An improved approach investigating epithelial ion transport in scleractinian corals

Coral epithelia control ion fluxes to the calcification site influencing biomineralization and proxy incorporation. However, data on in vivo characteristics of coral tissue such as permeability, selectivity, and active ion transport are scarce but important for calcification and proxy modeling. To investigate ion permeability and ion fluxes across coral tissues in vivo, we developed an electrophysiological approach for the assessment of active and passive epithelial transport properties. Growing Stylophora pistillata corals in a thin layer over permeable filters allowed ion exchange at the site of skeleton formation for reproducible measurements of electrophysiological properties of coral tissues in a modified Ussing chamber. Compared to former applications, electrical measurements on these coral filter units were dominated by tissue characteristics with minimal influence of skeleton or physical stress. Coral tissues were cation selective. Their overall high electrical resistance characterized them as tight epithelia indicating low paracellular permeability for passive ion diffusion. This includes ions relevant for calcification. A small short-circuit current indicates active charge transport across the entire coral tissue. The present approach is applicable to corals laterally overgrowing substrates. It allows the electrophysiological characterization of coral tissue in vivo in response to environmental conditions. This will improve our knowledge on transepithelial transport relevant for biomineralization in corals

Coral Host Cells Acidify Symbiotic Algal Microenvironment to Promote Photosynthesis

Symbiotic dinoflagellate algae residing inside coral tissues supply the host with the majority of their energy requirements through the translocation of photosynthetically fixed carbon. The algae, in turn, rely on the host for the supply of inorganic carbon. Carbon must be concentrated as CO2 in order for photosynthesis to proceed, and here we show that the coral host plays an active role in this process. The host-derived symbiosome membrane surrounding the algae abundantly expresses vacuolar H+-ATPase (VHA), which acidifies the symbiosome space down to pH ∼4. Inhibition of VHA results in a significant decrease in average H+ activity in the symbiosome of up to 75% and a significant reduction in O2 production rate, a measure of photosynthetic activity. These results suggest that host VHA is part of a previously unidentified carbon concentrating mechanism for algal photosynthesis and provide mechanistic evidence that coral host cells can actively modulate the physiology of their symbionts

Constraining calcium isotope fractionation (δ44/40Ca) in modern and fossil scleractinian coral skeleton

The present study investigates the influence of environmental (temperature, salinity) and biological (growth rate, inter-generic variations) parameters on calcium isotope fractionation (δ44/40Ca) in scleractinian coral skeleton to better constrain this record. Previous studies focused on the δ44/40Ca record in different marine organisms to reconstruct seawater composition or temperature, but only few studies investigated corals. This study presents measurements performed on modern corals from natural environments (from the Maldives for modern and from Tahiti for fossil corals) as well as from laboratory cultures (Centre Scientifique de Monaco). Measurements on Porites sp., Acropora sp., Montipora verrucosa and Stylophora pistillata allow constraining inter-generic variability. Our results show that the fractionation of δ44/40Ca ranges from 0.6 to 0.1‰, independent of the genus or the environmental conditions. No significant relationship between the rate of calcification and δ44/40Ca was found. The weak temperature dependence reported in earlier studies is most probably not the only parameter that is responsible for the fractionation. Indeed, sub-seasonal temperature variations reconstructed by δ18O and Sr/Ca ratio using a multi-proxy approach, are not mirrored in the coral's δ44/40Ca variations. The intergeneric variability and intrageneric variability among the studied samples are weak except for S. pistillata, which shows calcium isotopic values increasing with salinity. The variability between samples cultured at a salinity of 40 is higher than those cultured at a salinity of 36 for this species. The present study reveals a strong biological control of the skeletal calcium isotope composition by the polyp and a weak influence of environmental factors, specifically temperature and salinity (except for S. pistillata). Vital effects have to be investigated in situ to better constrain their influence on the calcium isotopic signal. If vital effects could be extracted from the isotopic signal, the calcium isotopic composition of coral skeletons could provide reliable information on the calcium composition and budget in ocean. Highlights ► Corals cultured in aquaria or from natural environment show the same Ca isotopic composition. ► δ44/40Ca of coral skeleton is independent of depositional setting environment. ► Strong influence of vital effects on coral skeleton δ44/40Ca composition and calcification mechanism

Genes Related to Ion-Transport and Energy Production Are Upregulated in Response to CO₂-Driven pH Decrease in Corals: New Insights from Transcriptome Analysis

Since the preindustrial era, the average surface ocean pH has declined by 0.1 pH units and is predicted to decline by an additional 0.3 units by the year 2100. Although subtle, this decreasing pH has profound effects on the seawater saturation state of carbonate minerals and is thus predicted to impact on calcifying organisms. Among these are the scleractinian corals, which are the main builders of tropical coral reefs. Several recent studies have evaluated the physiological impact of low pH, particularly in relation to coral growth and calcification. However, very few studies have focused on the impact of low pH at the global molecular level. In this context we investigated global transcriptomic modifications in a scleractinian coral (Pocillopora damicornis) exposed to pH 7.4 compared to pH 8.1 during a 3-week period. The RNAseq approach shows that 16% of our transcriptome was affected by the treatment with 6% of upregulations and 10% of downregulations. A more detailed analysis suggests that the downregulations are less coordinated than the upregulations and allowed the identification of several biological functions of interest. In order to better understand the links between these functions and the pH, transcript abundance of 48 candidate genes was quantified by q-RT-PCR (corals exposed at pH 7.2 and 7.8 for 3 weeks). The combined results of these two approaches suggest that pH ≥ 7.4 induces an upregulation of genes coding for proteins involved in calcium and carbonate transport, conversion of CO₂ into HCO₃⁻ and organic matrix that may sustain calcification. Concomitantly, genes coding for heterotrophic and autotrophic related proteins are upregulated. This can reflect that low pH may increase the coral energy requirements, leading to an increase of energetic metabolism with the mobilization of energy reserves. In addition, the uncoordinated downregulations measured can reflect a general trade-off mechanism that may enable energy reallocation

