Combined effects of short-term exposure to elevated CO2 and decreased O2 on the physiology and energy budget of the thick shell mussel Mytilus coruscus

Hypoxia and ocean acidification are two consequences of anthropogenic activities. These global trends occur on top of natural variability. In environments such as estuarine areas, short-term acute pH and O2 fluctuations are occurring simultaneously. The present study tested the combined effects of short-term seawater acidification and hypoxia on the physiology and energy budget of the thick shell mussel Mytilus coruscus. Mussels were exposed for 72 h to six combined treatments with three pH levels (8.1, 7.7 and 7.3) and two dissolved oxygen (DO) levels (2 mg/L, 6 mg/L). Clearance rate (CR), food absorption efficiency (AE), respiration rate (RR), ammonium excretion rate (ER), O:N ratio and scope for growth (SFG) were significantly reduced, and faecal organic dry weight ratio (E) was significantly increased at low DO. Low pH did not lead to a reduced SFG. Interactive effects of pH and DO were observed for CR, E and RR. Principal component analysis (PCA) revealed positive relationships among most physiological indicators, especially between SFG and CR under normal DO conditions. These results demonstrate that Mytilus coruscus was sensitive to short-term (72 h) exposure to decreased O2 especially if combined with decreased pH levels. In conclusion, the short-term oxygen and pH variation significantly induced physiological changes of mussels with some interactive effects.

Coral energy reserves and calcification in a high-CO2 world at two temperatures

Rising atmospheric CO2 concentrations threaten coral reefs globally by causing ocean acidification (OA) and warming. Yet, the combined effects of elevated pCO2 and temperature on coral physiology and resilience remain poorly understood. While coral calcification and energy reserves are important health indicators, no studies to date have measured energy reserve pools (i.e., lipid, protein, and carbohydrate) together with calcification under OA conditions under different temperature scenarios. Four coral species, Acropora millepora, Montipora monasteriata, Pocillopora damicornis, Turbinaria reniformis, were reared under a total of six conditions for 3.5 weeks, representing three pCO2 levels (382, 607, 741 µatm), and two temperature regimes (26.5, 29.0°C) within each pCO2 level. After one month under experimental conditions, only A. millepora decreased calcification (-53%) in response to seawater pCO2 expected by the end of this century, whereas the other three species maintained calcification rates even when both pCO2 and temperature were elevated. Coral energy reserves showed mixed responses to elevated pCO2 and temperature, and were either unaffected or displayed nonlinear responses with both the lowest and highest concentrations often observed at the mid-pCO2 level of 607 µatm. Biweekly feeding may have helped corals maintain calcification rates and energy reserves under these conditions. Temperature often modulated the response of many aspects of coral physiology to OA, and both mitigated and worsened pCO2 effects. This demonstrates for the first time that coral energy reserves are generally not metabolized to sustain calcification under OA, which has important implications for coral health and bleaching resilience in a high-CO2 world. Overall, these findings suggest that some corals could be more resistant to simultaneously warming and acidifying oceans than previously expected.

Differential impacts of elevated CO2 and acidosis on the energy budget of gill and liver cells from Atlantic cod

