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.

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.

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.

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.

Seawater carbonate chemistry, clearance rates, respiration rates, condition index and cellular turnover (RNA: DNA) of Juvenile King Scallop, Pecten maximus in a laboratory experiment

The decline in ocean water pH and changes in carbonate saturation states through anthropogenically mediated increases in atmospheric CO2 levels may pose a hazard to marine organisms. This may be particularly acute for those species reliant on calcareous structures like shells and exoskeletons. This is of particular concern in the case of valuable commercially exploited species such as the king scallop, Pecten maximus. In this study we investigated the effects on oxygen consumption, clearance rates and cellular turnover in juvenile P. maximus following 3 months laboratory exposure to four pCO2 treatments (290, 380, 750 and 1140 µatm). None of the exposure levels were found to have significant effect on the clearance rates, respiration rates, condition index or cellular turnover (RNA: DNA) of individuals. While it is clear that some life stages of marine bivalves appear susceptible to future levels of ocean acidification, particularly under food limiting conditions, the results from this study suggest that where food is in abundance, bivalves like juvenile P. maximus may display a tolerance to limited changes in seawater chemistry.

Interactive effects of salinity and elevated CO2 levels on juvenile eastern oysters, Crassostrea virginica

Rising levels of atmospheric CO2 lead to acidification of the ocean and alter seawater carbonate chemistry, which can negatively impact calcifying organisms, including mollusks. In estuaries, exposure to elevated CO2 levels often co-occurs with other stressors, such as reduced salinity, which enhances the acidification trend, affects ion and acid-base regulation of estuarine calcifiers and modifies their response to ocean acidification. We studied the interactive effects of salinity and partial pressure of CO2 (PCO2) on biomineralization and energy homeostasis in juveniles of the eastern oyster, Crassostrea virginica, a common estuarine bivalve. Juveniles were exposed for 11 weeks to one of two environmentally relevant salinities (30 or 15 PSU) either at current atmospheric PCO2 (400 µatm, normocapnia) or PCO2 projected by moderate IPCC scenarios for the year 2100 (700-800 µatm, hypercapnia). Exposure of the juvenile oysters to elevated PCO2 and/or low salinity led to a significant increase in mortality, reduction of tissue energy stores (glycogen and lipid) and negative soft tissue growth, indicating energy deficiency. Interestingly, tissue ATP levels were not affected by exposure to changing salinity and PCO2, suggesting that juvenile oysters maintain their cellular energy status at the expense of lipid and glycogen stores. At the same time, no compensatory upregulation of carbonic anhydrase activity was found under the conditions of low salinity and high PCO2. Metabolic profiling using magnetic resonance spectroscopy revealed altered metabolite status following low salinity exposure; specifically, acetate levels were lower in hypercapnic than in normocapnic individuals at low salinity. Combined exposure to hypercapnia and low salinity negatively affected mechanical properties of shells of the juveniles, resulting in reduced hardness and fracture resistance. Thus, our data suggest that the combined effects of elevated PCO2 and fluctuating salinity may jeopardize the survival of eastern oysters because of weakening of their shells and increased energy consumption.

Rapid acclimation of juvenile corals to CO2-mediated acidification by upregulation of heat shock protein and Bcl-2 genes

Corals play a key role in ocean ecosystems and carbonate balance, but their molecular response to ocean acidification remains unclear. The only previous whole-transcriptome study documented extensive disruption of gene expression, particularly of genes encoding skeletal organic matrix proteins, in juvenile corals (Acropora millepora) after short-term (3 d) exposure to elevated pCO2. In this study, whole-transcriptome analysis was used to compare the effects of such 'acute' (3 d) exposure to elevated pCO2 with a longer ('prolonged'; 9 d) period of exposure beginning immediately post-fertilization. Far fewer genes were differentially expressed under the 9-d treatment, and although the transcriptome data implied wholesale disruption of metabolism and calcification genes in the acute treatment experiment, expression of most genes was at control levels after prolonged treatment. There was little overlap between the genes responding to the acute and prolonged treatments, but heat shock proteins (HSPs) and heat shock factors (HSFs) were over-represented amongst the genes responding to both treatments. Amongst these was an HSP70 gene previously shown to be involved in acclimation to thermal stress in a field population of another acroporid coral. The most obvious feature of the molecular response in the 9-d treatment experiment was the upregulation of five distinct Bcl-2 family members, the majority predicted to be anti-apoptotic. This suggests that an important component of the longer term response to elevated CO2 is suppression of apoptosis. It therefore appears that juvenile A. millepora have the capacity to rapidly acclimate to elevated pCO2, a process mediated by upregulation of specific HSPs and a suite of Bcl-2 family members.

