Temperature Dependent Effects of Elevated CO2 on Shell Composition and Mechanical Properties of Hydroides elegans: Insights from a Multiple Stressor Experiment

The majority of marine benthic invertebrates protect themselves from predators by producing calcareous tubes or shells that have remarkable mechanical strength. An elevation of CO2 or a decrease in pH in the environment can reduce intracellular pH at the site of calcification and thus interfere with animal's ability to accrete CaCO3. In nature, decreased pH in combination with stressors associated with climate change may result in the animal producing severely damaged and mechanically weak tubes. This study investigated how the interaction of environmental drivers affects production of calcareous tubes by the serpulid tubeworm, Hydroides elegans. In a factorial manipulative experiment, we analyzed the effects of pH (8.1 and 7.8), salinity (34 and 27), and temperature (23°C and 29°C) on the biomineral composition, ultrastructure and mechanical properties of the tubes. At an elevated temperature of 29°C, the tube calcite/aragonite ratio and Mg/Ca ratio were both increased, the Sr/Ca ratio was decreased, and the amorphous CaCO3 content was reduced. Notably, at elevated temperature with decreased pH and reduced salinity, the constructed tubes had a more compact ultrastructure with enhanced hardness and elasticity compared to decreased pH at ambient temperature. Thus, elevated temperature rescued the decreased pH-induced tube impairments. This indicates that tubeworms are likely to thrive in early subtropical summer climate. In the context of climate change, tubeworms could be resilient to the projected near-future decreased pH or salinity as long as surface seawater temperature rise at least by 4°C.

Effects of elevated CO2 and temperature on an intertidal meiobenthic community

In the near future, the marine environment is likely to be subjected to simultaneous increases in temperature and decreased pH. The potential effects of these changes on intertidal, meiofaunal assemblages were investigated using a mesocosm experiment. Artificial Substrate Units containing meiofauna from the extreme low intertidal zone were exposed for 60 days to eight experimental treatments (four replicates for each treatment) comprising four pH levels: 8.0 (ambient control), 7.7 & 7.3 (predicted changes associated with ocean acidification), and 6.7 (CO2 point-source leakage from geological storage), crossed with two temperatures: 12 °C (ambient control) and 16 °C (predicted). Community structure, measured using major meiofauna taxa was significantly affected by pH and temperature. Copepods and copepodites showed the greatest decline in abundance in response to low pH and elevated temperature. Nematodes increased in abundance in response to low pH and temperature rise, possibly caused by decreased predation and competition for food owing to the declining macrofauna density. Nematode species composition changed significantly between the different treatments, and was affected by both seawater acidification and warming. Estimated nematode species diversity, species evenness, and the maturity index, were substantially lower at 16 °C, whereas trophic diversity was slightly higher at 16 °C except at pH 6.7. This study has demonstrated that the combination of elevated levels of CO2 and ocean warming may have substantial effects on structural and functional characteristics of meiofaunal and nematode communities, and that single stressor experiments are unlikely to encompass the complexity of abiotic and biotic interactions. At the same time, ecological interactions may lead to complex community responses to pH and temperature changes in the interstitial environment.

Metabolic costs of larval settlement and metamorphosis in the coral Seriatopora caliendrum under ambient and elevated pCO2

We tested the effects of pCO2 on Seriatopora caliendrum recruits over the first 5.3 d of post-settlement existence. In March 2011, 11-20 larvae were settled in glass vials (3.2 mL) and incubated at 24.0 °C and ~250 µmol quanta/m**2/s while supplied with seawater (at 1.4 mL/s) equilibrated with 51.6 Pa pCO2 (ambient) or 86.4 Pa pCO2. At 51.6 Pa pCO2, mean respiration 7 h post-settlement was 0.056 ± 0.007 nmol O2/recruit/min, but rose quickly to 0.095 ± 0.007 nmol O2/recruit/min at 3.3 d post-settlement, and thereafter declined to 0.075 ± 0.002 nmol O2/recruit/min at 5.3 d post-settlement (all ± SE). Elevated pCO2 depressed respiration of recruits by 19% after 3.3 d and 12% overall (i.e., integrated over 5.3 d), and while it had no effect on corallite area, elevated pCO2 was associated with weaker adhesion to the glass settlement surface and lower protein biomass. The unique costs of settlement and metamorphosis for S. caliendrum over 5.3 d are estimated to be 257 mJ/recruit at 51.6 Pa pCO2, which is less than the energy content of the larvae and recruits.

