Seawater carbonate chemistry and biological processes of oysters Crassostrea virginica during experiments, 2010

Estuarine organisms are exposed to periodic strong fluctuations in seawater pH driven by biological carbon dioxide (CO2) production, which may in the future be further exacerbated by the ocean acidification associated with the global rise in CO2. Calcium carbonate-producing marine species such as mollusks are expected to be vulnerable to acidification of estuarine waters, since elevated CO2 concentration and lower pH lead to a decrease in the degree of saturation of water with respect to calcium carbonate, potentially affecting biomineralization. Our study demonstrates that the increase in CO2 partial pressure (pCO2) in seawater and associated decrease in pH within the environmentally relevant range for estuaries have negative effects on physiology, rates of shell deposition and mechanical properties of the shells of eastern oysters Crassostrea virginica (Gmelin). High CO2 levels (pH ~7.5, pCO2 ~3500 µatm) caused significant increases in juvenile mortality rates and inhibited both shell and soft-body growth compared to the control conditions (pH ~8.2, pCO2 ~380 µatm). Furthermore, elevated CO2 concentrations resulted in higher standard metabolic rates in oyster juveniles, likely due to the higher energy cost of homeostasis. The high CO2 conditions also led to changes in the ultrastructure and mechanical properties of shells, including increased thickness of the calcite laths within the hypostracum and reduced hardness and fracture toughness of the shells, indicating that elevated CO2 levels have negative effects on the biomineralization process. These data strongly suggest that the rise in CO2 can impact physiology and biomineralization in marine calcifiers such as eastern oysters, threatening their survival and potentially leading to profound ecological and economic impacts in estuarine ecosystems.

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

Seawater carbonate chemistry and shell weights of Laternula elliptica, Yoldia eightsi, Nacella concinna and Liothyrella uva during experiments, 2009

Antarctic calcified macroorganisms are particularly vulnerable to ocean acidification because many are weakly calcified, the dissolution rates of calcium carbonate are inversely related to temperature, and high latitude seas are predicted to become undersaturated in aragonite by the year 2100. We examined the post-mortem dissolution rates of aragonitic and calcitic shells from four species of Antarctic benthic marine invertebrates (two bivalves, one limpet, one brachiopod) and the thallus of a limpet shell-encrusting coralline alga exposed to acidified pH (7.4) or non-acidified pH (8.2) seawater at a constant temperature of 4 C. Within a period of only 14-35 days, shells of all four species held in pH 7.4 seawater had suffered significant dissolution. Despite calcite being 35% less soluble in seawater than aragonite, there was surprisingly, no consistent pattern of calcitic shells having slower dissolution rates than aragonitic shells. Outer surfaces of shells held in pH 7.4 seawater exhibited deterioration by day 35, and by day 56 there was exposure of aragonitic or calcitic prisms within the shell architecture of three of the macroinvertebrate species. Dissolution of coralline algae was confirmed by differences in weight loss in limpet shells with and without coralline algae. By day 56, thalli of the coralline alga held in pH 7.4 displayed a loss of definition of the conceptacle pores and cracking was evident at the zone of interface with limpet shells. Experimental studies are needed to evaluate whether there are adequate compensatory mechanisms in these and other calcified Antarctic benthic macroorganisms to cope with anticipated ocean acidification. In their absence, these organisms, and the communities they comprise, are likely to be among the first to experience the cascading impacts of ocean acidification.

Effect of increased pCO2 level on early shell development in great scallop (Pecten maximus Lamarck) larvae

