Preliminary results with a torsion microbalance indicate that carbon dioxide and exposed carbonic anhydrase in the organic matrix are the basis of calcification on the skeleton surface of living corals
Ocean acidification is altering the ]]>
Agaricia agaricites were suspended from a torsion microbalance into gently stirred, temperaturecontrolled, seawater. Net calcification rates were monitored while light, temperature and pH were manipulated singly. The living coral pieces were sensitive to changes in conditions, especially light, and calcification was often suspended for one or two hours or overnight. The mean calcification rate increased from 0.06 in the dark to 0.10 mg.h-1.cm-2 ]]>
-1.m-2) and showed a positive linear relationship with temperature. With a reduction of mean pH from 8.2 to 7.6 the mean calcification rate in the light (65 μmol.s-1.m-2) increased from 0.19 to 0.28 mg.h-1.cm-2 (T test, n=8, p<0.05) indicating a dependency on carbon dioxide. After waterpiking and exposure of the skeletal surface/organic matrix to seawater, calcification showed an astonishing initial increase of more than an ]]>
-1.cm-2 (T test, n=11, p<0.02) again indicating a ]]>
-1.cm-2
(T test, n=6, p<0.05) indicating a dependence on carbonate. At a pH of 6.5 the skeleton lost weight at a rate of 1.8 mg.h-1.cm-2. The relationship between net calcification and pH (n=2) indicates that wt gain turns to loss at pH 7.4. These experiments confirm that calcification is a two-step process, involving secretion of a layer of ]]>
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Key words: coral calcification, CO2, pH, temperature, organic matrix, carbonic anhydrase.
Resumen ]]>
La acidificaión de los océanos está alterando la calcificón de los corales. Sin embargo, el mecanismo no es todavía claro. Para explorar que controla la calcificación piezas pequeñas del borde de láminas delgadas de Agaricia agaricites fueron suspendidas de una microbalanza de torsión en ]]>
-1 cm-2 (prueba T, n=8, p<0.01) en luminosidad baja (15 μmol s-1 m-2) y mostró una relación lineal positiva con la temperatura. Con una reducción en el pH promedio de 8.2 a 7.6 la tasa de calcificación media en la luz (65 μmol.s-1.m-2) aumentó de 0.19 a 0.28 mg h-1 cm-2 (prueba T, n=8, p<0.05) indicando una dependencia de dióxido de carbono. ]]>
-1 cm-2 (prueba T, n=11, p<0.02) de nuevo indicando la dependencia en el dióxido de carbono. La calcificación cesó en la presencia de azolamida un inhibidor de la anhidrasa carbónica. Tinciones confirmaron la presencia de anhidrasa carbónica, particularmente en las crestas de los septos. Después de sumergir esqueletos sin tejido ]]>
-1 cm-2 (prueba T, n=6, p<0.05) indicando una dependencia en carbonato. A un pH de 6.5 la tasa de pérdida de peso ]]>
-1 cm-2. La relación entre calcificaión neta y pH (n=2) indican que la gancia de peso se vuele pérdida a pH 7.4. Estos experimentos confirman que la calcificación es un proceso de dos pasos, involucrando la secreción de la capa de matriz orgánica que incorpora anhidrasa carbónica para producir una superficie de ]]>
Palabras clave: calcificación de corales, CO2, pH, temperatura, matriz orgánica, anhidrasa carbónica ]]>
The processes involved in the production of an aragonite (CaCO3) skeleton in corals are poorly known compared to calcification in other animals (Allemand et al. 2011). Seawater contains about 10.3 mmol.kg-1 of calcium ions ]]>
2+), dissolved inorganic carbon is present in seawater as carbon dioxide (CO2), carbonic acid (H2CO3), bicarbonate (HCO3-) and carbonate (CO32-), with the equilibria between them described by their equilibrium constants and their relationships with pH , alkalinity and temperature. What is still not clear is in what form dissolved ]]>
Ca2+ get there and to what extent, if at all, the calcifying surface is in direct contact with seawater. It seems to have been taken for granted that the formation of aragonite crystals would result from the combination of Ca2+ and CO32- dissolved in seawater. However, Lee et al. (2010) have reported crystalline aragonite deposition from a solution of CaCl2 containing carbonic anhydrase using CO2 directly from the atmosphere. The sources of calcium and carbon and how they reach the calcification site have been reviewed by Cohen & McConnaughey (2003), Furla et al. (2000) and Allemand et al. ]]>
et al. (2011). Carbon-dioxide which can move freely through cell membranes was proposed by McConnaughey (1989) as a substrate for calcification to account for 18O and 13C deficiencies in coral skeletons (McConnaughey, 2000). The response of corals to changes in CO2 partial pressure and temperature was reviewed by Reynaud et al. (2003). While calcification and photosynthesis may compete for the same DIC pool they are sometimes regarded as complementary (Gattuso et al. 1999) rather than in competition (Langdon & Atkinson, 2005). With a focus on ocean acidification as the result of increased anthropogenic carbon dioxide in the atmosphere, calcification has been increasingly linked to the calcium-carbonate saturation state ]]>