Larval and post-larval stages of pacific oyster (Crassostrea gigas) are resistant to elevated CO2

Rising anthropogenic carbon dioxide (CO2) dissolving into coastal waters is decreasing the pH and carbonate ion concentration, thereby lowering the saturation state of calcium carbonate (CaCO3) minerals through a process named ocean acidification (OA). The unprecedented threats posed by such low pH on calcifying larvae of several edible oyster species have not yet been fully explored. Effects of low pH (7.9, 7.6, 7.4) on the early growth phase of Portuguese oyster (Crassostrea angulata) veliger larvae was examined at ambient salinity (34 ppt) and the low-salinity (27 ppt) treatment. Additionally, the combined effect of pH (8.1, 7.6), salinity (24 and 34 ppt) and temperature (24 °C and 30 °C) was examined using factorial experimental design. Surprisingly, the early growth phase from hatching to 5-day-old veliger stage showed high tolerance to pH 7.9 and pH 7.6 at both 34 ppt and 27 ppt. Larval shell area was significantly smaller at pH 7.4 only in low-salinity. In the 3-factor experiment, shell area was affected by salinity and the interaction between salinity and temperature but not by other combinations. Larvae produced the largest shell at the elevated temperature in low-salinity, regardless of pH. Thus the growth of the Portuguese oyster larvae appears to be robust to near-future pH level (> 7.6) when combined with projected elevated temperature and low-salinity in the coastal aquaculture zones of South China Sea.

Elevated CO2 affects the behavior of an ecologically and economically important coral reef fish

We tested the effect of near-future CO2 levels (= 490, 570, 700, and 960 µatm CO2) on the olfactory responses and activity levels of juvenile coral trout, Plectropomus leopardus, a piscivorous reef fish that is also one of the most important fisheries species on the Great Barrier Reef, Australia. Juvenile coral trout reared for 4 weeks at 570 µatm CO2 exhibited similar sensory responses and behaviors to juveniles reared at 490 µatm CO2 (control). In contrast, juveniles reared at 700 and 960 µatm CO2 exhibited dramatically altered sensory function and behaviors. At these higher CO2 concentrations, juveniles became attracted to the odor of potential predators, as has been observed in other reef fishes. They were more active, spent less time in shelter, ventured further from shelter, and were bolder than fish reared at 490 or 570 µatm CO2. These results demonstrate that behavioral impairment of coral trout is unlikely if pCO2 remains below 600 µatm; however, at higher levels, there are significant impacts on juvenile performance that are likely to affect survival and energy budgets, with consequences for predator-prey interactions and commercial fisheries.

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.

Experiment: Elevated CO2 levels affect the activity of nitrate reductase and carbonic anhydrase in the calcifying rhodophyte Corallina officinalis

The concentration of CO2 in global surface ocean waters is increasing due to rising atmospheric CO2 emissions, resulting in lower pH and a lower saturation state of carbonate ions. Such changes in seawater chemistry are expected to impact calcification in calcifying marine organisms. However, other physiological processes related to calcification might also be affected, including enzyme activity. In a mesocosm experiment, macroalgal communities were exposed to three CO2 concentrations (380, 665, and 1486 µatm) to determine how the activity of two enzymes related to inorganic carbon uptake and nutrient assimilation in Corallina officinalis, an abundant calcifying rhodophyte, will be affected by elevated CO2 concentrations. The activity of external carbonic anhydrase, an important enzyme functioning in macroalgal carbon-concentrating mechanisms, was inversely related to CO2 concentration after long-term exposure (12 weeks). Nitrate reductase, the enzyme responsible for reduction of nitrate to nitrite, was stimulated by CO2 and was highest in algae grown at 665 µatm CO2. Nitrate and phosphate uptake rates were inversely related to CO2, while ammonium uptake was unaffected, and the percentage of inorganic carbon in the algal skeleton decreased with increasing CO2. The results indicate that the processes of inorganic carbon and nutrient uptake and assimilation are affected by elevated CO2 due to changes in enzyme activity, which change the energy balance and physiological status of C. officinalis, therefore affecting its competitive interactions with other macroalgae. The ecological implications of the physiological changes in C. officinalis in response to elevated CO2 are discussed.

