ALBERT

Description: Barnacles are dominant members of marine intertidal communities. Their success depends on firm attachment provided by their proteinaceous adhesive and protection imparted by their calcified shell plates. Little is known about how variations in the environment affect adhesion and shell formation processes in barnacles. Increased levels of atmospheric CO2 have led to a reduction in the pH of ocean waters (i.e., ocean acidification), a trend that is expected to continue into the future. Here, we assessed if a reduction in seawater pH, at levels predicted within the next 200 years, would alter physiology, adhesion, and shell formation in the cosmopolitan barnacle Amphibalanus (=Balanus) amphitrite. Juvenile barnacles, settled on silicone substrates, were exposed to one of three static levels of pHT, 8.01, 7.78, or 7.50, for 13 weeks. We found that barnacles were robust to reduced pH, with no effect of pH on physiological metrics (mortality, tissue mass, and presence of eggs). Likewise, adhesive properties (adhesion strength and adhesive plaque gross morphology) were not affected by reduced pH. Shell formation, however, was affected by seawater pH. Shell mass and base plate area were higher in barnacles exposed to reduced pH; barnacles grown at pHT 8.01 exhibited approximately 30% lower shell mass and 20% smaller base plate area as compared to those at pHT 7.50 or 7.78. Enhanced growth at reduced pH appears to be driven by the increased size of the calcite crystals that comprise the shell. Despite enhanced growth, mechanical properties of the base plate (but not the parietal plates) were compromised at the lowest pH level. Barnacle base plates at pHT 7.50 broke more easily and crack propagation, measured through microhardness testing, was significantly affected by seawater pH. Other shell metrics (plate thickness, relative crystallinity, and atomic disorder) were not affected by seawater pH. Hence, a reduction in pH resulted in larger barnacles but with base plates that would crack more readily. It is yet to be determined if such changes would alter the survival of A. amphitrite in the field, but changes in the abundance of this ecologically dominant species would undoubtedly affect the composition of biofouling communities.

Description: Ocean acidification (OA) adversely affects a broad range of marine calcifying organisms. Crustaceans, however, exhibit mixed responses to OA, with growth or survival negatively affected in some species, but unaffected or positively affected in others. In crustaceans, the mineralized cuticle resists mechanical loads, provides protection from the environment, and enables mobility, but little is known about how OA or interactions between OA and temperature affect its structure or function. Here, the effects of OA on the mechanics, structure, and composition of the cuticle in two Alaska king crab species was assessed. Juvenile blue king crabs (Paralithodes platypus) were exposed for a year to three pH levels, 8.1 (ambient), 7.8 and 7.5. Juvenile red king crabs (Paralithodes camtschaticus) were exposed for ~ 6 months to two pH levels, 8.0 and 7.8, at three temperatures: ambient, ambient + 2 °C, and ambient + 4 °C. Cuticle microhardness (a measure of resistance to permanent or plastic mechanical deformation), thickness, ultrastructure, and elemental composition were assessed in two body regions, the carapace and the crushing chela (claw). In both species tested, OA reduced endocuticle microhardness in the chela, but not in the carapace. There was no effect of pH or temperature on total procuticle thickness of the chela or carapace in either species. Reductions in microhardness were not driven by reduced calcium content of the shell. In fact, calcium content was significantly elevated in the carapace of blue king crabs and in the chela of red king crabs exposed to lower than ambient pH at ambient temperature, suggesting that calcium content alone is not a sufficient proxy for mechanical properties. Reduced chela microhardness, indicative of more compliant material, could compromise the utility of crushing chelae in feeding and defense.

