Metabolic responses to temperature stress under elevated pCO2 in Crepidula fornicata

In the current context of environmental change, ocean acidification is predicted to affect the cellular processes, physiology and behaviour of all marine organisms, impacting survival, growth and reproduction. In relation to thermal tolerance limits, the effects of elevated pCO2 could be expected to be more pronounced at the upper limits of the thermal tolerance window. Our study focused on Crepidula fornicata, an invasive gastropod which colonized shallow waters around European coasts during the 20th century. We investigated the effects of 10 weeks' exposure to current (380 µatm) and elevated (550, 750, 1,000 µatm) pCO2 on this engineer species using an acute temperature increase (1 °C/12 h) as the test. Respiration rates were measured on both males (small individuals) and females (large individuals). Mortality increased suddenly from 34 °C, particularly in females. Respiration rate in C. fornicata increased linearly with temperature between 18 and 34 °C, but no differences were detected between the different pCO2 conditions either in the regressions between respiration rate and temperature or in Q10 values. In the same way, condition indices were similar in all the pCO2 treatments at the end of the experiment, but decreased from the beginning of the experiment. This species was highly resistant to acute exposure to high temperature regardless of pCO2 levels, even though food was limited during the experiment. Crepidula fornicata appears to have either developed resistance mechanisms or a strong phenotypic plasticity to deal with fluctuations of physicochemical parameters in its habitat. This suggests that invasive species may be more resistant to future environmental changes than its native competitors.

Rates of egg production, grazing and mortatlity of Calanus finmarchicus measured experimentally in the North Atlantic during the Maria S. Merian cruise MSM26, spring 2013

An incubation experiment at five different temperatures was used to assess the potential for adaptation of Calanus finmarchicus to future warming of the ocean. During a short term (3 h) and long term (6 day) exposure of individual females to a gradient of temperature stress, egg production and fecal pellet production were monitored to indicate secondary production and grazing rates. A longer term (10 day) exposure to elevated temperatures followed by a return to ambient sea temperatures was used to assess the potential recovery of individuals exposed to temperature stress. Females were picked out from WP2 net samples and acclimatised in 2 L bottles of GFF filtered seawater with Thalassiosira weissflogii as prey for >48 h at ambient SST. Experimental bottles were filled with filtered seawater (GFF filtered from non-toxic seawater supply) and acclimated to experimental temperature overnight (0, 5, 10, 15 and 20 °C). Individual females were transferred into bottles using forceps and the bottles were inoculated with T. weissflogii to a final concentration of 5 µg chl L-1. Bottles were then placed into water baths and incubated for 3h or 6 d, and monitored for egg and fecal pellet production rates. A 10 day exposure experiment was used to test the potential for recovery from temperature stress, by returning females incubated at 5, 10, 15 and 20 °C back to 10 °C for 24 h and counting egg and fecal pellet production.

Colony-specific investigations reveal highly variable responses among individual corals to ocean acidification and warming

As anthropogenic climate change is an ongoing concern, scientific investigations on its impacts on coral reefs are increasing. Although impacts of combined ocean acidification (OA) and temperature stress (T) on reef-building scleractinian corals have been studied at the genus, species and population levels, there are little data available on how individual corals respond to combined OA and anomalous temperatures. In this study, we exposed individual colonies of Acropora digitifera, Montipora digitata and Porites cylindrica to four pCO2-temperature treatments including 400 µatm-28 °C, 400 µatm-31 °C, 1000 µatm-28 °C and 1000 µatm-31 °C for 26 days. Physiological parameters including calcification, protein content, maximum photosynthetic efficiency, Symbiodinium density, and chlorophyll content along with Symbiodinium type of each colony were examined. Along with intercolonial responses, responses of individual colonies versus pooled data to the treatments were investigated. The main results were: 1) responses to either OA or T or their combination were different between individual colonies when considering physiological functions; 2) tolerance to either OA or T was not synonymous with tolerance to the other parameter; 3) tolerance to both OA and T did not necessarily lead to tolerance of OA and T combined (OAT) at the same time; 4) OAT had negative, positive or no impacts on physiological functions of coral colonies; and 5) pooled data were not representative of responses of all individual colonies. Indeed, the pooled data obscured actual responses of individual colonies or presented a response that was not observed in any individual. From the results of this study we recommend improving experimental designs of studies investigating physiological responses of corals to climate change by complementing them with colony-specific examinations.