Impact of elevated pCO2 on paralytic shellfish poisoning toxin content and composition in Alexandrium tamarense

Ocean acidification is considered a major threat to marine ecosystems and may particularly affect primary producers. Here we investigated the impact of elevated pCO2 on paralytic shellfish poisoning toxin (PST) content and composition in two strains of Alexandrium tamarense, Alex5 and Alex2. Experiments were carried out as dilute batch to keep carbonate chemistry unaltered over time. We observed only minor changes with respect to growth and elemental composition in response to elevated pCO2. For both strains, the cellular PST content, and in particular the associated cellular toxicity, was lower in the high CO2 treatments. In addition, Alex5 showed a shift in its PST composition from a nonsulfated analogue towards less toxic sulfated analogues with increasing pCO2. Transcriptomic analyses suggest that the ability of A. tamarense to maintain cellular homeostasis is predominantly regulated on the post-translational level rather than on the transcriptomic level. Furthermore, genes associated to secondary metabolite and amino acid metabolism in Alex5 were down-regulated in the high CO2 treatment, which may explain the lower PST content. Elevated pCO2 also induced up-regulation of a putative sulfotransferase sxtN homologue and a substantial down-regulation of several sulfatases. Such changes in sulfur metabolism may explain the shift in PST composition towards more sulfated analogues. All in all, our results indicate that elevated pCO2 will have minor consequences for growth and elemental composition, but may potentially reduce the cellular toxicity of A. tamarense.

Parental exposure to elevated pCO2 influences the reproductive success of copepods

Substantial variations are reported for egg production and hatching rates of copepods exposed to elevated carbon dioxide concentrations (pCO2). One possible explanation, as found in other marine taxa, is that prior parental exposure to elevated pCO2 (and/or decreased pH) affects reproductive performance. Previous studies have adopted two distinct approaches, either (1) expose male and female copepoda to the test pCO2/pH scenarios, or (2) solely expose egg-laying females to the tests. Although the former approach is more realistic, the majority of studies have used the latter approach. Here, we investigated the variation in egg production and hatching success of Acartia tonsa between these two experimental designs, across five different pCO2 concentrations (385-6000 µatm pCO2). In addition, to determine the effect of pCO2 on the hatching success with no prior parental exposure, eggs produced and fertilized under ambient conditions were also exposed to these pCO2 scenarios. Significant variations were found between experimental designs, with approach (1) resulting in higher impacts; here >20% difference was seen in hatching success between experiments at 1000 µatm pCO2 scenarios (2100 year scenario), and >85% at 6000 µatm pCO2. This study highlights the potential to misrepresent the reproductive response of a species to elevated pCO2 dependent on parental exposure.

Experiment: Competition between calcifying and noncalcifying temperate marine macroalgae under elevated CO2 levels

Since pre-industrial times, uptake of anthropogenic CO2 by surface ocean waters has caused a documented change of 0.1 pH units. Calcifying organisms are sensitive to elevated CO2 concentrations due to their calcium carbonate skeletons. In temperate rocky intertidal environments, calcifying and noncalcifying macroalgae make up diverse benthic photoautotrophic communities. These communities may change as calcifiers and noncalcifiers respond differently to rising CO2 concentrations. In order to test this hypothesis, we conducted an 86?d mesocosm experiment to investigate the physiological and competitive responses of calcifying and noncalcifying temperate marine macroalgae to 385, 665, and 1486 µatm CO2. We focused on comparing 2 abundant red algae in the Northeast Atlantic: Corallina officinalis (calcifying) and Chondrus crispus (noncalcifying). We found an interactive effect of CO2 concentration and exposure time on growth rates of C. officinalis, and total protein and carbohydrate concentrations in both species. Photosynthetic rates did not show a strong response. Calcification in C. officinalis showed a parabolic response, while skeletal inorganic carbon decreased with increasing CO2. Community structure changed, as Chondrus crispus cover increased in all treatments, while C. officinalis cover decreased in both elevated-CO2 treatments. Photochemical parameters of other species are also presented. Our results suggest that CO2 will alter the competitive strengths of calcifying and noncalcifying temperate benthic macroalgae, resulting in different community structures, unless these species are able to adapt at a rate similar to or faster than the current rate of increasing sea-surface CO2 concentrations.

