Impact of ocean acidification and warming on respiration, heart rate and acid-base status of Mytilus edulis from the North Sea

Anthropogenic climate change confronts marine organisms with rapid trends of concomitant warming and CO2 induced ocean acidification. The survival and distribution of species partly depend on their ability to exploit their physiological plasticity during acclimatization. Therefore, in laboratory studies the effects of simulated future ocean acidification on thermal tolerance, energy metabolism and acid-base regulation capacity of the North Sea population of the blue mussel Mytilus edulis were examined. Following one month of pre-acclimation to 10 °C and control CO2 levels, mussels were exposed for two weeks to control and projected oceanic CO2 levels (390, 750 and 1120 µatm) before being subjected to a stepwise warming protocol between 10 °C and 31 °C (+ 3 °C each night). Oxygen consumption and heart rates, anaerobic metabolite levels and haemolymph acid-base status were determined at each temperature. CO2 exposure left oxygen consumption rate unchanged at acclimation temperature but caused a somewhat stronger increase during acute warming and thus mildly higher Q10-values than seen in controls. Interestingly, the thermally induced limitation of oxygen consumption rate set in earlier in normocapnic than in hypercapnic (1120 µatm CO2) mussels (25.2 °C vs. 28.8 °C), likely due to an onset of metabolic depression in the control group following warming. However, the temperature induced increase in heart rate became limited above 25 °C in both groups indicating an unchanged pejus temperature regardless of CO2 treatment. An upper critical temperature was reached above 28 °C in both treatments indicated by the accumulation of anaerobic metabolites in the mantle tissue, paralleled by a strong increase in haemolymph PCO2 at 31 °C. Ocean acidification caused a decrease in haemolymph pH. The extracellular acidosis remained largely uncompensated despite some bicarbonate accumulation. In all treatments animals developed a progressive warming-induced extracellular acidosis. A stronger pH drop at around 25 °C was followed by stagnating heart rates. However, normocapnic mussels enhanced bicarbonate accumulation at the critical limit, a strategy no longer available to hypercapnic mussels. In conclusion, CO2 has small effects on the response patterns of mussels to warming, leaving thermal thresholds largely unaffected. High resilience of adult North Sea mussels to future ocean acidification indicates that sensitivity to thermal stress is more relevant in shaping the response to future climate change.

Seawater carbonate chemistry and resource allocation and extracellular acid-base status in the sea urchin Strongylocentrotus droebachiensis during experiments, 2012

Anthropogenic CO2 emission will lead to an increase in seawater pCO2 of up to 80-100 Pa (800-1000 µatm) within this century and to an acidification of the oceans. Green sea urchins (Strongylocentrotus droebachiensis) occurring in Kattegat experience seasonal hypercapnic and hypoxic conditions already today. Thus, anthropogenic CO2 emissions will add up to existing values and will lead to even higher pCO2 values >200 Pa (>2000 µatm). To estimate the green sea urchins' potential to acclimate to acidified seawater, we calculated an energy budget and determined the extracellular acid base status of adult S. droebachiensis exposed to moderately (102 to 145 Pa, 1007 to 1431 µatm) and highly (284 to 385 Pa, 2800 to 3800 µatm) elevated seawater pCO2 for 10 and 45 days. A 45 - day exposure to elevated pCO2 resulted in a shift in energy budgets, leading to reduced somatic and reproductive growth. Metabolic rates were not significantly affected, but ammonium excretion increased in response to elevated pCO2. This led to decreased O:N ratios. These findings suggest that protein metabolism is possibly enhanced under elevated pCO2 in order to support ion homeostasis by increasing net acid extrusion. The perivisceral coelomic fluid acid-base status revealed that S. droebachiensis is able to fully (intermediate pCO2) or partially (high pCO2) compensate extracellular pH (pHe) changes by accumulation of bicarbonate (maximum increases 2.5 mM), albeit at a slower rate than typically observed in other taxa (10 day duration for full pHe compensation). At intermediate pCO2, sea urchins were able to maintain fully compensated pHe for 45 days. Sea urchins from the higher pCO2 treatment could be divided into two groups following medium-term acclimation: one group of experimental animals (29%) contained remnants of food in their digestive system and maintained partially compensated pHe (+2.3 mM HCO3), while the other group (71%) exhibited an empty digestive system and a severe metabolic acidosis (-0.5 pH units, -2.4 mM HCO3). There was no difference in mortality between the three pCO2 treatments. The results of this study suggest that S. droebachiensis occurring in the Kattegat might be pre-adapted to hypercapnia due to natural variability in pCO2 in its habitat. We show for the first time that some echinoderm species can actively compensate extracellular pH. Seawater pCO2 values of >200 Pa, which will occur in the Kattegat within this century during seasonal hypoxic events, can possibly only be endured for a short time period of a few weeks. Increases in anthropogenic CO2 emissions and leakages from potential sub-seabed CO2 storage (CCS) sites thus impose a threat to the ecologically and economically important species S. droebachiensis.