Description: Background. Ocean acidification as a result of increased anthropogenic CO2 emissions is occurring in marine and estuarine environments worldwide. The coastal ocean experiences additional daily and seasonal fluctuations in pH that can be lower than projected end of century open ocean pH reductions. Projected and current ocean acidification have wide-ranging effects on many aquatic organisms, however the exact mechanisms of the impacts of ocean acidification on many of these animals remains to be characterized. Methods. In order to assess the impact of ocean acidification on marine invertebrates, Pacific oysters (Crassostrea gigas) were exposed to one of four different pCO2 levels for four weeks: 400 µatm (pH 8.0), 800 µatm (pH 7.7), 1000 µatm (pH 7.6), or 2800 µatm (pH 7.3). At the end of 4 weeks a variety of physiological parameters were measured to assess the impacts of ocean acidification: tissue glycogen content and fatty acid profile, shell micromechanical properties, and response to acute heat shock. To determine the effects of ocean acidification on the underlying molecular physiology of oysters and their stress response, some of the oysters from 400 µatm and 2800 µatm were exposed to an additional mechanical stress and shotgun proteomics were done on oysters from high and low pCO2 and from with and without mechanical stress. Results. At the end of the four week exposure period, oysters in all four pCO2 environments deposited new shell, but growth rate was not different among the treatments. However, micromechanical properties of the new shell were compromised by elevated pCO2. Elevated pCO2 affected neither whole body fatty acid composition, nor glycogen content, nor mortality rate associated with acute heat shock. Shotgun proteomics revealed that several physiological pathways were significantly affected by ocean acidification, including antioxidant response, carbohydrate metabolism, and transcription and translation. Additionally, the proteomic response to a second stress differed with pCO2, with numerous processes significantly affected by mechanical stimulation at high versus low pCO2 (all proteomics data are available in the ProteomeXchange under the identifier PXD000835). Discussion. Oyster physiology is significantly altered by exposure to elevated pCO2, indicating changes in energy resource use. This is especially apparent in the assessment of the effects of pCO2 on the proteomic response to a second stress. The altered stress response illustrates that ocean acidification may impact how oysters respond to other changes in their environment. These data contribute to an integrative view of the effects of ocean acidification on oysters as well as physiological trade-offs during environmental stress.

Description: Ocean acidification can affect the ability of calcifying organisms to build and maintain mineralized tissue. In decapod crustaceans, the exoskeleton is a multilayered structure composed of chitin, protein, and mineral, predominately magnesian calcite or amorphous calcium carbonate (ACC). We investigated the effects of acidification on the exoskeleton of mature (post-terminal-molt) female southern Tanner crabs, Chionoecetes bairdi. Crabs were exposed to one of three pH levels—8.1, 7.8, or 7.5—for two years. Reduced pH led to a suite of body-region-specific effects on the exoskeleton. Microhardness of the claw was 38% lower in crabs at pH 7.5 compared with those at pH 8.1, but carapace microhardness was unaffected by pH. In contrast, reduced pH altered elemental content in the carapace (reduced calcium, increased magnesium), but not the claw. Diminished structural integrity and thinning of the exoskeleton was observed at reduced pH in both body regions; internal erosion of the carapace was present in most crabs at pH 7.5, and the claws of these crabs showed substantial external erosion, with tooth-like denticles nearly or completely worn away. Using infrared spectroscopy, we observed a shift in the phase of calcium carbonate present in the carapace of pH-7.5 crabs: a mix of ACC and calcite was found in the carapace of crabs at pH 8.1, whereas the bulk of calcium carbonate had transformed to calcite in pH-7.5 crabs. With limited capacity for repair, the exoskeleton of long-lived crabs that undergo a terminal molt, such as C. bairdi, may be especially susceptible to ocean acidification.

Description: We examined the effect of long-term (2 year) exposure to decreased seawater pH (7.8 and 7.5, PCO2 ~ 760 and 1550 µatm, respectively) on exoskeletal properties in post-terminal-molt female Chionoecetes opilio. Since the effects of OA vary among body regions in decapods, exoskeletal properties (microhardness, thickness, and elemental composition) were measured in five body regions: the carapace, both claws, and both third walking legs.

Description: 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.