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

Amino acids in the interstitial waters from ODP Hole 113-695A

Site 695 lies on the southeast margin of the South Orkney microcontinent on the northern margin of the Weddell Sea, at 62°23.48'S, 43°27.10'W in 1305 m water depth. The inorganic properties of interstitial waters at this site, including sulfate reduction, biogenic methane production, and high concentrations of ammonia and phosphate, imply high microbial activity. However, no clear relationship between amino acid composition and concentration and the type of microbial activity (e.g., sulfate reduction or methane production) can be identified. The THAA (total hydrolyzable amino acids) values range between 2.45 and 17.31 µmol/L, averaging 7.14 µmol/L. The mean concentrations and relative abundance values of acidic, basic, neutral, aromatic, and sulfur-containing amino acids are 1.34 (18%), 1.09 (15%), 3.93 (54%), 0.50 (8%), and 0.02 (0%) µmol/L, respectively. Glycine is the most abundant amino acid residue, with serine, glutamic acid, and ornithine next. The DFAA (dissolved free amino acids) values range from 0.10 to 12.73 µmol/L, averaging 4.07 µmol/L. The acidic, basic, neutral, aromatic, and sulfurcontaining amino acids are on average 0.21, 0.79, 2.56, 0.41, and 0.01 µmol/L, respectively. The relative abundances of acidic, basic, neutral, and aromatic amino acids average 4%, 18%, 58%, and 15%, respectively. Predominance of DFAA over DCAA (dissolved combined amino acids) in interstitial waters of Lithologic Units I and II is contrary to the predominance of DCAA over DFAA in other interstitial waters and seawater. The comparison of amino acid compositions between DCAA and siliceous plankton suggests that the DCAA in interstitial waters originally comes from amino acids derived from siliceous plankton. However, other sources which are much enriched in glutamic acid contribute to the DCAA composition.

Biogeochemical measurements of sediments collected at station MSM15/1_492-PUC2 (reference site 1) in the Black Sea during the MSM15/1 cruise in 2010

Biogeochemical measurements in sediment cores collected with the submersible JAGO (pusch cores) and a TV-MUC in the Black Sea during MSM15/1, Northwest Crimea (HYPOX Project), at water depths between 152-156 m. A series of microbial mats were sampled on the hypoxic region of the Crimean Shelf. Concentrations of organic carbon (Corg) and nitrogen (N) were measured on finely powdered, freeze-dried subsamples of sediment using a using a Fisons NA-1500 elemental analyzer. For organic carbon determination samples were pre-treated with 12.5% HCl to remove carbonates. Chlorophyll a (chl a), phaeopigments (PHAEO) and chloroplastic pigment equivalents (CPE) was measured according to Schubert et al., (2005) and total hydrolyzable amino acids (THAA) and single amino acid: ASP, GLU, SER, HIS, GLY, THR, ARG, ALA, TYR, MET, VAL, PHE, ILE, LEU, LYS following Dauwe et al., 1998.

Biogeochemical measurements of sediments collected at station MSM15/1_377-1 (microbial mat 4) in the Black Sea during the MSM15/1 cruise in 2010

Biogeochemical measurements in sediment cores collected with the submersible JAGO (pusch cores) and a TV-MUC in the Black Sea during MSM15/1, Northwest Crimea (HYPOX Project), at water depths between 152-156 m. A series of microbial mats were sampled on the hypoxic region of the Crimean Shelf. Concentrations of organic carbon (Corg) and nitrogen (N) were measured on finely powdered, freeze-dried subsamples of sediment using a using a Fisons NA-1500 elemental analyzer. For organic carbon determination samples were pre-treated with 12.5% HCl to remove carbonates. Chlorophyll a (chl a), phaeopigments (PHAEO) and chloroplastic pigment equivalents (CPE) was measured according to Schubert et al., (2005) and total hydrolyzable amino acids (THAA) and single amino acid: ASP, GLU, SER, HIS, GLY, THR, ARG, ALA, TYR, MET, VAL, PHE, ILE, LEU, LYS following Dauwe et al., 1998.