Ca2+] x [CO32-]) to the solubility product of the mineral deposited, in this case aragonite (Allemande et al. 2004). As acidity increases the relative concentration of carbonate in seawater is reduced and has been used to predict decreased calcification at the organism and community ]]>
et al., 1999; Gattuso et al., 1999, Marubini & Thake, 1999, Langdon, 2000, Langdon et al., 2000 Langdon et al. 2003 and Erez et al. 2011). However problems have emerged with the link between calcification and carbonate concentration or saturation state (Jury et al 2010, De Putron ]]>
et al. or with the predictions that have followed. From the banding of Florida corals from 1937 to 1996 Helmle et al., 2011 found that calcification was stable and the average rate during the most recent decade was not significantly different from those of the preceding 5 decades. Fabricius et al. (2011) found that as pH declined from 8.1 to 7.8 (the expected change by the end of the century), found reductions in coral diversity but not all corals were ]]>
et al. 2010 kept coral fragments in controlled aquarium conditions with normal and raised C02 levels equivalent to pH values of 8.09, 7.49 and 7.19 and the fragments all survived and added new skeleton. The Mediterranean coral Cladocera in low pH level did not show predicted reduced calcification (Rodolfo-Metalpa et al. 2010, 2011). There is a lot of information on the growth and dissolution kinetics of finely divided aragonite (Gutjahr et al., 1996, ]]>
et al.(2011) found that dead corals did not dissolve at a pH of 7.8 and measured dissolution rates at pH 7.4 (0.3-0.6 mg.g-1.d-1) and 6.8 (3.7-4.2 mg.g-1
.d-1). Rodolfo-Metalpa et al., (2011) suggested that coral tissues protect the skeletons from corrosive (Ω < 1) seawater. Villegas-Jiminez et al. (2009) reported large consistent proton/Ca2+ exchanges which ]]>
et al., 1996) the same problems of interpretation are likely to exist for aragonite.
Active calcification in ]]>
et al.1998 and Allemand et al. 2011) and calcium carbonate, crystallized in the form of aragonite, is deposited in the organic matrix with the involvement of structural proteins with a catalytic role similar to that of carbonic anhydrase (Tambutté et al. 2007), calcium binding compounds (Isa & Okazaki, 1987, Constantz & Wiener 1988 and Puverel
et al. 2005) and proteins regulating the biomineralization process along with Mg2+ which inhibits calcite formation (Rahman & Oomori, 2009). Aragonite deposition takes place from the extracellular calcifying fluid (ECF) or hydrogel-like medium (ECM) (Bryan & Hill, 1941 and Cuif et al. 2004a) onto the organic matrix framework on the surface of ]]>
Ca2+, HCO3- and CO32- cannot move passively through cell membranes so either active transport via a transcellular route, passive diffusion of ions or seawater using ]]>
et al. 2011, Cohen & McConnaughey, 2003). Ca2+, for example, may arrive from the calicoblastic layer as the result of the Ca2+ATPase pump ]]>
CO2 to the ECM (Adkins et al. 2003, Sandeman, 2008a). Allemand et al. 2011 reviewed the possible light enhancement mechanisms for calcification. The range of ratios observed for calcification rates in the light versus in the dark is ]]>
et al. 1999).
Calcification rates of corals are commonly derived by buoyant weighing techniques (Davies, 1989), uptake of the radioactive isotope 45Ca ( eg Moya et al. 2006, Al-Horani, 2007), the alkalinity anomaly technique (Smith & Key, 1975) or ]]>
Agaricia. A method was developed for calibration of the balance without the need to apply the complex equations (Davies, 1989 and Jokiel et al, 1978) used for determining the air weight of coral skeleton from its weight in seawater.
This study was ]]>
CO2 with the air. Also, removal of water for analysis changes the water level and interferes with balance function, the only means available of monitoring chemistry of the seawater in ]]>
CO2 or CO3 2- are used as the substrate for calcification. The ]]>
Materials and Methods
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Small pieces of Agaricia agaricites were snipped from the edge of thin plates of young colonies growing near the reef crest opposite the Discovery Bay Marine Laboratory. The pieces were immediately transported to the seawater tables where they were trimmed to a suitable size (1-2cm2) then suspended by loops ]]>
et al. (Table 1) using the CO@SYS program (Lewis and Wallis 1998) and the NBS Buffer scale.