Could the acid-base status of Antarctic sea urchins indicate a better-than-expected resilience to near-future ocean acidification?

Increasing atmospheric carbon dioxide concentration alters the chemistry of the oceans towards more acidic conditions. Polar oceans are particularly affected due to their low temperature, low carbonate content and mixing patterns, for instance upwellings. Calcifying organisms are expected to be highly impacted by the decrease in the oceans' pH and carbonate ions concentration. In particular, sea urchins, members of the phylum Echinodermata, are hypothesized to be at risk due to their high-magnesium calcite skeleton. However, tolerance to ocean acidification in metazoans is first linked to acid-base regulation capacities of the extracellular fluids. No information on this is available to date for Antarctic echinoderms and inference from temperate and tropical studies needs support. In this study, we investigated the acid-base status of 9 species of sea urchins (3 cidaroids, 2 regular euechinoids and 4 irregular echinoids). It appears that Antarctic regular euechinoids seem equipped with similar acid-base regulation systems as tropical and temperate regular euechinoids but could rely on more passive ion transfer systems, minimizing energy requirements. Cidaroids have an acid-base status similar to that of tropical cidaroids. Therefore Antarctic cidaroids will most probably not be affected by decreasing seawater pH, the pH drop linked to ocean acidification being negligible in comparison of the naturally low pH of the coelomic fluid. Irregular echinoids might not suffer from reduced seawater pH if acidosis of the coelomic fluid pH does not occur but more data on their acid-base regulation are needed. Combining these results with the resilience of Antarctic sea urchin larvae strongly suggests that these organisms might not be the expected victims of ocean acidification. However, data on the impact of other global stressors such as temperature and of the combination of the different stressors needs to be acquired to assess the sensitivity of these organisms to global change.

Seawater carbonate chemistry and conditions of Mytilus edulis extracellular body fluids and shell composition in a pH-treatment experiment: Acid-base status, trace elements and delta11B, 2012

Mytilus edulis were cultured for 3 months under six different seawater pCO2 levels ranging from 380 to 4000 µatm. Specimen were taken from Kiel Fjord (Western Baltic Sea, Germany) which is a habitat with high and variable seawater pCO2 and related shifts in carbonate system speciation (e.g., low pH and low CaCO3 saturation state). Hemolymph (HL) and extrapallial fluid (EPF) samples were analyzed for pH and total dissolved inorganic carbon (CT) to calculate pCO2 and [HCO3]. A second experiment was conducted for 2 months with three different pCO2 levels (380, 1400 and 4000 µatm). Boron isotopes (delta11B) were investigated by LA-MC-ICP-MS (Laser Ablation-Multicollector-Inductively Coupled Plasma-Mass Spectrometry) in shell portions precipitated during experimental treatment time. Additionally, elemental ratios (B/Ca, Mg/Ca and Sr/Ca) in the EPF of specimen from the second experiment were measured via ICP-OES (Inductively Coupled Plasma-Optical Emission Spectrometry). Extracellular pH was not significantly different in HL and EPF but systematically lower than ambient water pH. This is due to high extracellular pCO2 values, a prerequisite for metabolic CO2 excretion. No accumulation of extracellular [HCO3] was measured. Elemental ratios (B/Ca, Mg/Ca and Sr/Ca) in the EPF increased slightly with pH which is in accordance with increasing growth and calcification rates at higher seawater pH values. Boron isotope ratios were highly variable between different individuals but also within single shells. This corresponds to a high individual variability in fluid B/Ca ratios and may be due to high boron concentrations in the organic parts of the shell. The mean delta11B value shows no trend with pH but appears to represent internal pH (EPF) rather than ambient water pH.