As a result of high anthropogenic CO2 emissions, the concentration of CO2 in the oceans has increased, causing a decrease in pH, known as ocean acidification (OA). Numerous studies have shown negative effects on marine invertebrates, and also that the early life stages are the most sensitive to OA. We studied the effects of OA on embryos and unfed larvae of the great scallop (Pecten maximus Lamarck), at pCO(2) levels of 469 (ambient), 807, 1164, and 1599 µatm until seven days after fertilization. To our knowledge, this is the first study on OA effects on larvae of this species. A drop in pCO(2) level the first 12 h was observed in the elevated pCO(2) groups due to a discontinuation in water flow to avoid escape of embryos. When the flow was restarted, pCO(2) level stabilized and was significantly different between all groups. OA affected both survival and shell growth negatively after seven days. Survival was reduced from 45% in the ambient group to 12% in the highest pCO(2) group. Shell length and height were reduced by 8 and 15 %, respectively, when pCO(2) increased from ambient to 1599 µatm. Development of normal hinges was negatively affected by elevated pCO(2) levels in both trochophore larvae after two days and veliger larvae after seven days. After seven days, deformities in the shell hinge were more connected to elevated pCO(2) levels than deformities in the shell edge. Embryos stained with calcein showed fluorescence in the newly formed shell area, indicating calcification of the shell at the early trochophore stage between one and two days after fertilization. Our results show that P. maximus embryos and early larvae may be negatively affected by elevated pCO(2) levels within the range of what is projected towards year 2250, although the initial drop in pCO(2) level may have overestimated the effect of the highest pCO(2) levels. Future work should focus on long-term effects on this species from hatching, throughout the larval stages, and further into the juvenile and adult stages.

Seawater carbonate chemistry, shell linear extension and calcification during calcein staining and 45Ca experiments with pteropod Limacina helicina, 2009

Thecosome pteropods (shelled pelagic molluscs) can play an important role in the food web of various ecosystems and play a key role in the cycling of carbon and carbonate. Since they harbor an aragonitic shell, they could be very sensitive to ocean acidification driven by the increase of anthropogenic CO2 emissions. The impact of changes in the carbonate chemistry was investigated on Limacina helicina, a key species of Arctic ecosystems. Pteropods were kept in culture under controlled pH conditions corresponding to pCO2 levels of 350 and 760 µatm. Calcification was estimated using a fluorochrome and the radioisotope 45Ca. It exhibits a 28 % decrease at the pH value expected for 2100 compared to the present pH value. This result supports the concern for the future of pteropods in a high-CO2 world, as well as of those species dependent upon them as a food resource. A decline of their populations would likely cause dramatic changes to the structure, function and services of polar ecosystems.

Seawater carbonate chemistry and proportion of different dissolution levels in live juvenile Limacina helicina antarctica from the natural environment and ship-board incubations

The carbonate chemistry of the surface ocean is rapidly changing with ocean acidification, a result of human activities. In the upper layers of the Southern Ocean, aragonite-a metastable form of calcium carbonate with rapid dissolution kinetics-may become undersaturated by 2050. Aragonite undersaturation is likely to affect aragonite-shelled organisms, which can dominate surface water communities in polar regions. Here we present analyses of specimens of the pteropod Limacina helicina antarctica that were extracted live from the Southern Ocean early in 2008. We sampled from the top 200 m of the water column, where aragonite saturation levels were around 1, as upwelled deep water is mixed with surface water containing anthropogenic CO2. Comparing the shell structure with samples from aragonite-supersaturated regions elsewhere under a scanning electron microscope, we found severe levels of shell dissolution in the undersaturated region alone. According to laboratory incubations of intact samples with a range of aragonite saturation levels, eight days of incubation in aragonite saturation levels of 0.94-1.12 produces equivalent levels of dissolution. As deep-water upwelling and CO2 absorption by surface waters is likely to increase as a result of human activities, we conclude that upper ocean regions where aragonite-shelled organisms are affected by dissolution are likely to expand.

