins than controls and was more resistant to further copepod grazing. Ingestion of non-toxic alternative prey was not affected by the presence of induced A. minutum. The ability of A. minutum to sense and respond to the presence of grazers by increased PST production and increased resistance to grazing may facilitate the formation of harmful algal blooms in the sea.

Will life find a way? Evolution of marine species under global change.

Projections of marine biodiversity and implementation of effective actions for its maintenance in the face of current rapid global environmental change are constrained by our limited understanding of species' adaptive responses, including transgenerational plasticity, epigenetics and natural selection. This special issue presents 13 novel studies, which employ experimental and modelling approaches to (i) investigate plastic and evolutionary responses of marine species to major global change drivers; (ii) ask relevant broad eco-evolutionary questions, implementing multiple species and populations studies; (iii) show the advantages of using advanced experimental designs and tools; (iv) construct novel model organisms for marine evolution; (v) help identifying future challenges for the field; and (vi) highlight the importance of incorporating existing evolutionary theory into management solutions for the marine realm. What emerges is that at least some populations of marine species have the ability to adapt to future global change conditions. However, marine organisms' capacity for adaptation appears finite, due to evolutionary trade-offs and possible rapid losses in genetic diversity. This further corroborates the idea that acquiring an evolutionary perspective on how marine life will respond to the selective pressure of future global changes will guide us in better identifying which conservation efforts will be most needed and most effective.

Selection on oxidative phosphorylation and ribosomal structure as a multigenerational response to ocean acidification in the common copepod Pseudocalanus acuspes.

Ocean acidification is expected to have dramatic impacts on oceanic ecosystems, yet surprisingly few studies currently examine long-term adaptive and plastic responses of marine invertebrates to pCO 2 stress. Here, we exposed populations of the common copepod Pseudocalanus acuspes to three pCO 2 regimes (400, 900, and 1550 µatm) for two generations, after which we conducted a reciprocal transplant experiment. A de novo transcriptome was assembled, annotated, and gene expression data revealed that genes involved in RNA transcription were strongly down-regulated in populations with long-term exposure to a high pCO 2 environment, even after transplantation back to control levels. In addition, 747 000 SNPs were identified, out of which 1513 showed consistent changes in nucleotide frequency between replicates of control and high pCO 2 populations. Functions involving RNA transcription and ribosomal function, as well as ion transport and oxidative phosphorylation, were highly overrepresented. We thus conclude that pCO 2 stress appears to impose selection in copepods on RNA synthesis and translation, possibly modulated by helicase expression. Using a physiological hypothesis-testing strategy to mine gene expression data, we herein increase the power to detect cellular targets of ocean acidification. This novel approach seems promising for future studies of effects of environmental changes in ecologically important nonmodel organisms.

Seawater pH Predicted for the Year 2100 Affects the Metabolic Response to Feeding in Copepodites of the Arctic Copepod Calanus glacialis.

Widespread ocean acidification (OA) is transforming the chemistry of the global ocean, and the Arctic is recognised as a region where the earliest and strongest impacts of OA are expected. In the present study, metabolic effects of OA and its interaction with food availability was investigated in Calanus glacialis from the Kongsfjord, West Spitsbergen. We measured metabolic rates and RNA/DNA ratios (an indicator of biosynthesis) concurrently in fed and unfed individuals of copepodite stages CII-CIII and CV subjected to two different pH levels representative of present day and the "business as usual" IPCC scenario (RCP8.5) prediction for the year 2100. The copepods responded more strongly to changes in food level than to decreasing pH, both with respect to metabolic rate and RNA/DNA ratio. However, significant interactions between effects of pH and food level showed that effects of pH and food level act in synergy in copepodites of C. glacialis. While metabolic rates in copepodites stage CII-CIII increased by 78% as a response to food under present day conditions (high pH), the increase was 195% in CII-CIIIs kept at low pH-a 2.5 times greater increase. This interaction was absent for RNA/DNA, so the increase in metabolic rates were clearly not a reaction to changing biosynthesis at low pH per se but rather a reaction to increased metabolic costs per unit of biosynthesis. Interestingly, we did not observe this difference in costs of growth in stage CV. A 2.5 times increase in metabolic costs of growth will leave the copepodites with much less energy for growth. This may infer significant changes to the C. glacialis population during future OA.

Effects of selected PAHs on reproduction and survival of the calanoid copepod Acartia tonsa.

The effects of selected polycyclic aromatic hydrocarbons on the marine calanoid copepod Acartia tonsa were tested in laboratory short-term toxicity tests in order to facilitate risk assessment of those compounds to the marine pelagic environment. Photo-induced toxicity of pyrene was also investigated under naturally relevant UV light regimes. Lethal and sublethal effects on egg production rate, hatching and potential recruitment rate were evaluated after 48 h exposure to fluoranthene, phenanthrene and pyrene. The 48 h-median lethal concentrations (LC(50)) reducing survival by 50% were 594, 2,366 and >640 nM for fluoranthene, phenanthrene and pyrene, respectively, whilst lower concentrations induced different sublethal effects. Median effective concentrations (EC(50)) affecting the egg production rate and the recruitment rate were 433 and 385 (fluoranthene), 1,245 and 1,012 (phenanthrene) and 306 and 295 nM (pyrene), respectively. An increase in toxicity of pyrene was detected after incubation under UV light, resulting in LC(50) values of 201 nM (24 h) and 138 nM (48 h) and EC(50) values of 79 nM (egg production rate) and 41 nM (recruitment rate). Finally, a comparison between effective concentrations and worst-case environmental concentrations reported in literature indicated that pyrene may pose a threat to A. tonsa from exposure in the field, and that the risk of adverse effects is high for fluoranthene.