The effects of thermal and high-CO2 stresses on the metabolism and surrounding microenvironment of the coral Galaxea fascicularis

The effects of elevated temperature and high pCO2 on the metabolism of Galaxea fascicularis were studied with oxygen and pH microsensors. Photosynthesis and respiration rates were evaluated from the oxygen fluxes from and to the coral polyps. High-temperature alone lowered both photosynthetic and respiration rates. High pCO2 alone did not significantly affect either photosynthesis or respiration rates. Under a combination of high-temperature and high-CO2, the photosynthetic rate increased to values close to those of the controls. The same pH in the diffusion boundary layer was observed under light in both (400 and 750 ppm) CO2 treatments, but decreased significantly in the dark as a result of increased CO2. The ATP contents decreased with increasing temperature. The effects of temperature on the metabolism of corals were stronger than the effects of increased CO2. The effects of acidification were minimal without combined temperature stress. However, acidification combined with higher temperature may affect coral metabolism due to the amplification of diel variations in the microenvironment surrounding the coral and the decrease in ATP contents.

Stable isotope ratios of planktonic foraminifera from North Atlantic surface sediments

We measured the oxygen isotopic composition of the deep-dwelling foraminiferal species G. inflata, G. truncatulinoides dextral and sinistral, and P. obliquiloculata in 29 modern core tops raised from the North Atlantic Ocean. We compared calculated isotopic temperatures with atlas temperatures and defined ecological models for each species. G. inflata and G. truncatulinoides live preferentially at the base of the seasonal thermocline. Under temperature stress, i.e., when the base of the seasonal thermocline is warmer than 16°C, G. inflata and G. truncatulinoides live deeper in the main thermocline. P. obliquiloculata inhabits the seasonal thermocline in warm regions. We tested our model using 10 cores along the Mauritanian upwelling and show that the comparison of d18O variations registered by the surficial species G. ruber and G. bulloides and the deep-dwelling species G. inflata evidences significant glacial-interglacial shifts of the Mauritanian upwelling cells.

Physiological and ecological variables measured at the high and low pCO2 reef sections

Experiments have shown that ocean acidification due to rising atmospheric carbon dioxide concentrations has deleterious effects on the performance of many marine organisms. However, few empirical or modelling studies have addressed the long-term consequences of ocean acidification for marine ecosystems. Here we show that as pH declines from 8.1 to 7.8 (the change expected if atmospheric carbon dioxide concentrations increase from 390 to 750 ppm, consistent with some scenarios for the end of this century) some organisms benefit, but many more lose out. We investigated coral reefs, seagrasses and sediments that are acclimatized to low pH at three cool and shallow volcanic carbon dioxide seeps in Papua New Guinea. At reduced pH, we observed reductions in coral diversity, recruitment and abundances of structurally complex framework builders, and shifts in competitive interactions between taxa. However, coral cover remained constant between pH 8.1 and ~7.8, because massive Porites corals established dominance over structural corals, despite low rates of calcification. Reef development ceased below pH 7.7. Our empirical data from this unique field setting confirm model predictions that ocean acidification, together with temperature stress, will probably lead to severely reduced diversity, structural complexity and resilience of Indo-Pacific coral reefs within this century.

Area-averaged mean seasonal cycles of chlorophyll-a concentration, sea surface temperature, alongshore wind stress, westward wind speed and dust component of the aerosol optical depth at 550 nm