Seawater carbonate chemistry and biological processes of Pandalus borealis during experiments, 2011

Ocean acidification (OA) resulting from anthropogenic emissions of carbon dioxide (CO2) has already lowered and is predicted to further lower surface ocean pH. There is a particular need to study effects of OA on organisms living in cold-water environments due to the higher solubility of CO2 at lower temperatures. Mussel larvae (Mytilus edulis) and shrimp larvae (Pandalus borealis) were kept under an ocean acidification scenario predicted for the year 2100 (pH 7.6) and compared against identical batches of organisms held under the current oceanic pH of 8.1, which acted as a control. The temperature was held at a constant 10°C in the mussel experiment and at 5°C in the shrimp experiment. There was no marked effect on fertilization success, development time, or abnormality to the D-shell stage, or on feeding of mussel larvae in the low-pH (pH 7.6) treatment. Mytilus edulis larvae were still able to develop a shell in seawater undersaturated with respect to aragonite (a mineral form of CaCO3), but the size of low-pH larvae was significantly smaller than in the control. After 2 mo of exposure the mussels were 28% smaller in the pH 7.6 treatment than in the control. The experiment with Pandalus borealis larvae ran from 1 through 35 days post hatch. Survival of shrimp larvae was not reduced after 5 wk of exposure to pH 7.6, but a significant delay in zoeal progression (development time) was observed.

Seawater carbonate chemistry and biological processes of bivalve shellfish Mercenaria mercenaria and Argopecten irradians during experiments, 2011

The combustion of fossil fuels has enriched levels of CO2 in the world's oceans and decreased ocean pH. Although the continuation of these processes may alter the growth, survival, and diversity of marine organisms that synthesize CaCO3shells, the effects of ocean acidification since the dawn of the industrial revolution are not clear. Here we present experiments that examined the effects of the ocean's past, present, and future (21st and 22nd centuries) CO2concentrations on the growth, survival, and condition of larvae of two species of commercially and ecologically valuable bivalve shellfish (Mercenaria mercenariaand Argopecten irradians). Larvae grown under near preindustrial CO2concentrations (250 ppm) displayed significantly faster growth and metamorphosis as well as higher survival and lipid accumulation rates compared with individuals reared under modern day CO2 levels. Bivalves grown under near preindustrial CO2 levels displayed thicker, more robust shells than individuals grown at present CO2 concentrations, whereas bivalves exposed to CO2 levels expected later this century had shells that were malformed and eroded. These results suggest that the ocean acidification that has occurred during the past two centuries may be inhibiting the development and survival of larval shellfish and contributing to global declines of some bivalve populations.

Seawater carbonate chemistry and benthic marine community during experiments, 2011

Ocean acidification is predicted to impact all areas of the oceans and affect a diversity of marine organisms. However, the diversity of responses among species prevents clear predictions about the impact of acidification at the ecosystem level. Here, we used shallow water CO2 vents in the Mediterranean Sea as a model system to examine emergent ecosystem responses to ocean acidification in rocky reef communities. We assessed in situ benthic invertebrate communities in three distinct pH zones (ambient, low, and extreme low), which differed in both the mean and variability of seawater pH along a continuous gradient. We found fewer taxa, reduced taxonomic evenness, and lower biomass in the extreme low pH zones. However, the number of individuals did not differ among pH zones, suggesting that there is density compensation through population blooms of small acidification-tolerant taxa. Furthermore, the trophic structure of the invertebrate community shifted to fewer trophic groups and dominance by generalists in extreme low pH, suggesting that there may be a simplification of food webs with ocean acidification. Despite high variation in individual species' responses, our findings indicate that ocean acidification decreases the diversity, biomass, and trophic complexity of benthic marine communities. These results suggest that a loss of biodiversity and ecosystem function is expected under extreme acidification scenarios.

Seawater carbonate chemistry and biological processes during experiments with calcifiing organisms, 2009