Live Tissue Imaging Shows Reef Corals Elevate pH under Their Calcifying Tissue Relative to Seawater

The threat posed to coral reefs by changes in seawater pH and carbonate chemistry (ocean acidification) raises the need for a better mechanistic understanding of physiological processes linked to coral calcification. Current models of coral calcification argue that corals elevate extracellular pH under their calcifying tissue relative to seawater to promote skeleton formation, but pH measurements taken from the calcifying tissue of living, intact corals have not been achieved to date. We performed live tissue imaging of the reef coral Stylophora pistillata to determine extracellular pH under the calcifying tissue and intracellular pH in calicoblastic cells. We worked with actively calcifying corals under flowing seawater and show that extracellular pH (pHe) under the calicoblastic epithelium is elevated by ∼0.5 and ∼0.2 pH units relative to the surrounding seawater in light and dark conditions respectively. By contrast, the intracellular pH (pHi) of the calicoblastic epithelium remains stable in the light and dark. Estimates of aragonite saturation states derived from our data indicate the elevation in subcalicoblastic pHe favour calcification and may thus be a critical step in the calcification process. However, the observed close association of the calicoblastic epithelium with the underlying crystals suggests that the calicoblastic cells influence the growth of the coral skeleton by other processes in addition to pHe modification. The procedure used in the current study provides a novel, tangible approach for future investigations into these processes and the impact of environmental change on the cellular mechanisms underpinning coral calcification

Calcification is not the Achilles' heel of cold-water corals in an acidifying ocean.

Ocean acidification is thought to be a major threat to coral reefs: laboratory evidence and CO2 seep research has shown adverse effects on many coral species, although a few are resilient. There are concerns that cold-water corals are even more vulnerable as they live in areas where aragonite saturation (Ωara ) is lower than in the tropics and is falling rapidly due to CO2 emissions. Here, we provide laboratory evidence that net (gross calcification minus dissolution) and gross calcification rates of three common cold-water corals, Caryophyllia smithii, Dendrophyllia cornigera, and Desmophyllum dianthus, are not affected by pCO2 levels expected for 2100 (pCO2  1058 μatm, Ωara 1.29), and nor are the rates of skeletal dissolution in D. dianthus. We transplanted D. dianthus to 350 m depth (pHT 8.02; pCO2  448 μatm, Ωara 2.58) and to a 3 m depth CO2 seep in oligotrophic waters (pHT 7.35; pCO2  2879 μatm, Ωara 0.76) and found that the transplants calcified at the same rates regardless of the pCO2 confirming their resilience to acidification, but at significantly lower rates than corals that were fed in aquaria. Our combination of field and laboratory evidence suggests that ocean acidification will not disrupt cold-water coral calcification although falling aragonite levels may affect other organismal physiological and/or reef community processes

Biochemical characterization of the skeletal matrix of the massive coral, Porites australiensis – The saccharide moieties and their localization

To construct calcium carbonate skeletons of sophisticated architecture, scleractinian corals secrete an extracellular skeletal organic matrix (SOM) from aboral ectodermal cells. The SOM, which is composed of proteins, saccharides, and lipids, performs functions critical for skeleton formation. Even though polysaccharides constitute the major component of the SOM, its contribution to coral skeleton formation is poorly understood. To this end, we analyzed the SOM of the massive colonial coral, Porites australiensis, the skeleton of which has drawn great research interest because it records environmental conditions throughout the life of the colony. The coral skeleton was extensively cleaned, decalcified with acetic acid, and organic fractions were separated based on solubility. These fractions were analyzed using various techniques, including SDS-PAGE, FT-IR, in vitro crystallization, CHNS analysis, chromatography analysis of monosaccharide and enzyme-linked lectin assay (ELLA). We confirmed the acidic nature of SOM and the presence of sulphate, which is thought to initiate CaCO3 crystallization. In order to analyze glycan structures, we performed ELLA on the soluble SOM for the first time and found that it exhibits strong specificity to Datura stramonium lectin (DSL). Furthermore, using biotinylated DSL with anti-biotin antibody conjugated to nanogold, in situ localization of DSL-binding polysaccharides in the P. australiensis skeleton was performed. Signals were distributed on the surfaces of fiber-like crystals of the skeleton, suggesting that polysaccharides may modulate crystal shape. Our study emphasizes the importance of sugar moieties in biomineralization of scleractinian corals

Light enhanced calcification in Stylophora pistillata: effects of glucose, glycerol and oxygen

Zooxanthellate corals have long been known to calcify faster in the light than in the dark, however the mechanism underlying this process has been uncertain. Here we tested the effects of oxygen under controlled pCO2 conditions and fixed carbon sources on calcification in zooxanthellate and bleached microcolonies of the branching coral Stylophora pistillata. In zooxanthellate microcolonies, oxygen increased dark calcification rates to levels comparable to those measured in the light. However in bleached microcolonies oxygen alone did not enhance calcification, but when combined with a fixed carbon source (glucose or glycerol), calcification increased. Respiration rates increased in response to oxygen with greater increases when oxygen is combined with fixed carbon. ATP content was largely unaffected by treatments, with the exception of glycerol which decreased ATP levels