Nutrient supply in the area off Northwest Africa is mainly regulated by two processes, coastal upwelling and deposition of Saharan dust. In the present study, both processes were analyzed and evaluated by different methods, including cross-correlation, multiple correlation, and event statistics, using remotely sensed proxies of the period from 2000 to 2008 to investigate their influence on the marine environment. The remotely sensed chlorophyll-a concentration was used as a proxy for the phytoplankton biomass stimulated by nutrient supply into the euphotic zone from deeper water layers and from the atmosphere. Satellite-derived alongshore wind stress and sea-surface temperature were applied as proxies for the strength and reflection of coastal upwelling processes. The westward wind and the dust component of the aerosol optical depth describe the transport direction of atmospheric dust and the atmospheric dust column load. Alongshore wind stress and induced upwelling processes were most significantly responsible for the surface chlorophyll-a variability, accounting for about 24% of the total variance, mainly in the winter and spring due to the strong north-easterly trade winds. The remotely sensed proxies allowed determination of time lags between biological response and its forcing processes. A delay of up to 16 days in the surface chlorophyll-a concentration due to the alongshore wind stress was determined in the northern winter and spring. Although input of atmospheric iron by dust storms can stimulate new phytoplankton production in the study area, only 5% of the surface chlorophyll-a variability could be ascribed to the dust component in the aerosol optical depth. All strong desert storms were identified by an event statistics in the time period from 2000 to 2008. The 57 strong storms were studied in relation to their biological response. Six events were clearly detected in which an increase of chlorophyll-a was caused by Saharan dust input and not by coastal upwelling processes. Time lags of <8 days, 8 days, and 16 days were determined. An increase in surface chlorophyll-a concentration of up to 2.4 mg m**3 after dust storms in which the dust component of the aerosol optical depth was up to 0.9 was observed.

Effects of seawater pCO2 and temperature on shell growth, shell stability, condition and cellular stress of Western Baltic Sea Mytilus edulis (L.) and Arctica islandica (L.)

Acidification of the World's oceans may directly impact reproduction, performance and shell formation of marine calcifying organisms. In addition, since shell production is costly and stress in general draws on an organism's energy budget, shell growth and stability of bivalves should indirectly be affected by environmental stress. The aim of this study was to investigate whether a combination of warming and acidification leads to increased physiological stress (lipofuscin accumulation and mortality) and affects the performance [shell growth, shell breaking force, condition index (Ci)] of young Mytilus edulis and Arctica islandica from the Baltic Sea. We cultured the bivalves in a fully-crossed 2-factorial experimental setup (seawater (sw) pCO2 levels "low", "medium" and "high" for both species, temperature levels 7.5, 10, 16, 20 and 25 °C for M. edulis and 7.5, 10 and 16 °C for A. islandica) for 13 weeks in summer. Mytilus edulis and A. islandica appeared to tolerate wide ranges of sw temperature and pCO2. Lipofuscin accumulation of M. edulis increased with temperature while the Ci decreased, but shell growth of the mussels only sharply decreased while its mortality increased between 20 and 25 °C. In A. islandica, lipofuscin accumulation increased with temperature, whereas the Ci, shell growth and shell breaking force decreased. The pCO2 treatment had only marginal effects on the measured parameters of both bivalve species. Shell growth of both bivalve species was not impaired by under-saturation of the sea water with respect to aragonite and calcite. Furthermore, independently of water temperatures shell breaking force of both species and shell growth of A. islandica remained unaffected by the applied elevated sw pCO2 for several months. Only at the highest temperature (25 °C), growth arrest of M. edulis was recorded at the high sw pCO2 treatment and the Ci of M. edulis was slightly higher at the medium sw pCO2 treatment than at the low and high sw pCO2 treatments. The only effect of elevated sw pCO2 on A. islandica was an increase in lipofuscin accumulation at the high sw pCO2 treatment compared to the medium sw pCO2 treatment. Our results show that, despite this robustness, growth of both M. edulis and A. islandica can be reduced if sw temperatures remain high for several weeks in summer. As large body size constitutes an escape from crab and sea star predation, this can make bivalves presumably more vulnerable to predation with possible negative consequences on population growth. In M. edulis, but not in A. islandica, this effect is amplified by elevated sw pCO2. We follow that combined effects of elevated sw pCO2 and ocean warming might cause shifts in future Western Baltic Sea community structures and ecosystem services; however, only if predators or other interacting species do not suffer as strong from these stressors.