Anthropogenic elevation of atmospheric carbon dioxide (pCO2) is making the oceans more acidic, thereby reducing their degree of saturation with respect to calcium carbonate (CaCO3). There is mounting concern over the impact that future CO2-induced reductions in the CaCO3 saturation state of seawater will have on marine organisms that construct their shells and skeletons from this mineral. Here, we present the results of 60 d laboratory experiments in which we investigated the effects of CO2-induced ocean acidification on calcification in 18 benthic marine organisms. Species were selected to span a broad taxonomic range (crustacea, cnidaria, echinoidea, rhodophyta, chlorophyta, gastropoda, bivalvia, annelida) and included organisms producing aragonite, low-Mg calcite, and high-Mg calcite forms of CaCO3. We show that 10 of the 18 species studied exhibited reduced rates of net calcification and, in some cases, net dissolution under elevated pCO2. However, in seven species, net calcification increased under the intermediate and/or highest levels of pCO2, and one species showed no response at all. These varied responses may reflect differences amongst organisms in their ability to regulate pH at the site of calcification, in the extent to which their outer shell layer is protected by an organic covering, in the solubility of their shell or skeletal mineral, and whether they utilize photosynthesis. Whatever the specific mechanism(s) involved, our results suggest that the impact of elevated atmospheric pCO2 on marine calcification is more varied than previously thought.

Grazing under experimental hypercapnia and elevated temperature does not affect the radula of a chiton (Mollusca, Polyplacophora, Lepidopleurida)

Chitons (class Polyplacophora) are benthic grazing molluscs with an eight-part aragonitic shell armature. The radula, a serial tooth ribbon that extends internally more than half the length of the body, is mineralised on the active feeding teeth with iron magnetite apparently as an adaptation to constant grazing on rocky substrates. As the anterior feeding teeth are eroded they are shed and replaced with a new row. The efficient mineralisation and function of the radula could hypothetically be affected by changing oceans in two ways: changes in seawater chemistry (pH and pCO2) may impact the biomineralisation pathway, potentially leading to a weaker or altered density of the feeding teeth; rising temperatures could increase activity levels in these ectothermic animals, and higher feeding rates could increase wear on the feeding teeth beyond the animals' ability to synthesise, mineralise, and replace radular rows. We therefore examined the effects of pH and temperature on growth and integrity in the radula of the chiton Leptochiton asellus. Our experiment implemented three temperature (10, 15, 20 °C) and two pCO2 treatments (400 µatm, pH 8.0; 2000 µatm, pH 7.5) for six treatment groups. Animals (n = 50) were acclimated to the treatment conditions for a period of 4 weeks. This is sufficient time for growth of ca. 7-9 new tooth rows or 20% turnover of the mineralised portion. There was no significant difference in the number of new (non-mineralised) teeth or total tooth row count in any treatment. Examination of the radulae via SEM revealed no differences in microwear or breakage on the feeding cusps correlating to treatment groups. The shell valves also showed no signs of dissolution. As a lineage, chitons have survived repeated shifts in Earth's climate through geological time, and at least their radulae may be robust to future perturbations.

Projected near-future CO2 levels increase activity and alter defensive behaviours in the tropical squid Idiosepius pygmaeus

Carbon dioxide (CO2) levels projected to occur in the oceans by the end of this century cause a range of behavioural effects in fish, but whether other highly active marine organisms, such as cephalopods, are similarly affected is unknown. We tested the effects of projected future CO2 levels (626 and 956 µatm) on the behaviour of male two-toned pygmy squid, Idiosepius pygmaeus. Exposure to elevated CO2 increased the number of active individuals by 19-25% and increased movement (number of line-crosses) by nearly 3 times compared to squid at present-day CO2. Squid vigilance and defensive behaviours were also altered by elevated CO2 with >80% of individuals choosing jet escape responses over defensive arm postures in response to a visual startle stimulus, compared with 50% choosing jet escape responses at control CO2. In addition, more escape responses were chosen over threat behaviours in body pattern displays at elevated CO2 and individuals were more than twice as likely to use ink as a defence strategy at 956 µatm CO2, compared with controls. Increased activity could lead to adverse effects on energy budgets as well as increasing visibility to predators. A tendency to respond to a stimulus with escape behaviours could increase survival, but may also be energetically costly and could potentially lead to more chases by predators compared with individuals that use defensive postures. These results demonstrate that projected future ocean acidification affects the behaviours of a tropical squid species.