Ocean acidification impacts fish and other marine species through increased seawater PCO2 levels (hypercapnia). Knowledge of the physiological mechanisms mediating effects in various tissues of fish is incomplete. Here we tested the effects of extracellular hypercapnia and acidosis on energy metabolism of gill and liver cells of Atlantic cod. Exposure media mimicked blood conditions in vivo, either during normo- or hypercapnia and at control or acidic extracellular pH (pHe). We determined metabolic rate and energy expenditure for protein biosynthesis, Na+/K+-ATPase and H+-ATPase and considered nutrition status by measurements of metabolic rate and protein biosynthesis in media with and without free amino acids (FAA). Addition of FAA stimulated hepatic but not branchial oxygen consumption. Normo- and hypercapnic acidosis as well as hypercapnia at control pHe depressed metabolic stimulation of hepatocytes. In gill cells, acidosis depressed respiration independent of PCO2 and FAA levels. For both cell types, depressed respiration was not correlated with the same reduction in energy allocated to protein biosynthesis or Na+/K+-ATPase. Hepatic energy expenditure for protein synthesis and Na+/K+- ATPase was even elevated at acidic compared to control pHe suggesting increased costs for ion regulation and cel- lular reorganization. Hypercapnia at control pHe strongly reduced oxygen demand of branchial Na+/K+-ATPase with a similar trend for H+-ATPase. We conclude that extracellular acidosis triggers metabolic depression in gill and metabolically stimulated liver cells. Additionally, hypercapnia itself seems to limit capacities for metabolic usage of amino acids in liver cells while it decreases the use and costs of ion regulatory ATPases in gill cells.

Physiological advantages of dwarfing in surviving extinctions in high-CO2 oceans

Excessive CO2 in the present-day ocean-atmosphere system is causing ocean acidification, and is likely to cause a severe biodiversity decline in the future, mirroring effects in many past mass extinctions. Fossil records demonstrate that organisms surviving such events were often smaller than those before, a phenomenon called the Lilliput effect. Here, we show that two gastropod species adapted to acidified seawater at shallow-water CO2 seeps were smaller than those found in normal pH conditions and had higher mass-specific energy consumption but significantly lower whole-animal metabolic energy demand. These physiological changes allowed the animals to maintain calcification and to partially repair shell dissolution. These observations of the long-term chronic effects of increased CO2 levels forewarn of changes we can expect in marine ecosystems as CO2 emissions continue to rise unchecked, and support the hypothesis that ocean acidification contributed to past extinction events. The ability to adapt through dwarfing can confer physiological advantages as the rate of CO2 emissions continues to increase.

Microalgal CO2 Sequestering - Modeling Microalgae Production Costs

Microalgae CO2 sequestering facilities might become an industrial reality if microalgae biomass could be produced at cost below $500.00 t-1. We develop a model for estimation of total production costs of microalgae as a function of known production-specific expenses, and incorporate into the model the effects of uncontrollable factors which affect known production-specific expenses. Random fluctuations were intentionally incorporated into the model, consequently into generated cost/technology scenarios, because each and every logically interconnected equipment/operation that is used in design/construction/operation/maintenance of a production process is inevitably subject to random cost/price fluctuations which can neither be eliminated nor a priori controlled. A total of 152 costs/technology scenarios were evaluated to find forty four scenarios in which Predicted Total Production Costs of Microalgae (PTPCM) was in the range $200 to $500 t-1 ha-1 y-1. An additional 24 scenarios were found with PTCPM in the range of $102 to $200 t-1 ha-1 y-1. These findings suggest that microalgae CO2 sequestering and the production of commercial compounds from microalgal biomass can be economically viable venture even today when microalgae production technology is still far from its optimum.

Increased CO2 modifies the carbon balance and the photosynthetic yield of two common Arctic brown seaweeds: Desmarestia aculeata and Alaria esculenta

Ocean acidification affects with special intensity Arctic ecosystems, being marine photosynthetic organisms a primary target, although the consequences of this process in the carbon fluxes of Arctic algae are still unknown. The alteration of the cellular carbon balance due to physiological acclimation to an increased CO2 concentration (1300 ppm) in the common Arctic brown seaweeds Desmarestia aculeata and Alaria esculenta from Kongsfjorden (Svalbard) was analysed. Growth rate of D. aculeata was negatively affected by CO2 enrichment, while A. esculenta was positively affected, as a result of a different reorganization of the cellular carbon budget in both species. Desmarestia aculeata showed increased respiration, enhanced accumulation of storage biomolecules and elevated release of dissolved organic carbon, whereas A. esculenta showed decreased respiration and lower accumulation of storage biomolecules. Gross photosynthesis (measured both as O2 evolution and 14C fixation) was not affected in any of them, suggesting that photosynthesis was already saturated at normal CO2 conditions and did not participate in the acclimation response. However, electron transport rate changed in both species in opposite directions, indicating different energy requirements between treatments and species specificity. High CO2 levels also affected the N-metabolism, and 13C isotopic discrimination values from algal tissue pointed to a deactivation of carbon concentrating mechanisms. Since increased CO2 has the potential to modify physiological mechanisms in different ways in the species studied, it is expected that this may lead to changes in the Arctic seaweed community, which may propagate to the rest of the food web.