Population and life-stage specific sensitivities to temperature and salinity stress in barnacles

Temperature and salinity shape the distribution and genetic structure of marine communities. Future warming and freshening will exert an additional stress to coastal marine systems. The extent to which organisms respond to these shifts will, however, be mediated by the tolerances of all life-stages and populations of species and their potential to adapt. We investigated nauplius and cypris larvae of the barnacle Balanus (Amphibalanus) improvisus from the Swedish west coast with respect to temperature (12, 20, and 28 °C) and salinity (5, 15, and 30) tolerances. Warming accelerated larval development and increased overall survival and subsequent settlement success. Nauplii developed and metamorphosed best at intermediate salinity. This was also observed in cypris larvae when the preceding nauplii stages had been reared at a salinity of 30. Direct comparisons of the present findings with those on a population from the more brackish Baltic Sea demonstrate contrasting patterns. We conclude that i) B. improvisus larvae within the Baltic region will be favoured by near-future seawater warming and freshening, that ii) salinity tolerances of larvae from the two different populations reflect salinities in their native habitats, but are nonetheless suboptimal and that iii) this species is generally highly plastic with regard to salinity.

Impact of thermal stress on evolutionary trajectories of pathogen resistance in three-spined stickleback (Gasterosteus aculeatus), supplementary material

Background: Pathogens are a major regulatory force for host populations, especially under stressful conditions. Elevated temperatures may enhance the development of pathogens, increase the number of transmission stages, and can negatively influence host susceptibility depending on host thermal tolerance. As a net result, this can lead to a higher prevalence of epidemics during summer months. These conditions also apply to marine ecosystems, where possible ecological impacts and the population-specific potential for evolutionary responses to changing environments and increasing disease prevalence are, however, less known. Therefore, we investigated the influence of thermal stress on the evolutionary trajectories of disease resistance in three marine populations of three-spined sticklebacks Gasterosteus aculeatus by combining the effects of elevated temperature and infection with a bacterial strain of Vibrio sp. using a common garden experiment. Results: We found that thermal stress had an impact on fish weight and especially on survival after infection after only short periods of thermal acclimation. Environmental stress reduced genetic differentiation (QST) between populations by releasing cryptic within-population variation. While life history traits displayed positive genetic correlations across environments with relatively weak genotype by environment interactions (GxE), environmental stress led to negative genetic correlations across environments in pathogen resistance. This reversal of genetic effects governing resistance is probably attributable to changing environment-dependent virulence mechanisms of the pathogen interacting differently with host genotypes, i.e. GPathogenxGHostxE or (GPathogenxE)x(GHostxE) interactions, rather than to pure host genetic effects, i.e. GHostxE interactions. Conclusion: To cope with climatic changes and the associated increase in pathogen virulence, host species require wide thermal tolerances and pathogen-resistant genotypes. The higher resistance we found for some families at elevated temperatures showed that there is evolutionary potential for resistance to Vibrio sp. in both thermal environments. The negative genetic correlation of pathogen resistance between thermal environments, on the other hand, indicates that adaptation to current conditions can be a weak predictor for performance in changing environments. The observed feedback on selective gradients exerted on life history traits may exacerbate this effect, as it can also modify the response to selection for other vital components of fitness.

Diurnal and seasonal measurements of seawater chemistry, temperature and PAR in Hurricane Hole, U.S. Virgin Islands - 2010-2012

Risk analyses indicate that more than 90% of the world's reefs will be threatened by climate change and local anthropogenic impacts by the year 2030 under "business-as-usual" climate scenarios. Increasing temperatures and solar radiation cause coral bleaching that has resulted in extensive coral mortality. Increasing carbon dioxide reduces seawater pH, slows coral growth, and may cause loss of reef structure. Management strategies include establishment of marine protected areas with environmental conditions that promote reef resiliency. However, few resilient reefs have been identified, and resiliency factors are poorly defined. Here we characterize the first natural, non-reef coral refuge from thermal stress and ocean acidification and identify resiliency factors for mangrove-coral habitats. We measured diurnal and seasonal variations in temperature, salinity, photosynthetically active radiation (PAR), and seawater chemistry; characterized substrate parameters; and examined water circulation patterns in mangrove communities where scleractinian corals are growing attached to and under mangrove prop roots in Hurricane Hole, St. John, US Virgin Islands. Additionally, we inventoried the coral species and quantified incidences of coral bleaching, mortality, and recovery for two major reef-building corals, Colpophyllia natans and Diploria labyrinthiformis, growing in mangrove-shaded and exposed (unshaded) areas. Over 30 species of scleractinian corals were growing in association with mangroves. Corals were thriving in low-light (more than 70% attenuation of incident PAR) from mangrove shading and at higher temperatures than nearby reef tract corals. A higher percentage of C. natans colonies were living shaded by mangroves, and no shaded colonies were bleached. Fewer D. labyrinthiformis colonies were shaded by mangroves, however more unshaded colonies were bleached. A combination of substrate and habitat heterogeneity, proximity of different habitat types, hydrographic conditions, and biological influences on seawater chemistry generate chemical conditions that buffer against ocean acidification. This previously undocumented refuge for corals provides evidence for adaptation of coastal organisms and ecosystem transition due to recent climate change. Identifying and protecting other natural, non-reef coral refuges is critical for sustaining corals and other reef species into the future.