For all experiments, single pieces of coral were suspended by the monofilament loop and a 5-6 cm length of 0.05 mm diam. stainless steel (s/s) wire from the beam of the torsion balance (Fig. 1) into a vessel with 1 or 2l of seawater that had been passed through activated charcoal and millepore filtered (0.45µ). The temperature of the water in the chamber except where otherwise stated was maintained at 28ºC and was regulated to within 0.1ºC by a temperature sensing thermistor, regulator circuit and insulated heater coil. A small magnetic stirrer (1.0x0.7cm) gently and continuously circulated the water in the vessel. The pH was changed by adding hydrochloric acid (adjusted to the density of seawater), or by exchanging some of the seawater in the vessel with seawater that had a high dissolved CO2 or NaCO3 content. For higher pH levels water in the vessel was exchanged with seawater that had been bubbled with CO2 free air. A pH meter (IQ200, Scientific Instruments), accurate to 2 decimal places, and calibrated daily, was used to monitor changes of pH. Room lighting was 15 μmol.s-1.m-2 and additional light was provided by a metal halide spotlight (Philips 25W, 10°) above the beaker. Irradiance levels were measured with a LI-COR Quantum/Radiometer (LI-250). The room was otherwise darkened and draughts were excluded as far as possible. The balance was protected from air movement by cardboard baffles and readings were only taken when the room air-conditioning unit was not active. The magnetic stirrer was turned off about three minutes before readings were taken. After suspending a piece of coral in the system the position of the balance beam was adjusted by the counter balance weight and/or rotating the fixed end of the torsion wire so that the laser beam was near the appropriate end the scale. Changes in weight as aragonite skeleton was deposited result in movement of the laser beam/spot on the scale and readings were taken at intervals of about ten minutes. The course of an experiment was followed by plotting the readings on graph paper. Rates of weight gain or loss were calculated by regression analysis of series of 4-6 readings. After a set of control reading a single experimental parameter (irradiance, temperature or pH) was then changed and after an hour for acclimation a new set of reading could be taken at 10 minute intervals to give a new rate. When rates of change were very low, longer periods between reading and more readings were taken.
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A series of small solid aragonite cubes, cut from Agaricia skeletons (1-3mg) of which the dry weights were accurately known were prepared beforehand. Each cube was attached to a 20 cm length of extremely fine monofilament consisting of a single strand from dental floss. Attachment was by dipping the end of the filament into cyanoacrylate and touching it to the cube. A cube was used to calibrate the system for each experiment by dropping it, ]]>
cm2, mean wt 0.42 g, n=56). The vessels used in the study were large (1 or 2 l beakers), so that changes in the composition of the sea water during an experiment were minimised. The greatest change affecting experiments was probably due to evaporation ]]>
CO2] and [CO3] at the control and treatment pH used can be seen in Table 1.
To examine the role ]]>
For the higher calcification rates of the waterpiked coral thicker wire was used in the balance to reduce its sensitivity so that the laser spot stayed on scale. The calcification rates of the freshly waterpiked coral were not linear and a different procedure had to be used to compare control and post treatment rates. Readings of weight were taken for 4-6 hours to ]]>
In order to see if there were any ]]>
CO2. Dried skeletons taken to Trent University, Canada, were used to determine the relationship between weight gain and loss across the pH range 8.2-6.0. For these experiments ]]>
NCSS statistical software (Number Crunching Statistical Systems, Dr Jerry Heintze, Kaysville, Utah) was used for obtaining a best fit for the growth curves.
Results ]]>
Live coral: Pieces of live Agaricia did not start calcifying until about 24 hours after collection but if newly collected pieces werem suspended on the torsion balance overnight in the dark calcification usually started before morning. Corals appeared to be very sensitive to any changes in conditions. Changes in light level or chemistry of the water were often ]]>
-1.m-2) were not used in the study because of the formation of oxygen bubbles which were sometimes formed and interfered with the functioning of ]]>
-1.m-2 calcification was positively correlated with temperature (Fig. 2). Over the range 27-29.5ºC the calcification rate increased by about 15% per 1ºC change (n=1). The mean calcification rate for Agaricia ]]>
increased by 60% from 0.063 mg.hr-1. cm-2 in the dark to 0.101 mg.hr-1.cm-2.in ambi ent laboratory lighting of 15 ]]>
-1.m-2 (T test, n=8, p < 0.01). When the irradiance level was increased to 65 μmol.s-1.m-2
further calcification often ceased for an hour or two then sometimes increased to a higher level but often to a lower level (mean 0.80 mg.hr-1.cm-2, T test, n=10, p > 0.05) than under the laboratory lighting. When the mean pH of the seawater was lowered from 8.2 to 7.6 the mean calcification rate increased from 0.19 mg.hr-1.cm-2 to 0.28 mg.hr-1.cm-2 (T test, n=7, p< 0.05).