Acid-base balance and changes in haemolymph properties of the South African rock lobsters, Jasus lalandii, a palinurid decapod, during chronic hypercapnia

Few studies exist reporting on long-term exposure of crustaceans to hypercapnia. We exposed juvenile South African rock lobsters, Jasus lalandii, to hypercapnic conditions of pH 7.3 for 28 weeks and subsequently analysed changes in the extracellular fluid (haemolymph). Results revealed, for the first time, adjustments in the haemolymph of a palinurid crustacean during chronic hypercapnic exposure: 1) acid-base balance was adjusted and sustained by increased bicarbonate and 2) quantity and oxygen binding properties of haemocyanin changed. Compared with lobsters kept under normocapnic conditions (pH 8.0), during prolonged hypercapnia, juvenile lobsters increased bicarbonate buffering of haemolymph. This is necessary to provide optimum pH conditions for oxygen binding of haemocyanin and functioning of respiration in the presence of a strong Bohr Effect. Furthermore, modification of the intrinsic structure of the haemocyanin molecule, and not the presence of molecular modulators, seems to improve oxygen affinity under conditions of elevated pCO2.

Impact of elevated pCO2 on acid-base regulation of the sea urchin Echinometra mathaei and its relation to resistance to ocean acidification: A study in mesocosms

Due to their low metabolism and apparent poor ion regulation ability, sea urchins could be particularly sensitive to ocean acidification resulting from increased dissolution of atmospheric carbon dioxide. Therefore, we evaluated the acid-base regulation ability of the coral reef sea urchin Echinometra mathaei and the impact of decreased pH on its growth and respiration activity. The study was conducted in two identical artificial reef mesocosms during seven weeks. Experimental tanks were maintained respectively at mean pHT 7.7 and 8.05 (with field-like night and day variations). The major physico-chemical parameters were identical, only pCO2 and pHT differed. Results indicate that E. mathaei can regulate the pH of its coelomic fluid in the considered range of pH, allowing a sustainable growth and ensuring an unaffected respiratory metabolism, at least at short term.

Acid-base regulation in the Dungeness crab (Metacarcinus magister)

Homeostatic regulation allows organisms to secure basic physiological processes in a varying environment. To counteract fluctuations in ambient carbonate system speciation due to elevated seawater pCO2 (hypercapnia), many aquatic crustaceans excrete/accumulate acid-base equivalents through their gills; however, not much is known about the role of ammonia in this response. The present study investigated the effects of hypercapnia on acid-base and ammonia regulation in the Dungeness crab, Metacarcinus magister on the whole animal and isolated gill levels. Hemolymph pCO2 and [HCO3]- increased in M. magister acclimated to elevated pCO2 (330 Pa), while pH remained stable. Additionally, hemolymph [Na+], [Ca2+], and [SO4]2- were significantly increased. When challenged with varying pH during gill perfusion, the pH of the artificial hemolymph remained relatively unchanged. Overall, ammonia production and excretion, as well as oxygen consumption, were reduced in crabs acclimated to elevated pCO2, demonstrating that either (amino acid) oxidation is reduced in response to this particular stress, or nitrogenous wastes are excreted in an alternative form.

Acid-base physiology response to ocean acidification of two ecologically and economically important holothuroids from contrasting habitats, Holothuria scabra and Holothuria parva

Sea cucumbers are dominant invertebrates in several ecosystems such as coral reefs, seagrass meadows and mangroves. As bioturbators, they have an important ecological role in making available calcium carbonate and nutrients to the rest of the community. However, due to their commercial value, they face overexploitation in the natural environment. On top of that, occurring ocean acidification could impact these organisms, considered sensitive as echinoderms are osmoconformers, high-magnesium calcite producers and have a low metabolism. As a first investigation of the impact of ocean acidification on sea cucumbers, we tested the impact of short-term (6 to 12 days) exposure to ocean acidification (seawater pH 7.7 and 7.4) on two sea cucumbers collected in SW Madagascar, Holothuria scabra, a high commercial value species living in the seagrass meadows, and H. parva, inhabiting the mangroves. The former lives in a habitat with moderate fluctuations of seawater chemistry (driven by day-night differences) while the second lives in a highly variable intertidal environment. In both species, pH of the coelomic fluid was significantly negatively affected by reduced seawater pH, with a pronounced extracellular acidosis in individuals maintained at pH 7.7 and 7.4. This acidosis was due to an increased dissolved inorganic carbon content and pCO2 of the coelomic fluid, indicating a limited diffusion of the CO2 towards the external medium. However, respiration and ammonium excretion rates were not affected. No evidence of accumulation of bicarbonate was observed to buffer the coelomic fluid pH. If this acidosis stays uncompensated for when facing long-term exposure, other processes could be affected in both species, eventually leading to impacts on their ecological role.

