Ion transporter gene expression is linked to the thermal sensitivity of calcification in the reef coral Stylophora pistillata.

Coral calcification underpins biodiverse reef ecosystems, but the physiology underlying the thermal sensitivity of corals to changing seawater temperatures remains unclear. Furthermore, light is also a key factor in modulating calcification rates, but a mechanistic understanding of how light interacts with temperature to affect coral calcification is lacking. Here, we characterized the thermal performance curve (TPC) of calcification of the wide-spread, model coral species Stylophora pistillata, and used gene expression analysis to investigate the role of ion transport mechanisms in thermally-driven declines in day and nighttime calcification. Focusing on genes linked to transport of dissolved inorganic carbon (DIC), calcium and H+, our study reveals a high degree of coherence between physiological responses (e.g. calcification and respiration) with distinct gene expression patterns to the different temperatures in day and night conditions. At low temperatures, calcification and gene expression linked to DIC transport processes were downregulated, but showed little response to light. By contrast, at elevated temperature, light had a positive effect on calcification and stimulated a more functionally diverse gene expression response of ion transporters. Overall, our findings highlight the role of mechanisms linked to DIC, calcium and H+ transport in the thermal sensitivity of coral calcification and how this sensitivity is influenced by light.

Effects of light and darkness on pH regulation in three coral species exposed to seawater acidification.

The resilience of corals to ocean acidification has been proposed to rely on regulation of extracellular calcifying medium pH (pHECM), but few studies have compared the capacity of coral species to control this parameter at elevated pCO2. Furthermore, exposure to light and darkness influences both pH regulation and calcification in corals, but little is known about its effect under conditions of seawater acidification. Here we investigated the effect of acidification in light and darkness on pHECM, calcifying cell intracellular pH (pHI), calcification, photosynthesis and respiration in three coral species: Stylophora pistillata, Pocillopora damicornis and Acropora hyacinthus. We show that S. pistillata was able to maintain pHECM under acidification in light and darkness, but pHECM decreased in P. damicornis and A. hyacinthus to a much greater extent in darkness than in the light. Acidification depressed calcifying cell pHI in all three species, but we identified an unexpected positive effect of light on pHI. Calcification rate and pHECM decreased together under acidification, but there are inconsistencies in their relationship indicating that other physiological parameters are likely to shape how coral calcification responds to acidification. Overall our study reveals interspecies differences in coral regulation of pHECM and pHI when exposed to acidification, influenced by exposure to light and darkness.

In vivo pH measurement at the site of calcification in an octocoral.

Calcareous octocorals are ecologically important calcifiers, but little is known about their biomineralization physiology, relative to scleractinian corals. Many marine calcifiers promote calcification by up-regulating pH at calcification sites against the surrounding seawater. Here, we investigated pH in the red octocoral Corallium rubrum which forms sclerites and an axial skeleton. To achieve this, we cultured microcolonies on coverslips facilitating microscopy of calcification sites of sclerites and axial skeleton. Initially we conducted extensive characterisation of the structural arrangement of biominerals and calcifying cells in context with other tissues, and then measured pH by live tissue imaging. Our results reveal that developing sclerites are enveloped by two scleroblasts and an extracellular calcifying medium of pH 7.97 ± 0.15. Similarly, axial skeleton crystals are surrounded by cells and a calcifying medium of pH 7.89 ± 0.09. In both cases, calcifying media are more alkaline compared to calcifying cells and fluids in gastrovascular canals, but importantly they are not pH up-regulated with respect to the surrounding seawater, contrary to what is observed in scleractinians. This points to a potential vulnerability of this species to decrease in seawater pH and is consistent with reports that red coral calcification is sensitive to ocean acidification.

Coral calcifying fluid pH is modulated by seawater carbonate chemistry not solely seawater pH.

Reef coral calcification depends on regulation of pH in the internal calcifying fluid (CF) in which the coral skeleton forms. However, little is known about calcifying fluid pH (pHCF) regulation, despite its importance in determining the response of corals to ocean acidification. Here, we investigate pHCF in the coral Stylophora pistillata in seawater maintained at constant pH with manipulated carbonate chemistry to alter dissolved inorganic carbon (DIC) concentration, and therefore total alkalinity (AT). We also investigate the intracellular pH of calcifying cells, photosynthesis, respiration and calcification rates under the same conditions. Our results show that despite constant pH in the surrounding seawater, pHCF is sensitive to shifts in carbonate chemistry associated with changes in [DIC] and [AT], revealing that seawater pH is not the sole driver of pHCF Notably, when we synthesize our results with published data, we identify linear relationships of pHCF with the seawater [DIC]/[H+] ratio, [AT]/ [H+] ratio and [[Formula: see text]]. Our findings contribute new insights into the mechanisms determining the sensitivity of coral calcification to changes in seawater carbonate chemistry, which are needed for predicting effects of environmental change on coral reefs and for robust interpretations of isotopic palaeoenvironmental records in coral skeletons.

Morphological plasticity of the coral skeleton under CO2-driven seawater acidification.