Table 1. Pressure, temperature and isotope ratios between H2O and CO2 for all experiments

Traditionally, the application of stable isotopes in Carbon Capture and Storage (CCS) projects has focused on d13C values of CO2 to trace the migration of injected CO2 in the subsurface. More recently the use of d18O values of both CO2 and reservoir fluids has been proposed as a method for quantifying in situ CO2 reservoir saturations due to O isotope exchange between CO2 and H2O and subsequent changes in d18OH2O values in the presence of high concentrations of CO2. To verify that O isotope exchange between CO2 and H2O reaches equilibrium within days, and that d18OH2O values indeed change predictably due to the presence of CO2, a laboratory study was conducted during which the isotope composition of H2O, CO2, and dissolved inorganic C (DIC) was determined at representative reservoir conditions (50°C and up to 19 MPa) and varying CO2 pressures. Conditions typical for the Pembina Cardium CO2 Monitoring Pilot in Alberta (Canada) were chosen for the experiments. Results obtained showed that d18O values of CO2 were on average 36.4±2.2 per mil (1 sigma, n=15) higher than those of water at all pressures up to and including reservoir pressure (19 MPa), in excellent agreement with the theoretically predicted isotope enrichment factor of 35.5 per mil for the experimental temperatures of 50°C. By using 18O enriched water for the experiments it was demonstrated that changes in the d18O values of water were predictably related to the fraction of O in the system sourced from CO2 in excellent agreement with theoretical predictions. Since the fraction of O sourced from CO2 is related to the total volumetric saturation of CO2 and water as a fraction of the total volume of the system, it is concluded that changes in d18O values of reservoir fluids can be used to calculate reservoir saturations of CO2 in CCS settings given that the d18O values of CO2 and water are sufficiently distinct.

Temperature tolerance of different larval stages of the spider crab Hyas araneus exposed to elevated seawater PCO2

Exposure to elevated seawater PCO2 limits the thermal tolerance of crustaceans but the underlying mechanisms have not been comprehensively explored. Larval stages of crustaceans are even more sensitive to environmental hypercapnia and possess narrower thermal windows than adults. In a mechanistic approach, we analysed the impact of high seawater CO2 on parameters at different levels of biological organization, from the molecular to the whole animal level. At the whole animal level we measured oxygen consumption, heart rate and activity during acute warming in zoea and megalopa larvae of the spider crab Hyas araneus exposed to different levels of seawater PCO2. Furthermore, the expression of genes responsible for acid-base regulation and mitochondrial energy metabolism, and cellular responses to thermal stress (e.g. the heat shock response) was analysed before and after larvae were heat shocked byrapidly raising the seawater temperature from 10°C rearing temperature to 20°C. Zoea larvae showed a high heat tolerance, which decreased at elevated seawater PCO2, while the already low heat tolerance of megalopa larvae was not limited further by hypercapnic exposure. There was a combined effect of elevated seawater CO2 and heat shock in zoea larvae causing elevated transcript levels of heat shock proteins. In all three larval stages, hypercapnic exposure elicited an up-regulation of genes involved in oxidative phosphorylation, which was, however, not accompanied by increased energetic demands. The combined effect of seawater CO2 and heat shock on the gene expression of heat shock proteins reflects the downward shift in thermal limits seen on the whole animal level and indicates an associated capacity to elicit passive thermal tolerance. The up-regulation of genes involved in oxidative phosphorylation might compensate for enzyme activities being lowered through bicarbonate inhibition and maintain larval standard metabolic rates at high seawater CO2 levels. The present study underlines the necessity to align transcriptomic data with physiological responses when addressing mechanisms affected by an interaction of elevated seawater PCO2 and temperature extremes.