Experiment: Food availability outweighs ocean acidification effects in juvenile Mytilus edulis

Ocean acidification is expected to decrease calcification rates of bivalves. Nevertheless in many coastal areas high pCO2 variability is encountered already today. Kiel Fjord (Western Baltic Sea) is a brackish (12-20 g kg-1) and CO2 enriched habitat, but the blue mussel Mytilus edulis dominates the benthic community. In a coupled field and laboratory study we examined the annual pCO2 variability in this habitat and the combined effects of elevated pCO2 and food availability on juvenile M. edulis growth and calcification. In the laboratory experiment, mussel growth and calcification were found to chiefly depend on food supply, with only minor impacts of pCO2 up to 3350 µatm. Kiel Fjord was characterized by strong seasonal pCO2 variability. During summer, maximal pCO2 values of 2500 µatm were observed at the surface and >3000 µatm at the bottom. However, the field growth experiment revealed seven times higher growth and calcification rates of M. edulis at a high pCO2 inner fjord field station (mean pCO2 ca. 1000 µatm) in comparison to a low pCO2 outer fjord station (ca. 600 µatm). In addition, mussels were able to outcompete the barnacle Amphibalanus improvisus at the high pCO2 site. High mussel productivity at the inner fjord site was enabled by higher particulate organic carbon concentrations. Kiel Fjord is highly impacted by eutrophication, which causes bottom water hypoxia and consequently high seawater pCO2. At the same time, elevated nutrient concentrations increase the energy availability for filter feeding organisms such as mussels. Thus M. edulis can dominate over a seemingly more acidification resistant species such as A. improvisus. We conclude that benthic stages of M. edulis tolerate high ambient pCO2 when food supply is abundant and that important habitat characteristics such as species interactions and energy availability need to be considered to predict species vulnerability to ocean acidification.

Seawater carbonate chemistry and Ruditapes decussatus biological processes during experiments, 2011

We investigated the effects of ocean acidification on juvenile clams Ruditapes decussatus (average shell length 10.24 mm) in a controlled CO2 perturbation experiment. The carbonate chemistry of seawater was manipulated by diffusing pure CO2, to attain two reduced pH levels (by -0.4 and -0.7 pH units), which were compared to unmanipulated seawater. After 75 days we found no differences among pH treatments in terms of net calcification, size or weight of the clams. The naturally elevated total alkalinity of local seawater probably contributed to buffer the effects of increased pCO2 and reduced pH. Marine organisms may, therefore, show diverse responses to ocean acidification at local scales, particularly in coastal, estuarine and transitional waters, where the physical-chemical characteristics of seawater are most variable. Mortality was significantly reduced in the acidified treatments. This trend was probably related to the occurrence of spontaneous spawning events in the control and intermediate acidification treatments. Spawning, which was unexpected due to the small size of the clams, was not observed for the pH -0.7 treatment, suggesting that the increased survival under acidified conditions may have been associated with a delay in the reproductive cycle of the clams. Future research about the impacts of ocean acidification on marine biodiversity should be extended to other types of biological and ecological processes, apart from biological calcification.

Effects of ocean acidification on overwintering juvenile Arctic pteropods Limacina helicina

Pteropods are planktonic mollusks that play an important role in the food web of various ecosystems, particularly at high latitudes. Because they produce an aragonitic shell, pteropods are expected to be very sensitive to ocean acidification driven by anthropogenic CO2 emissions. The effect of ocean acidification was investigated using juveniles of the Arctic pteropod Limacina helicina from the Canada Basin of the Arctic Ocean. The animals were maintained in 3 controlled pH conditions (total scale pH [pHT] = 8.05, 7.90 or 7.75) for 8 d, and their mortality and the linear extension of their shell were monitored. The pH did not impact the mortality rate, but the linear extension of the shell decreased as a function of declining pH. Surprisingly, the pteropods were still able to extend their shell at an aragonite saturation state as low as 0.6. Nevertheless, dissolution marks were visible on the whole shell, indicating that calcium carbonate dissolution had also occurred, casting doubts on the ability of the pteropods to maintain a positive balance between precipitation and dissolution of calcium carbonate under corrosive conditions.