Differential acid-base regulation in various gills of the green crab Carcinus maenas: Effects of elevated environmental pCO2

Euryhaline decapod crustaceans possess an efficient regulation apparatus located in the gill epithelia, providing a high adaptation potential to varying environmental abiotic conditions. Even though many studies focussed on the osmoregulatory capacity of the gills, acid-base regulatory mechanisms have obtained much less attention. In the present study, underlying principles and effects of elevated pCO2 on acid-base regulatory patterns were investigated in the green crab Carcinus maenas acclimated to diluted seawater. In gill perfusion experiments, all investigated gills 4-9 were observed to up-regulate the pH of the hemolymph by 0.1-0.2 units. Anterior gills, especially gill 4, were identified to be most efficient in the equivalent proton excretion rate. Ammonia excretion rates mirrored this pattern among gills, indicating a linkage between both processes. In specimen exposed to elevated pCO2 levels for at least 7 days, mimicking a future ocean scenario as predicted until the year 2300, hemolymph K+ and ammonia concentrations were significantly elevated, and an increased ammonia excretion rate was observed. A detailed quantitative gene expression analysis revealed that upon elevated pCO2 exposure, mRNA levels of transcripts hypothesized to be involved in ammonia and acid-base regulation (Rhesus-like protein, membrane-bound carbonic anhydrase, Na+/K+-ATPase) were affected predominantly in the non-osmoregulating anterior gills.

Seawater carbonate chemistry and biological processes of mussel Crassostrea gigas during experiments, 2011

Climate change with increasing temperature and ocean acidification (OA) poses risks for marine ecosystems. According to Pörtner and Farrell [1], synergistic effects of elevated temperature and CO2-induced OA on energy metabolism will narrow the thermal tolerance window of marine ectothermal animals. To test this hypothesis, we investigated the effect of an acute temperature rise on energy metabolism of the oyster, Crassostrea gigas chronically exposed to elevated CO2 levels (partial pressure of CO2 in the seawater ~0.15 kPa, seawater pH ~ 7.7). Within one month of incubation at elevated PCO2 and 15 °C hemolymph pH fell (pHe = 7.1 ± 0.2 (CO2-group) vs. 7.6 ± 0.1 (control)) and PeCO2 values in hemolymph increased (0.5 ± 0.2 kPa (CO2-group) vs. 0.2 ± 0.04 kPa (control)). Slightly but significantly elevated bicarbonate concentrations in the hemolymph of CO2-incubated oysters ([HCO-3]e = 1.8 ± 0.3 mM (CO2-group) vs. 1.3 ± 0.1 mM (control)) indicate only minimal regulation of extracellular acid-base status. At the acclimation temperature of 15 °C the OA-induced decrease in pHe did not lead to metabolic depression in oysters as standard metabolism rates (SMR) of CO2-exposed oysters were similar to controls. Upon acute warming SMR rose in both groups, but displayed a stronger increase in the CO2-incubated group. Investigation in isolated gill cells revealed a similar temperature-dependence of respiration between groups. Furthermore, the fraction of cellular energy demand for ion regulation via Na+/K+-ATPase was not affected by chronic hypercapnia or temperature. Metabolic profiling using 1H-NMR spectroscopy revealed substantial changes in some tissues following OA exposure at 15 °C. In mantle tissue alanine and ATP levels decreased significantly whereas an increase in succinate levels was observed in gill tissue. These findings suggest shifts in metabolic pathways following OA-exposure. Our study confirms that OA affects energy metabolism in oysters and suggests that climate change may affect populations of sessile coastal invertebrates such as mollusks

Grazing rates and body mass of Calanus finmarchicus and Calanus glacialis incubated under elevated pCO2