Elevated CO2 alters larval proteome and its phosphorylation status in the commercial oyster, Crassostrea hongkongensis

Ocean acidification (OA) is beginning to have noticeable negative impact on calcification rate, shell structure and physiological energy budgeting of several marine organisms; these alter the growth of many economically important shellfish including oysters. Early life stages of oysters may be particularly vulnerable to OA-driven low pH conditions because their shell is made up of the highly soluble form of calcium carbonate (CaCO3) mineral, aragonite. Our long-term CO2 perturbation experiment showed that larval shell growth rate of the oyster species Crassostrea hongkongensis was significantly reduced at pH < 7.9 compared to the control (8.2). To gain new insights into the underlying mechanisms of low-pH-induced delays in larval growth, we have examined the effect of pH on the protein expression pattern, including protein phosphorylation status at the pediveliger larval stage. Using two-dimensional electrophoresis and mass spectrometry, we demonstrated that the larval proteome was significantly altered by the two low pH treatments (7.9 and 7.6) compared to the control pH (8.2). Generally, the number of expressed proteins and their phosphorylation level decreased with low pH. Proteins involved in larval energy metabolism and calcification appeared to be down-regulated in response to low pH, whereas cell motility and production of cytoskeletal proteins were increased. This study on larval growth coupled with proteome change is the first step toward the search for novel Protein Expression Signatures indicative of low pH, which may help in understanding the mechanisms involved in low pH tolerance.

Coral-algae metabolism and diurnal changes in the CO2-carbonate system of bulk sea water

Precise measurements were conducted in continuous flow seawater mesocosms located in full sunlight that compared metabolic response of coral, coral-macroalgae and macroalgae systems over a diurnal cycle. Irradiance controlled net photosynthesis (Pnet), which in turn drove net calcification (Gnet), and altered pH. Pnet exerted the dominant control on [CO3]2- and aragonite saturation state (Omega arag) over the diel cycle. Dark calcification rate decreased after sunset, reaching zero near midnight followed by an increasing rate that peaked at 03:00 h. Changes in Omega arag and pH lagged behind Gnet throughout the daily cycle by two or more hours. The flux rate Pnet was the primary driver of calcification. Daytime coral metabolism rapidly removes dissolved inorganic carbon (DIC) from the bulk seawater and photosynthesis provides the energy that drives Gnet while increasing the bulk water pH. These relationships result in a correlation between Gnet and Omega arag, with Omega arag as the dependent variable. High rates of H+ efflux continued for several hours following mid-day peak Gnet suggesting that corals have difficulty in shedding waste protons as described by the Proton Flux Hypothesis. DIC flux (uptake) followed Pnet and Gnet and dropped off rapidly following peak Pnet and peak Gnet indicating that corals can cope more effectively with the problem of limited DIC supply compared to the problem of eliminating H+. Over a 24 h period the plot of total alkalinity (AT) versus DIC as well as the plot of Gnet versus Omega arag revealed a circular hysteresis pattern over the diel cycle in the coral and coral-algae mesocosms, but not the macroalgae mesocosm. Presence of macroalgae did not change Gnet of the corals, but altered the relationship between Omega arag and Gnet. Predictive models of how future global changes will effect coral growth that are based on oceanic Omega arag must include the influence of future localized Pnet on Gnet and changes in rate of reef carbonate dissolution. The correlation between Omega arag and Gnet over the diel cycle is simply the response of the CO2-carbonate system to increased pH as photosynthesis shifts the equilibria and increases the [CO3]2- relative to the other DIC components of [HCO3]- and [CO2]. Therefore Omega arag closely tracked pH as an effect of changes in Pnet, which also drove changes in Gnet. Measurements of DIC flux and H+ flux are far more useful than concentrations in describing coral metabolism dynamics. Coral reefs are systems that exist in constant disequilibrium with the water column.