Ocean acidification reduces the crystallographic control in juvenile mussel shells

Global climate change threatens the oceans as anthropogenic carbon dioxide causes ocean acidification and reduced carbonate saturation. Future projections indicate under saturation of aragonite, and potentially calcite, in the oceans by 2100. Calcifying organisms are those most at risk from such ocean acidification, as carbonate is vital in the biomineralisation of their calcium carbonate protective shells. This study highlights the importance of multi-generational studies to investigate how marine organisms can potentially adapt to future projected global climate change. Mytilus edulis is an economically important marine calcifier vulnerable to decreasing carbonate saturation as their shells comprise two calcium carbonate polymorphs: aragonite and calcite. M. edulis specimens were cultured under current and projected pCO2 (380, 550, 750 and 1000 µatm), following 6 months of experimental culture, adults produced second generation juvenile mussels. Juvenile mussel shells were examined for structural and crystallographic orientation of aragonite and calcite. At 1000 µatm pCO2, juvenile mussels spawned and grown under this high pCO2 do not produce aragonite which is more vulnerable to carbonate under-saturation than calcite. Calcite and aragonite were produced at 380, 550 and 750 µatm pCO2. Electron back scatter diffraction analyses reveal less constraint in crystallographic orientation with increased pCO2. Shell formation is maintained, although the nacre crystals appear corroded and crystals are not so closely layered together. The differences in ultrastructure and crystallography in shells formed by juveniles spawned from adults in high pCO2 conditions may prove instrumental in their ability to survive ocean acidification.

Effect of ocean acidification on growth and otolith condition of juvenile scup, Stenotomus chrysops

Increasing amounts of atmospheric carbon dioxide (CO2) from human industrial activities are causing changes in global ocean carbonate chemistry, resulting in a reduction in pH, a process termed "ocean acidification." It is important to determine which species are sensitive to elevated levels of CO2 because of potential impacts to ecosystems, marine resources, biodiversity, food webs, populations, and effects on economies. Previous studies with marine fish have documented that exposure to elevated levels of CO2 caused increased growth and larger otoliths in some species. This study was conducted to determine whether the elevated partial pressure of CO2 (pCO2) would have an effect on growth, otolith (ear bone) condition, survival, or the skeleton of juvenile scup, Stenotomus chrysops, a species that supports both important commercial and recreational fisheries. Elevated levels of pCO2 (1200-2600 µatm) had no statistically significant effect on growth, survival, or otolith condition after 8 weeks of rearing. Field data show that in Long Island Sound, where scup spawn, in situ levels of pCO2 are already at levels ranging from 689 to 1828 µatm due to primary productivity, microbial activity, and anthropogenic inputs. These results demonstrate that ocean acidification is not likely to cause adverse effects on the growth and survivability of every species of marine fish. X-ray analysis of the fish revealed a slightly higher incidence of hyperossification in the vertebrae of a few scup from the highest treatments compared to fish from the control treatments. Our results show that juvenile scup are tolerant to increases in seawater pCO2, possibly due to conditions this species encounters in their naturally variable environment and their well-developed pH control mechanisms.

Impacts of groundwater discharge at Myora Springs (North Stradbroke Island, Australia) on the phenolic metabolism of eelgrass, Zostera muelleri, and grazing by the juvenile rabbitfish, Siganus fuscescens

Myora Springs is one of many groundwater discharge sites on North Stradbroke Island (Queensland, Australia). Here spring waters emerge from wetland forests to join Moreton Bay, mixing with seawater over seagrass meadows dominated by eelgrass, Zostera muelleri. We sought to determine how low pH / high CO2 conditions near the spring affect these plants and their interactions with the black rabbitfish (Siganus fuscescens), a co-occurring grazer. In paired-choice feeding trials S. fuscescens preferentially consumed Z. muelleri shoots collected nearest to Myora Springs. Proximity to the spring did not significantly alter the carbon and nitrogen contents of seagrass tissues but did result in the extraordinary loss of soluble phenolics, including Folin-reactive phenolics, condensed tannins, and phenolic acids by ?87%. Conversely, seagrass lignin contents were, in this and related experiments, unaffected or increased, suggesting a shift in secondary metabolism away from the production of soluble, but not insoluble, (poly)phenolics. We suggest that groundwater discharge sites such as Myora Springs, and other sites characterized by low pH, are likely to be popular feeding grounds for seagrass grazers seeking to reduce their exposure to soluble phenolics.