Waterpiked coral: Freshly waterpiked pieces of coral suspended in seawater on the torsion balance were found to have an astonishing initial calcification rate (n=30) more than an order of magnitude higher than the same piece equation from enzyme kinetics (Lopez et al. 2000), with time replacing substrate level: Wt = (Wmax .t)/(K + t) (Note: Wt is the increase of weight at time t, Wmax is the asymptotic or maximum potential value of W and K is the time for half maximum growth).
With longer (t > 6 hours) experiments (Fig.4A, B, C) It became apparant that Wt does not actually reach an asymptote or maximum but continues to rise at a constant rate. The Michaelis-Menten general formula was of coral when alive. For example Agaricia #20 had a calcification rate of 0.029-0.063 mg.hr-1. cm-2 when alive but after waterpiking the initial calcification rate was over 1.0 mg.hr-1.cm-2. ]]>
The rate however decreases exponentially with time (Fig. 3). The weight/time data give a good fit (R2 = 0.9952) to the monomolecular growth equation, Wt = Wmax (1-ek.t) and a better fit (R2= 0.9987) to the generalized Michaelis-Menten modified to: Wt = ((Wmax .t)/(K + t)) + (Cb . t) where Cb is the constant rate of increase or slope of the line (Fig. 4B) and an even better fit to the data is obtained. For coral # 3a (Figs. 4A, B.) the fit to the modified formula (R2 = 0.9987) is better than that of the unmodified formula (R2 = 0.9983). At 27ºC (Fig. 5A) the predicted calcification rate was 0.48 mg.hr-1.cm-2, compared to the actual calcification rate at 29 ºC of 0.65 mg.hr-1.cm-2. This represents a 17.7 % increase per ºC which is fairly close to the 15% change per ºC for calcification obtained for live corals. The effect of changes of pH can be seen in Fig. 5B for a typical experiment. For a mean change of pH from 8.1 to 7.6 Agaricia responded with mean predicted calcification rate of 0.18 mg.hr-1.cm-2 compared to an actual mean calcification rate 0.32 mg.hr-1.cm-2. This represents a 78% increase in calcification rate (T test, n=11, p < .02). Light did not affect the calcification rate. The initial (first hour) calcification rates of freshly waterpicked coral obtained from the NCSS curve-fitting software (Fig. 7) are shown plotted against the time of day at which each coral piece was waterpiked (n=20). These results show a steady decrease in activity through the daylight light hours but a massive increase in the late afternoon.
Carbonic anhydrase: Experiments were undertaken to verify the presence of carbonic anhydrase in the exposed skeletal surface. When the carbonic anhydrase inhibitor, acetazolamide, was added to give a 100µMol solution calcification ceased completely (Fig. 5C).
Pieces of Agaricia skeleton that had been waterpiked and dried were also tested for carbonic anhydrase with the technique of Ridge way & Moffatt (1986). The tips of ridges and septae showed the quite distinctive blackening that indicates the presence of carbonic anhydrase (Fig. 6).