It is currently under debate whether organisms that regulate their acid-base status under environmental hypercapnia demand additional energy. This could impair animal fitness, but might be compensated for via increased ingestion rates when food is available. No data are yet available for dominant Calanus spp. from boreal and Arctic waters. To fill this gap, we incubated C. glacialis at 390, 1120 and 3000 µatm for 16 days with Thalassiosira weissflogii (diatom) as food source on-board RV Polarstern in Fram Strait in 2012. Every four days copepods were sub-sampled from all CO2 treatments and clearance and ingestion rates were determined. During the SOPRAN mesocosm experiment in Bergen, Norway, 2011, we weekly collected C. finmarchicus from mesocosms initially adjusted to 390 and 3000 µatm CO2 and measured grazing at low and high pCO2. In addition, copepods were deep frozen for body mass analyses. Elevated pCO2 did not directly affect grazing activities and body mass, suggesting that the copepods did not have additional energy demands for coping with acidification, neither during long-term exposure nor after immediate changes in pCO2. Shifts in seawater pH thus do not seem to challenge these copepod species.

Seawater carbonate chemistry and biological processes during experiments with the spider crab Hyas araneus in Kongsfjorden, Svalbard, 2009

With global climate change, ocean warming and acidification occur concomitantly. In this study, we tested the hypothesis that increasing CO2 levels affect the acid-base balance and reduce the activity capacity of the Arctic spider crab Hyas araneus, especially at the limits of thermal tolerance. Crabs were acclimated to projected oceanic CO2 levels for 12 days (today: 380, towards the year 2100: 750 and 1,120 and beyond: 3,000 ?atm) and at two temperatures (1 and 4 °C). Effects of these treatments on the righting response (RR) were determined (1) at acclimation temperatures followed by (2) righting when exposed to an additional acute (15 min) heat stress at 12 °C. Prior to (resting) and after the consecutive stresses of combined righting activity and heat exposure, acid-base status and lactate contents were measured in the haemolymph. Under resting conditions, CO2 caused a decrease in haemolymph pH and an increase in oxygen partial pressure. Despite some buffering via an accumulation of bicarbonate, the extracellular acidosis remained uncompensated at 1 °C, a trend exacerbated when animals were acclimated to 4 °C. The additional combined exposure to activity and heat had only a slight effect on blood gas and acid-base status. Righting activity in all crabs incubated at 1 and 4 °C was unaffected by elevated CO2 levels or acute heat stress but was significantly reduced when both stressors acted synergistically. This impact was much stronger in the group acclimated at 1 °C where some individuals acclimated to high CO2 levels stopped responding. Lactate only accumulated in the haemolymph after combined righting and heat stress. In the group acclimated to 1 °C, lactate content was highest under normocapnia and lowest at the highest CO2 level in line with the finding that RR was largely reduced. In crabs acclimated to 4 °C, the RR was less affected by CO2 such that activity caused lactate to increase with rising CO2 levels. In line with the concept of oxygen and capacity limited thermal tolerance, all animals exposed to temperature extremes displayed a reduction in scope for performance, a trend exacerbated by increasing CO2 levels. Additionally, the differences seen between cold- and warm-acclimated H. araneus after heat stress indicate that a small shift to higher acclimation temperatures also alleviates the response to temperature extremes, indicating a shift in the thermal tolerance window which reduces susceptibility to additional CO2 exposure.

Data compilation on the biological response to ocean acidification: environmental and experimental context of data sets and related literature

The exponential growth of studies on the biological response to ocean acidification over the last few decades has generated a large amount of data. To facilitate data comparison, a data compilation hosted at the data publisher PANGAEA was initiated in 2008 and is updated on a regular basis (doi:10.1594/PANGAEA.149999). By January 2015, a total of 581 data sets (over 4 000 000 data points) from 539 papers had been archived. Here we present the developments of this data compilation five years since its first description by Nisumaa et al. (2010). Most of study sites from which data archived are still in the Northern Hemisphere and the number of archived data from studies from the Southern Hemisphere and polar oceans are still relatively low. Data from 60 studies that investigated the response of a mix of organisms or natural communities were all added after 2010, indicating a welcomed shift from the study of individual organisms to communities and ecosystems. The initial imbalance of considerably more data archived on calcification and primary production than on other processes has improved. There is also a clear tendency towards more data archived from multifactorial studies after 2010. For easier and more effective access to ocean acidification data, the ocean acidification community is strongly encouraged to contribute to the data archiving effort, and help develop standard vocabularies describing the variables and define best practices for archiving ocean acidification data.