Tolerance of juvenile barnacles (Amphibalanus improvisus) to warming and elevated pCO2

We investigated the impacts of warming and elevated pCO2 on newly settled Amphibalanus improvisus from Kiel Fjord, an estuarine ecosystem characterized by significant natural pCO2 variability. In two experiments, juvenile barnacles were maintained at two temperature and three pCO2 levels (20/24°C, 700-2.140 µatm) for 8 weeks in a batch culture and at four pCO2 levels (20°C, 620-2.870 µatm) for 12 weeks in a water flow-through system. Warming as well as elevated pCO2 hardly affected growth or the condition index of barnacles, although some factor combinations led to temporal significances in enhanced or reduced growth with an increase in pCO2. While warming increased the shell strength of A. improvisus individuals, elevated pCO2 had only weak effects. We demonstrate a strong tolerance of juvenile A. improvisus to mean acidification levels of about 1,000 µatm pCO2 as is already naturally experienced by the investigated barnacle population.

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.

Elevated CO2 levels do not affect the shell structure of the bivalve Arctica islandica from the western Baltic

Shells of the bivalve Arctica islandica are used to reconstruct paleo-environmental conditions (e.g. temperature) via biogeochemical proxies, i.e. biogenic components that are related closely to environmental parameters at the time of shell formation. Several studies have shown that proxies like element and isotope-ratios can be affected by shell growth and microstructure. Thus it is essential to evaluate the impact of changing environmental parameters such as high pCO2 and consequent changes in carbonate chemistry on shell properties to validate these biogeochemical proxies for a wider range of environmental conditions. Growth experiments with Arctica islandica from the Western Baltic Sea kept under different pCO2 levels (from 380 to 1120 µatm) indicate no affect of elevated pCO2 on shell growth or crystal microstructure, indicating that A. islandica shows an adaptation to a wider range of pCO2 levels than reported for other species. Accordingly, proxy information derived from A. islandica shells of this region contains no pCO2 related bias.

Multiple stressor effects of near-future elevated seawater temperature and decreased pH on righting and escape behaviors of two common Antarctic gastropods

Warming seawater temperatures and ocean acidification on the coastal western Antarctic Peninsula pose unique challenges to stenothermal marine invertebrates. The present study examines prospective sub-lethal effects of elevated temperature, pCO2, and resultant decrease in seawater pH, on righting behavior and maximal escape speeds for two common gastropods, the limpet Nacella concinna (Strebel) and mesogastropod snail Margarella antarctica (Lamy). Replicate individuals held in individual containers were exposed to four combinations of seawater temperature (1.5 °C-current average, 3.5 °C-projected average by 2100) and pH (pH 8.0-current average, pH 7.8-projected average by 2100 as a result of elevated pCO2 levels) for a period of 6 weeks. Following this chronic exposure, righting behavior, determined for the limpets as proportion to right over 24 h and for snails as time to right, as well as maximum escape speed following contact with a sea star predator were measured. We found no significant differences in proportions of limpets displaying the capacity to right among the four temperature-pH treatments. However, there was a significant temperature-pH interaction effect for mean righting times in snails, indicating that the effect of pH on the time to right is dependent on temperature. We found no significant effects of temperature or pH on mean maximal escape speed in limpets. Additionally, we observed a significant temperature-pH interaction effect for mean maximal escape speed in snails. These interactive effects make it difficult to make clear predictions about how these environmental factors may impact behavioral responses.