Dead coral (Waterpiked coral after soaking in seawater): A third series of experiments was undertaken to investigate what happens after a waterpiked coral has reached its asymptotic maximum Wmax. After waterpiking and suspension in seawater for 48-72 hours it was found, surprisingly, that rates of deposition were linear and the same order of magnitude as those of the live coral. Unlike the situation with freshly waterpiked corals the rate of deposition this time was positively correlated with pH. (Figs. 8A, B). When the mean pH of the seawater was reduced from 8.2 to 7.5 the mean calcification rate decreased from 0.25 to 0.13 mg..h-1.cm-2. This 50% reduction is statistically significant (T test, n=6, p < 0.05). With a larger change in pH from 8.25 to 6.5 the deposition rate (n=1) (Fig. 8C) changed from a gain of 1.2 mg.hr-1.cm-2 to a loss of 1.8 mg.hr-1.cm-2. Interpolation between the two rates indicates that the point at which the change from deposition to dissolution is at around pH 7.5. Again the deposition rate of CaCO3 was very sensitive to temperature (n=1), showing a 57% increase per ºC but there was no change in response to changes of light. (n=6). In the first of two more detailed explorations of the relationship of calcification/dissolution rate and pH the plot of calcification rate against pH (Fig. 9A) had a sigmoidal form similar to a hyperbolic sine function sinh = ½(ex –e-x). The data (20 points) give a good fit (R2= 0.965) to the formula R = (ex.f-e-x.b)/K where R is the net Wt gain/loss, x is pHr -pH0 (pH0 the pH at which R = 0) and f and b are constants (comparable to reaction order or dissolution rate constants) for the forward (weight gain) and backward (weight loss) parameters and K is a constant. Growth/ dissolution results are commonly presented as a function of Ω, for example r = K(1-Ω)n (Cubillas et al. 2005) but the treatment used here is similar to the approach of Lopez et al. (2009) and DePaulo (2011) who regard the net calcification rate R as the sum of the forwards rate (gain) Rf and backwards rate (loss) Rb. The relationship between growth/dissolution and pH of a second Agaricia skeleton (Fig. 9B) showed the same hyperbolic sine shape but the data (23 data points) gave a less good fit to the formula (R2= 0.730). However the difference in position of the data points from the descending pH changes and the ascending pH series are suggestive of the hysteresis effect, depending on which way their experiments were run, described by Gutjahr et al. (1996) for finely divided aragonite and calcite. In their comparison of the growth and dissolution rates of finely divided calcite and aragonite plotted against pH the curves, especially for aragonite, are similar to those found in this study. The pH at which R was zero for both skeletons in this study (Fig. 9A, B) was very close to a pH of 7.4. The same crossover point found by Gutjahr et al. (1996) was close to a pH of 7.8. One effect noticed in this study but not followed up was that when a piece of coral was moved into seawater of lower pH (eg from 8.1 to 7.5) the pH increased by about 0.01.h-1.
Discussion
In the experiments in which the pH of the experimental medium was reduced from an average of 8.2 to 7.6 (estimated [CO2] increases from 10.1 µmol kg-1
to 49.7 while estimated [CO32-] decreases from 241.5 µmol kg-1 to 75) calcification in the living corals increased significantly This provides good evidence that CO2 is the substrate for calcification rather than CO32-. The report by Lee et al. (2010) that crystalline aragonite is deposited from a solution of CaCl2 containing carbonic anhydrase using CO2 ]]>
CO2 can be the substrate for calcification.
Light enhanced calcification has been shown to be 3x the dark level (Allemand et al. 2011). ]]>
Agaricia agaricites the dark calcification rate is almost doubled in very low light, but for higher light levels, results were inconsistent. In this study when light levels were increased calcification usually ceased immediately for a period of hours then settled at a new level which could be higher or lower that the original. At higher light levels oxygen bubbles interfered with the experimental technique and the hyperbolic tangent relationship between irradiance and calcification rate found by Marubini et ]]>
(2004) or Moya et al. (2006) could not be confirmed. Sandeman (2008a) showed that the Ca2+ATPase/proton pump may be light sensitive as suggested by Cohen & McConnaughey (2003) and proposed (Sandeman, 2008b) that H2O2 produced by zooxanthellae in high light conditions makes the plasma membrane leaky to Ca2+
with the result that more Ca2+ could reach the ECM. The inconsistent response of calcification to light found in this and other studies is difficult to explain. However, if CO2 is indeed the substrate ]]>
CO2 between calcification and photosynthesis.
The (x25) higher ]]>
Agaricia and the response to acetazolamide (Fig. 5C). Inspection showed the accumulation of a thin ]]>
et al. 2007), has calcium binding properties (Isa & Okazaki 1987; Constantz & Wiener, 1988 and Puverel et al. 2005) and acidic proteins regulating the biomineralization process and the mineralization process(Rahman & Oomori, 2009) may involve all of these. Confirmation for the presence of carbonic anhydrase on the surface of waterpiked corals comes from the experiment involving inhibition of calcification by acetazolamide (Fig. ]]>
) and the demonstration by staining of carbonic anhydrase in the most rapidly growing areas of the skeleton (Fig. 6). The basic shape of the deposition versus time curves for waterpiked corals (Figs. 3, 4A, B) is probably the result of the exposed carbonic anhydrase or other active ingredients of the organic matrix being covered as new aragonite ]]>
Wmax is reached when the deposition rate is reduced to zero when all the carbonic anhydrase is obliterated by deposited aragonite. Some confirmation for this comes from the lowering of the initial calcification rate (Fig. 7) that takes place during the day. The ]]>
CO2] increases from 13.4 ]]>
-1 to 49.7, while estimated [CO32-] decreases from 202.5 µmol kg-1 to 75) the actual calcification rates showed an increase ]]>
CO2 rather than CO32- is the form in which DIC reaches the site of calcification as suggested by McConnaughey (1989). As reported by Dauphin et al. (2006) and Tambutte et al. (2007) the enzyme ]]>
Montastrea annularis skeleton from the museum collection showed some activity.