Biogeochemical measurements of sediments collected at station MSM15/1_492-PUC3 (microbial mat 2) in the Black Sea during the MSM15/1 cruise in 2010

Biogeochemical measurements in sediment cores collected with the submersible JAGO (pusch cores) and a TV-MUC in the Black Sea during MSM15/1, Northwest Crimea (HYPOX Project), at water depths between 152-156 m. A series of microbial mats were sampled on the hypoxic region of the Crimean Shelf. Concentrations of organic carbon (Corg) and nitrogen (N) were measured on finely powdered, freeze-dried subsamples of sediment using a using a Fisons NA-1500 elemental analyzer. For organic carbon determination samples were pre-treated with 12.5% HCl to remove carbonates. Chlorophyll a (chl a), phaeopigments (PHAEO) and chloroplastic pigment equivalents (CPE) was measured according to Schubert et al., (2005) and total hydrolyzable amino acids (THAA) and single amino acid: ASP, GLU, SER, HIS, GLY, THR, ARG, ALA, TYR, MET, VAL, PHE, ILE, LEU, LYS following Dauwe et al., 1998.

Biogeochemical measurements of sediments collected at station MSM15/1_492-PUC5 (reference site 2) in the Black Sea during the MSM15/1 cruise in 2010

Biogeochemical measurements in sediment cores collected with the submersible JAGO (pusch cores) and a TV-MUC in the Black Sea during MSM15/1, Northwest Crimea (HYPOX Project), at water depths between 152-156 m. A series of microbial mats were sampled on the hypoxic region of the Crimean Shelf. Concentrations of organic carbon (Corg) and nitrogen (N) were measured on finely powdered, freeze-dried subsamples of sediment using a using a Fisons NA-1500 elemental analyzer. For organic carbon determination samples were pre-treated with 12.5% HCl to remove carbonates. Chlorophyll a (chl a), phaeopigments (PHAEO) and chloroplastic pigment equivalents (CPE) was measured according to Schubert et al., (2005) and total hydrolyzable amino acids (THAA) and single amino acid: ASP, GLU, SER, HIS, GLY, THR, ARG, ALA, TYR, MET, VAL, PHE, ILE, LEU, LYS following Dauwe et al., 1998.

Biogeochemical measurements of sediments collected at station MSM15/1_492-PUC9 (microbial mat 3) in the Black Sea during the MSM15/1 cruise in 2010

Biogeochemical measurements in sediment cores collected with the submersible JAGO (pusch cores) and a TV-MUC in the Black Sea during MSM15/1, Northwest Crimea (HYPOX Project), at water depths between 152-156 m. A series of microbial mats were sampled on the hypoxic region of the Crimean Shelf. Concentrations of organic carbon (Corg) and nitrogen (N) were measured on finely powdered, freeze-dried subsamples of sediment using a using a Fisons NA-1500 elemental analyzer. For organic carbon determination samples were pre-treated with 12.5% HCl to remove carbonates. Chlorophyll a (chl a), phaeopigments (PHAEO) and chloroplastic pigment equivalents (CPE) was measured according to Schubert et al., (2005) and total hydrolyzable amino acids (THAA) and single amino acid: ASP, GLU, SER, HIS, GLY, THR, ARG, ALA, TYR, MET, VAL, PHE, ILE, LEU, LYS following Dauwe et al., 1998.

Ex situ microsensor profiles of sulfide concentration and pH in/outside mat 1 (MSM15/1_492-PUC1) measured during the MSM15/1 cruise in 2010 (Black Sea)

Biogeochemical measurements in sediment cores collected with the submersible JAGO (pusch cores) and a TV-MUC in the Black Sea during MSM15/1, Northwest Crimea (HYPOX Project), at water depths between 152-156 m. A series of microbial mats were sampled on the hypoxic region of the Crimean Shelf. Concentrations of organic carbon (Corg) and nitrogen (N) were measured on finely powdered, freeze-dried subsamples of sediment using a using a Fisons NA-1500 elemental analyzer. For organic carbon determination samples were pre-treated with 12.5% HCl to remove carbonates. Chlorophyll a (chl a), phaeopigments (PHAEO) and chloroplastic pigment equivalents (CPE) was measured according to Schubert et al., (2005; doi:10.1029/2004GC000837) and total hydrolyzable amino acids (THAA) and single amino acid: ASP, GLU, SER, HIS, GLY, THR, ARG, ALA, TYR, MET, VAL, PHE, ILE, LEU, LYS following Dauwe et al., 1998. High-resolution ex situ sulfide and pH microprofiles, were assessed only for station MSM15/1_492_PUC1. "in mat 1, 2 and 3" refers to 3 different profiles in 3 different spots of the microbial mat, whereas "outside mat", a profile outside the microbial mat.