Ocean acidification causes corals to calcify at reduced rates, but current understanding of the underlying processes is limited. Here, we conduct a mechanistic study into how seawater acidification alters skeletal growth of the coral Stylophora pistillata. Reductions in colony calcification rates are manifested as increases in skeletal porosity at lower pH, while linear extension of skeletons remains unchanged. Inspection of the microstructure of skeletons and measurements of pH at the site of calcification indicate that dissolution is not responsible for changes in skeletal porosity. Instead, changes occur by enlargement of corallite-calyxes and thinning of associated skeletal elements, constituting a modification in skeleton architecture. We also detect increases in the organic matrix protein content of skeletons formed under lower pH. Overall, our study reveals that seawater acidification not only causes decreases in calcification, but can also cause morphological change of the coral skeleton to a more porous and potentially fragile phenotype.

Coral calcifying fluid pH dictates response to ocean acidification.

Ocean acidification driven by rising levels of CO2 impairs calcification, threatening coral reef growth. Predicting how corals respond to CO2 requires a better understanding of how calcification is controlled. Here we show how spatial variations in the pH of the internal calcifying fluid (pHcf) in coral (Stylophora pistillata) colonies correlates with differential sensitivity of calcification to acidification. Coral apexes had the highest pHcf and experienced the smallest changes in pHcf in response to acidification. Lateral growth was associated with lower pHcf and greater changes with acidification. Calcification showed a pattern similar to pHcf, with lateral growth being more strongly affected by acidification than apical. Regulation of pHcf is therefore spatially variable within a coral and critical to determining the sensitivity of calcification to ocean acidification.

Imaging intracellular pH in a reef coral and symbiotic anemone.

The challenges corals and symbiotic cnidarians face from global environmental change brings new urgency to understanding fundamental elements of their physiology. Intracellular pH (pHi) influences almost all aspects of cellular physiology but has never been described in anthozoans or symbiotic cnidarians, despite its pivotal role in carbon concentration for photosynthesis and calcification. Using confocal microscopy and the pH sensitive probe carboxy SNARF-1, we mapped pHi in short-term light and dark-incubated cells of the reef coral Stylophora pistillata and the symbiotic anemone Anemonia viridis. In all cells isolated from both species, pHi was markedly lower than the surrounding seawater pH of 8.1. In cells that contained symbiotic algae, mean values of pHi were significantly higher in light treated cells than dark treated cells (7.41 +/- 0.22 versus 7.13 +/- 0.24 for S. pistillata; and 7.29 +/- 0.15 versus 7.01 +/- 0.27 for A. viridis). In contrast, there was no significant difference in pHi in light and dark treated cells without algal symbionts. Close inspection of the interface between host cytoplasm and algal symbionts revealed a distinct area of lower pH adjacent to the symbionts in both light and dark treated cells, possibly associated with the symbiosome membrane complex. These findings are significant developments for the elucidation of models of inorganic carbon transport for photosynthesis and calcification and also provide a cell imaging procedure for future investigations into how pHi and other fundamental intracellular parameters in corals respond to changes in the external environment such as reductions in seawater pH.

Importance of time and place: patterns in abundance of Symbiodinium clades A and B in the tropical sea anemone Condylactis gigantea.

The capacity of some corals and other cnidarians to form symbioses with multiple algae (Symbiodinium) is a candidate route by which these symbioses tolerate variable environmental conditions. On Bermuda, the coral reef dwelling anemone Condylactis gigantea bears Symbiodinium of clades A and B. At thermally variable inshore and nearshore sites, clade A predominates (as sole symbiont or in mixed infection with clade B), whereas animals at offshore sites with more uniform temperatures bear only clade B or mixed infections. Individual animals at one nearshore site monitored over a year by sampling tentacles showed increased prevalence of clade A in March-November, when sea waters were warm (average 26 degrees C), and increased clade B in November-March when cool waters prevailed (average 18.5 degrees C). In laboratory analyses of excised tentacles, the symbiosis with clade B, but not clade A, bleached at elevated temperature (32 degrees C), suggesting that thermal tolerance may contribute to the higher prevalence of clade A at inshore/nearshore sites and in the summer. The temporal changes in the algal complement were not accompanied by bleaching, and Symbiodinium density fluctuated in hosts with stable Symbiodinium composition but not in hosts with variable composition. This suggests that changes in the relative abundance of Symbiodinium clades do not require bleaching and may even protect the symbiosis from large fluctuations in algal density.

Photosynthetic symbioses in animals.

Animals acquire photosynthetically-fixed carbon by forming symbioses with algae and cyanobacteria. These associations are widespread in the phyla Porifera (sponges) and Cnidaria (corals, sea anemones etc.) but otherwise uncommon or absent from animal phyla. It is suggested that one factor contributing to the distribution of animal symbioses is the morphologically-simple body plan of the Porifera and Cnidaria with a large surface area:volume relationship well-suited to light capture by symbiotic algae in their tissues. Photosynthetic products are released from living symbiont cells to the animal host at substantial rates. Research with algal cells freshly isolated from the symbioses suggests that low molecular weight compounds (e.g. maltose, glycerol) are the major release products but further research is required to assess the relevance of these results to the algae in the intact symbiosis. Photosynthesis also poses risks for the animal because environmental perturbations, especially elevated temperature or irradiance, can lead to the production of reactive oxygen species, damage to membranes and proteins, and 'bleaching', including breakdown of the symbiosis. The contribution of non-photochemical quenching and membrane lipid composition of the algae to bleaching susceptibility is assessed. More generally, the development of genomic techniques to help understand the processes underlying the function and breakdown of function in photosynthetic symbioses is advocated.