]]>
If the (x25) higher initial calcification rate of the freshly waterpiked coral skeletons compared to that of the same surface while alive is indeed the result of an enzyme or catalyst incorporation in the exposed surface it indicates that the calicoblastic layers restrict calcification and contact between the living calcifying surface and seawater delivery via a paracellular route must be small. A model that does not require a paracellular pathway is that of McConnaughey (1989) and Adkins
et al. (2003). It is based on dissolved CO2 as the substrate for calcification reaching the ECM by diffusion and Ca2+ ions are transported into the ECM and protons transported out by the ]]>
2+ATPase/proton pump. CO2 which can pass freely through lipid membranes reaches the ECM directly, with its passage enhanced by the pH difference between the ECM and calicoblastic layer created by movement of protons as the result of by the Ca2+ATPase/proton pump. In the presence of calcium binding proteins and carbonic anhydrase in the organic matrix calcium carbonate is deposited as aragonite: Ca2+ + H2CO3 → CaCO3 + 2H+
After waterpiking and soaking in seawater the pieces of dead coral skeleton showed a quite different response to lower pH. When the average pH was reduced from 8.2 to 7.5 (estimated [CO2] increases from 10.1 µmol kg-1 to 49.7, while estimated [CO32-] decreases from 241.5 µmol kg-1 to 75) calcification rate ]]>
Figs. 9A,B) gain weight and below 7.4 lose weight. The weight gains found in this study seem to be problematical. Emerson & Hedges (2009) state what appears to be a commonly held belief that when Ω >1 precipitation should occur “but this is rare because high concentrations of Mg. block nucleation sites on the mineral surface”, the results of this study suggest otherwise. The linear component of the weight gain Cb of ]]>
Fig. 9A, B) above and below 0, (described by b and f in the equations) are different. This was also found for finely divided aragonite by Gutjhar et al. (1996). The ]]>
Fig. 9B), also reported by Gutjhar et al. (1996) may be the result of the Ca2+/proton exchange reported by Villegas-Jiménez et al. (2009). In this ]]>
-1) indicating uptake of protons from the medium as described by Villegas-Jiménez et al. (2009). This effect as pointed out by Villegas-Jiménez et al. (2009) should be investigated because it could have important consequences for the ]]>
This study confirms that there are ]]>
et al. 2011). This was seen in living and and freshly waterpiked coral skeleton and is based on the enzyme carbonic anhydrase in the organic matrix, dissolved CO2 and Ca2+. Second, the deposition and dissolution seen in the freshly waterpiked pieces soaked in seawater (dead coral skeleton) with a non-living mineralization process which is carbonate-based and probably linked to the calcium carbonate saturation state ω.
In their survey ]]>
CO2 Reynaud et al. (2003) found -3% to -79% changes in calcification rate. Their own experiments indicated that, at normal temperatures, there was no response to elevated pCO2 but when temperature and pCO2 were both elevated, calcification dropped by 50%. Yii et al. (2009) found an increase in calcification rate for Galaxea fascicularis and a decrease for Porites cylindrica with increased
CO2 levels. Muehllehner & Edmunds (2008) reported a small (+5%) increase in calcification rate at 29ºC for Porites rus (n=11) and a larger (+100%) increase for Pocillopora meandrina (n=9) with increased pCO2; however at 27ºC there were reductions in calcification rate for ]]>
CO32-] and the carbonate-saturation state ω. In this study live corals and freshly waterpiked skeletons responded to lower pH or (elevated CO2) with increased calcification indicating that the substrate for calcification is CO2. It is perhaps important to consider why this result is at odds with the majority of studies that have reported coral calcification to decrease with increased pCO2. The explanation for the ]]>
Agaricia agaricites, were used. These, except for the thin broken edges, were completely covered with living tissue. In other studies the coral colonies used in calcification studies may have had dead exposed skeleton or a more porous skeleton with internal structural spaces, or channels and cavities from boring animals and which may present a significant area of skeleton in direct contact with seawater. The rate ]]>
Table 2). The difference between calcification rates in seawater of pH 8.2 and 7.8 range from +30% for coral with no dead areas to -21.5% for coral with ]]>
Cohen & McConnaughey (2003) posed the question “Why do coral reefs calcify so fast?”. One might also ask why corals calcify so slowly? In ]]>
Agaricia varied, at normal pH, from a mean of 0.063 mg.hr-1.cm-2 in the dark to 0.19 mg.hr-1.cm-2 in light while potential rates for newly exposed surface is over 1.0 mg.hr-1.cm-2. The reason is ]]>
Fig. 6 is distributed with the highest concentration on the fast growing septal ridges. This adds weight to the views of Wainwright (1963) and Allemand et al. (1989, 2011) that it is the organic matrix that determines the patterns of calcification and provides a mechanism for realizing the complex architecture of corals. The results obtained in this study are also consistent with the hypothetical mechanism presented by Allemand et ]]>
(2011) for the assembly of nanograins within the dynamic interface (Colfen & Mann, 2003) provided by the organic matrix.