Geographical variation in shell morphology of juvenile snails (Concholepas concholepas) along the physical-chemical gradient of the Chilean coast

Changes in phenotypic traits, such as mollusc shells, are indicative of variations in selective pressure along environmental gradients. Recently, increased sea surface temperature (SST) and ocean acidification (OA) due to increased levels of carbon dioxide in the seawater have been described as selective agents that may affect the biological processes underlying shell formation in calcifying marine organisms. The benthic snail Concholepas concholepas (Muricidae) is widely distributed along the Chilean coast, and so is naturally exposed to a strong physical-chemical latitudinal gradient. In this study, based on elliptical Fourier analysis, we assess changes in shell morphology (outlines analysis) in juvenile C. concholepas collected at northern (23°S), central (33°S) and southern (39°S) locations off the Chilean coast. Shell morphology of individuals collected in northern and central regions correspond to extreme morphotypes, which is in agreement with both the observed regional differences in the shell apex outlines, the high reclassification success of individuals (discriminant function analysis) collected in these regions, and the scaling relationship in shell weight variability among regions. However, these extreme morphotypes showed similar patterns of mineralization of calcium carbonate forms (calcite and aragonite). Geographical variability in shell shape of C. concholepas described by discriminant functions was partially explained by environmental variables (pCO2, SST). This suggests the influence of corrosive waters, such as upwelling and freshwaters penetrating into the coastal ocean, upon spatial variation in shell morphology. Changes in the proportion of calcium carbonate forms precipitated by C. concholepas across their shells and its susceptibility to corrosive coastal waters are discussed.

Mixed effects of elevated pCO2 on fertilisation, larval and juvenile development and adult responses in the mobile subtidal scallop Mimachlamys asperrima (Lamarck, 1819)

Ocean acidification is predicted to have severe consequences for calcifying marine organisms especially molluscs. Recent studies, however, have found that molluscs in marine environments with naturally elevated or fluctuating CO2 or with an active, high metabolic rate lifestyle may have a capacity to acclimate and be resilient to exposures of elevated environmental pCO2. The aim of this study was to determine the effects of near future concentrations of elevated pCO2 on the larval and adult stages of the mobile doughboy scallop, Mimachlamys asperrima from a subtidal and stable physio-chemical environment. It was found that fertilisation and the shell length of early larval stages of M. asperrima decreased as pCO2 increased, however, there were less pronounced effects of elevated pCO2 on the shell length of later larval stages, with high pCO2 enhancing growth in some instances. Byssal attachment and condition index of adult M. asperrima decreased with elevated pCO2, while in contrast there was no effect on standard metabolic rate or pHe. The responses of larval and adult M. asperrima to elevated pCO2 measured in this study were more moderate than responses previously reported for intertidal oysters and mussels. Even this more moderate set of responses are still likely to reduce the abundance of M. asperrima and potentially other scallop species in the world's oceans at predicted future pCO2 levels.

Seawater carbonate chemistry, thickness and carbonate elemental composition of the test of juvenile sea urchins in a laboratory experiment

Continuous anthropogenic CO2 emissions to the atmosphere and uptake by the oceans will cause a reduction of seawater pH and saturation state (Omega) of CaCO3 minerals from which marine calcifiers build their shells and skeletons. Sea urchins use the most soluble form of calcium carbonate, high-magnesium calcite, to build their skeleton, spines and grazing apparatus. In order to highlight the effects of increased pCO2 on the test thickness and carbonate elemental composition of juvenile sea urchins and potential differences in their responses linked to the diet, we performed a laboratory experiment on juvenile Paracentrotus lividus, grazing on calcifying (Corallina elongata) and non-calcifying (Cystoseira amentacea, Dictyota dichotoma) macroalgae, under different pH (corresponding to pCO2 values of 390, 550, 750 and 1000 µatm). Results highlighted the importance of the diet in determining sea urchin size irrespectively of the pCO2 level, and the relevance of macroalgal diet in modulating urchin Mg/Ca ratio. The present study provides relevant clues both in terms of the mechanism of mineral incorporation and in terms of bottom-up processes (algal diet) affecting top-down ones (fish predation) in rocky subtidal communities