Biogeochemical investigation of microbial mats from the hypoxic region of the Crimean Shelf (Black Sea)

Biogeochemical measurements in sediment cores collected with the submersible JAGO (pusch cores) and a TV-MUC in the Black Sea during MSM15/1, Northwest Crimea (HYPOX Project), at water depths between 152-156 m. A series of microbial mats were sampled on the hypoxic region of the Crimean Shelf. Concentrations of organic carbon (Corg) and nitrogen (N) were measured on finely powdered, freeze-dried subsamples of sediment using a using a Fisons NA-1500 elemental analyzer. For organic carbon determination samples were pre-treated with 12.5% HCl to remove carbonates. Chlorophyll a (chl a), phaeopigments (PHAEO) and chloroplastic pigment equivalents (CPE) was measured according to Schubert et al., (2005) and total hydrolyzable amino acids (THAA) and single amino acid: ASP, GLU, SER, HIS, GLY, THR, ARG, ALA, TYR, MET, VAL, PHE, ILE, LEU, LYS following Dauwe et al., 1998. High-resolution ex situ sulfide and pH microprofiles, were assessed only for station MSM15/1_492_PUC1. "in mat 1, 2 and 3" refers to 3 different profiles in 3 different spots of the microbial mat, whereas "outside mat", a profile outside the microbial mat.

Biogeochemical measurements of sediments collected at station MSM15/1_492-PUC1 (microbial mat 1) in the Black Sea during the MSM15/1 cruise in 2010

Biogeochemical measurements in sediment cores collected with the submersible JAGO (pusch cores) and a TV-MUC in the Black Sea during MSM15/1, Northwest Crimea (HYPOX Project), at water depths between 152-156 m. A series of microbial mats were sampled on the hypoxic region of the Crimean Shelf. Concentrations of organic carbon (Corg) and nitrogen (N) were measured on finely powdered, freeze-dried subsamples of sediment using a using a Fisons NA-1500 elemental analyzer. For organic carbon determination samples were pre-treated with 12.5% HCl to remove carbonates. Chlorophyll a (chl a), phaeopigments (PHAEO) and chloroplastic pigment equivalents (CPE) was measured according to Schubert et al., (2005) and total hydrolyzable amino acids (THAA) and single amino acid: ASP, GLU, SER, HIS, GLY, THR, ARG, ALA, TYR, MET, VAL, PHE, ILE, LEU, LYS following Dauwe et al., 1998.

Biogeochemical measurements of sediments collected at station MSM15/1_492-PUC6 (reference site 3) in the Black Sea during the MSM15/1 cruise in 2010

Biogeochemical measurements in sediment cores collected with the submersible JAGO (pusch cores) and a TV-MUC in the Black Sea during MSM15/1, Northwest Crimea (HYPOX Project), at water depths between 152-156 m. A series of microbial mats were sampled on the hypoxic region of the Crimean Shelf. Concentrations of organic carbon (Corg) and nitrogen (N) were measured on finely powdered, freeze-dried subsamples of sediment using a using a Fisons NA-1500 elemental analyzer. For organic carbon determination samples were pre-treated with 12.5% HCl to remove carbonates. Chlorophyll a (chl a), phaeopigments (PHAEO) and chloroplastic pigment equivalents (CPE) was measured according to Schubert et al., (2005) and total hydrolyzable amino acids (THAA) and single amino acid: ASP, GLU, SER, HIS, GLY, THR, ARG, ALA, TYR, MET, VAL, PHE, ILE, LEU, LYS following Dauwe et al., 1998.

Seawater carbonate chemistry and resource allocation and extracellular acid-base status in the sea urchin Strongylocentrotus droebachiensis during experiments, 2012