There are implications from this study for understanding what is likely to happen to corals as the result of acidification and/or warming of the world’s ]]>
CO2] is higher, and increased temperature, will both enhance active biotic calcification, at least to a point at which other processes such as bleaching take over. For skeleton directly exposed to seawater (dead areas of the coral), as pCO2 increases and ω becomes lower, dissolution ]]>
et al. (2011) and Ries (2011) will affect survival by seriously weakening the supporting structure. Those species with smaller areas of exposed dead surface and a stronger Calcium/ ]]>
References
Adkins, J.F., E.A. Boyle, W B. Curry & A. Lutringer. 2003. Stable isotopes in deep-sea corals and ]]>
1129-1143. [ Links ]
Al-Horani, F.A., É. Tambutté & D. Allemand 2007. Dark calcification and the daily rhythm of calcification in the scleratinian coral, Galaxea fascicularis. Coral Reefs 26: 531-53. [ Links ]
Allemand, D., É. Tambutté, J-P. Girard & J. Jaubert. 1998. Organic matrix synthesis in the scleratinian coral Stylophora pistillata: role in biomineralization and potential target of the organotin tributyltin. J. Exp. Biol. 201: 2001-2009. [ Links ]
Allemand, D., C. Ferrier-Pagès, P. Furla, F. Houlbrèque, S. Puverel, S. Reynaud, É Tambutté, S. Tambutté, & D. Zoccola. 2004. Biomineralisation in reef-building corals: from molecular mechanisms to environmental control. C. R. Palévol 3: 453-467. [ Links ]
Allemand, D., É. Tambutté, D. Zoccola & S. Tambutté. 2011. Coral calcification, cells to reefs, p. 119-150. In Z, Dubinsky & N. Stambler (Eds). Coral Reefs: An Ecosystem in Transition. Springer Science+Business Media, Berlin. [ Links ]
Atkinson, M.J. & C. Bingham. 1999. Elemental composition of commercial seasalts. J. Aquaricult. Aquat. Sci. 8: 39-43. [ Links ]
Bryan, W.H. & D. Hill. 1941. Spherulitic crystallization as a mechanism of skeletal growth in the hexacorals. Proc. R. Soc. Queensl. 52: 78-91. [ Links ]
Cai, W.-J., X. Hu, W.-J. Huang, L.-Q. Jiang, Y. Wang, T.-H. Peng & X. Zhang. 2010. Alkalinity distribution in the western North Atlantic Ocean margins, J. Geophys. Res. 115, C08014, doi:10.1029/2009JC005482. [ Links ]
Cohen, A. & T.A. McConnaughey. 2003. Geochemical perspectives on coral mineralization. Rev. Min. Geochem. 54: 151-187. [ Links ]
Cölfen, H. & S. Mann. 2003. Higher-order organization by mesoscale self-assembly and transformation of hybrid nanostructures. Angew. Chem. Int. Ed. 42: 2350–2365. [ Links ]
Constantz, B.R. & S. Weiner. 1988. Acidic macromolecules associated with the mineral phase of scleratinian coral skeletons. J. Exp. Zool. 248: 253-258. [ Links ]
Cubillas, P., S. Köhler, M. Prieto, C Chaïrat and E.H. Oelkers. 2005. Experimental determination of the dissolution rates of calcite, aragonite, and bivalves. Chem. Geol. 216: 59-77. [ Links ]
Cuif, J-P. & Y. Dauphin. 2005a The environment recording unit in corals skeletons – a synthesis of structural and chemical evidences for a biochemically driven, stepping-growth process in fibres. Biogeosciences 2: 61–73. [ Links ]
Cuif, J.-P. & Y. Dauphin. 2005b. The two-step mode of growth in the scleractinian coral skeletons from the micrometre to the overall scale. J. Struct. Biol. 150: 319–331. [ Links ]
Dauphin, Y., J-P. Cuif & P. Massard. 2006. Persistent organic components in heated coral aragonitic skeletons-Implications for palaeoenvironmental reconstructions. Chem. Geol. 231: 26-37. [ Links ]