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.

Effects of increased CO2 on fish gill and plasma proteome

Ocean acidification and warming are both primarily caused by increased levels of atmospheric CO2, and marine organisms are exposed to these two stressors simultaneously. Although the effects of temperature on fish have been investigated over the last century, the long-term effects of moderate CO2 exposure and the combination of both stressors are almost entirely unknown. A proteomics approach was used to assess the adverse physiological and biochemical changes that may occur from the exposure to these two environmental stressors. We analysed gills and blood plasma of Atlantic halibut (Hippoglossus hippoglossus) exposed to temperatures of 12°C (control) and 18°C (impaired growth) in combination with control (400 µatm) or high-CO2 water (1000 µatm) for 14 weeks. The proteomic analysis was performed using two-dimensional gel electrophoresis (2DE) followed by Nanoflow LC-MS/MS using a LTQ-Orbitrap. The high-CO2 treatment induced the up-regulation of immune system-related proteins, as indicated by the up-regulation of the plasma proteins complement component C3 and fibrinogen beta chain precursor in both temperature treatments. Changes in gill proteome in the high-CO2 (18°C) group were mostly related to increased energy metabolism proteins (ATP synthase, malate dehydrogenase, malate dehydrogenase thermostable, and fructose-1,6-bisphosphate aldolase), possibly coupled to a higher energy demand. Gills from fish exposed to high-CO2 at both temperature treatments showed changes in proteins associated with increased cellular turnover and apoptosis signalling (annexin 5, eukaryotic translation elongation factor 1 gamma, receptor for protein kinase C, and putative ribosomal protein S27). This study indicates that moderate CO2-driven acidification, alone and combined with high temperature, can elicit biochemical changes that may affect fish health.

Combined effects of different CO2 levels and N sources on the diazotrophic cyanobacterium Trichodesmium

To predict effects of climate change and possible feedbacks, it is crucial to understand the mechanisms behind CO2 responses of biogeochemically relevant phytoplankton species. Previous experiments on the abundant N2 fixers Trichodesmium demonstrated strong CO2 responses, which were attributed to an energy reallocation between its carbon (C) and nitrogen (N) acquisition. Pursuing this hypothesis, we manipulated the cellular energy budget by growing Trichodesmium erythraeum IMS101 under different CO2 partial pressure (pCO2) levels (180, 380, 980 and 1400?µatm) and N sources (N2 and NO3-). Subsequently, biomass production and the main energy-generating processes (photosynthesis and respiration) and energy-consuming processes (N2 fixation and C acquisition) were measured. While oxygen fluxes and chlorophyll fluorescence indicated that energy generation and its diurnal cycle was neither affected by pCO2 nor N source, cells differed in production rates and composition. Elevated pCO2 increased N2 fixation and organic C and N contents. The degree of stimulation was higher for nitrogenase activity than for cell contents, indicating a pCO2 effect on the transfer efficiency from N2 to biomass. pCO2-dependent changes in the diurnal cycle of N2 fixation correlated well with C affinities, confirming the interactions between N and C acquisition. Regarding effects of the N source, production rates were enhanced in NO3-grown cells, which we attribute to the higher N retention and lower ATP demand compared with N2 fixation. pCO2 effects on C affinity were less pronounced in NO3- users than N2 fixers. Our study illustrates the necessity to understand energy budgets and fluxes under different environmental conditions for explaining indirect effects of rising pCO2.

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.

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.