Anthropogenic CO2 emission will lead to an increase in seawater pCO2 of up to 80-100 Pa (800-1000 µatm) within this century and to an acidification of the oceans. Green sea urchins (Strongylocentrotus droebachiensis) occurring in Kattegat experience seasonal hypercapnic and hypoxic conditions already today. Thus, anthropogenic CO2 emissions will add up to existing values and will lead to even higher pCO2 values >200 Pa (>2000 µatm). To estimate the green sea urchins' potential to acclimate to acidified seawater, we calculated an energy budget and determined the extracellular acid base status of adult S. droebachiensis exposed to moderately (102 to 145 Pa, 1007 to 1431 µatm) and highly (284 to 385 Pa, 2800 to 3800 µatm) elevated seawater pCO2 for 10 and 45 days. A 45 - day exposure to elevated pCO2 resulted in a shift in energy budgets, leading to reduced somatic and reproductive growth. Metabolic rates were not significantly affected, but ammonium excretion increased in response to elevated pCO2. This led to decreased O:N ratios. These findings suggest that protein metabolism is possibly enhanced under elevated pCO2 in order to support ion homeostasis by increasing net acid extrusion. The perivisceral coelomic fluid acid-base status revealed that S. droebachiensis is able to fully (intermediate pCO2) or partially (high pCO2) compensate extracellular pH (pHe) changes by accumulation of bicarbonate (maximum increases 2.5 mM), albeit at a slower rate than typically observed in other taxa (10 day duration for full pHe compensation). At intermediate pCO2, sea urchins were able to maintain fully compensated pHe for 45 days. Sea urchins from the higher pCO2 treatment could be divided into two groups following medium-term acclimation: one group of experimental animals (29%) contained remnants of food in their digestive system and maintained partially compensated pHe (+2.3 mM HCO3), while the other group (71%) exhibited an empty digestive system and a severe metabolic acidosis (-0.5 pH units, -2.4 mM HCO3). There was no difference in mortality between the three pCO2 treatments. The results of this study suggest that S. droebachiensis occurring in the Kattegat might be pre-adapted to hypercapnia due to natural variability in pCO2 in its habitat. We show for the first time that some echinoderm species can actively compensate extracellular pH. Seawater pCO2 values of >200 Pa, which will occur in the Kattegat within this century during seasonal hypoxic events, can possibly only be endured for a short time period of a few weeks. Increases in anthropogenic CO2 emissions and leakages from potential sub-seabed CO2 storage (CCS) sites thus impose a threat to the ecologically and economically important species S. droebachiensis.

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.

Organic properties of sediments and amino acids in iterstitial waters of ODP Holes 111-677A and 111-678B

The basement at Ocean Drilling Program (ODP) Sites 677 and 678 originated from the Galapagos spreading center of the Costa Rica Rift and has moved about 200 km over the last 6 m.y. (Fig. 1) (Shipboard Scientific Party, 1987, 1988; Scientific Drilling Party, 1987). Sediments about 300 m thick cover basement so young that basal sediments at Sites 677 and 678 have been reheated up to 60?-70?C at Site 677 and altered to limestone and/or chert (Shipboard Scientific Party, 1988). Sediments from both sites indicate (1) a high sedimentation rate (about 48 m/m.y.) and (2) biogenic silica and carbonate as the main constituents of sediments (Table 1) (Shipboard Scientific Party, 1988). Heatflow observations and measurements of interstitial water chemistry around the sites show that Site 677 is in a lower heatflow zone (166 mW/m**2; 1°12.14'N, 83°44.22'W) whereas Site 678 is located in a zone of higher heat flow (250 mW/m**2; 1°13.01'N, 83°43.39'W) (Langseth et al., 1988; Shipboard Scientific Party, 1988). In the flank hydrothermal systems, circulating solution is moving upward through the sedimentary column in zones of higher heat flow while it is moving downward in zones of lower heat flow (Anderson and Skilbeck, 1981). The chemistry of the interstitial waters is modified by several processes such as (1) diagenetic reactions and (2) advective and (3) diffusive transports of dissolved constituents. Analyses of Ca2+ and Mg2+ in interstitial waters from Sites 677 and 678 show that their profiles are mainly controlled by advective transport (Shipboard Scientific Party, 1988). In contrast, the interstitial-water profiles for NH4+, Si, and PO4[3-] are highly affected by reactions in the sediments. Site 677 offers a good opportunity to investigate amino acids in the interstitial waters because sediments of similar compositions have been deposited at constant rates of sedimentation. There are few previous works on amino acid distributions in interstitial waters (Henrichs and Parrington, 1979; Michaelis et al., 1982; Henrichs et al., 1984; Henrichs and Farrington, 1987; Ishizuka et al., 1988). In this chapter, we report (1) Rock-Eval analysis and (2) the composition of total hydrolyzable and dissolved free amino acids (THAA and DFAA, respectively) in the interstitial waters. Our objectives are to discuss (1) the possible origin of organic materials, (2) the characteristics of THAA and DFAA, and (3) their relationships in interstitial waters.