Davies, P. S. 1989. Short-term growth measurements of corals using an accurate buoyant weighing technique. Mar. Biol.101: 389-395. [ Links ]
DePaolo, D.J., 2011 Surface kinetic model for isotopic and trace element fractionation during precipitation of calcite from aqueous solutions. Geochim. Cosmochim. Acta 75: 1039-1056. [ Links ]
de Putron, S.J., D.C. McCorkle, A.L. Cohen & A.B. Dillon. 2011. The impact of seawater saturation state and bicarbonate ion concentration on calcification by new recruits of two Atlantic corals. Coral Reefs 30: 321-328. [ Links ]
Emerson, S. & J. Hedges. 2008. Chemical Oceanography and the Marine Carbon Cycle. Cambridge Univ., Cambridge, UK. [ Links ]
Erez, J., S. Reynaud, J. Silverman, K. Schneider & D. Allemand. 2011. Coral calcification under ocean acidification and global change, p. 151-176. In Z. Dubinsky and N. Stambler (eds). Coral Reefs: An Ecosystem in Transition, Springer Science+Business Media, berlin. [ Links ]
Fabricius, K.E., C. Langdon, S. Uthicke, C. Humphrey, S. Noonan, G. De’ath, R. Okaziki, N Muehllehner, M.S. Glas & J.M. Lough. 2011. Losers and winners in coral reefs acclimatized to elevated carbon dioxide concentrations. Nature Clim. Change 1: 165-169. [ Links ]
Franzisket, L. 1964. Die Stoffwechselintensität der Riffkorallen und ihre ökologische phylogenetische und soziologische Bedeutung. Z. Vergleich. Physiol. 49: 91-113. [ Links ]
Furla, P., I. Galgani, I. Durand & D. Allemand. 2000. Sources and mechanisms of inorganic carbon transport for coral calcification and photosynthesis. J. Exp. Biol. 203: 3445-3457. [ Links ]
Gattuso, J-P., D. Allemand & M. Frankignoulle. 1999. Photosynthesis and calcification at cellular, organismal and community levels in coral reefs: a review on interactions and control by carbonate chemistry. Amer. Zool. 39: 160-183. [ Links ]
Gischler, E. & W. Oschmann. 2005. Historical climate variation in Belize (Central America) as recorded in scleratinian corals. Palaios 20: 159-174. [ Links ]
Gutjahr, A., H. Dabringhaus and R Lacmann. 1996. Studies on growth and dissolution kinetics of the CaCO3 polymorphs calcite and aragonite. I. Growth and dissolution in water. J. Crystal Growth 158: 246-309. [ Links ]
Helmle, K.P., R.E. Dodge, P.K. Swart, D.K. Gledhill & C.M. Eakin. 2011. Growth rates of Florida corals from 1937 to 1996 and their responses to climate change. Nature Comm. 2: 215 doi: 10.1038/ ncomms1222. [ Links ]
Isa, Y. & M. Okazaki. 1987. Some observations on the Ca2+-binding phospholipids from scleratinian coral skeletons. Comp. Biochem. Physiol. 87B: 507-512. [ Links ]
Jury, C.P., R.F. Whitehead & A. Szmant. 2010. Effects of variations in carbonate chemistry on the calcification rates of Madracis auretenra (= Madracis mirabilis sensu Wells, 1973): bicarbonate concentrations best predict calcification rates. Glob. Change Biol.16: 1632-1644. [ Links ]
Kesling, R.V. & F.C. Crafts. 1962. Ontogenetic increase in archimedian weight of the ostracod Clamidotheca unispinosa (Baird). Am. Midl. Nat. 68: 149-153. [ Links ]
Krief, S., E. J. Hendy, M. Fine, R. Yam, A.Meibom, G. L. Foster & A. Shemesh. 2010. Physiological and isotopic responses of scleractinian corals to ocean acidification. Geochim. Cosmochim. Acta. 74 : 4988-5001. [ Links ]
Kleypas, J.A., R.R. Buddemeier, D. Archer, J.P. Gattuso, C. Langdon, and B.N. Opdike. 1999. Geochemical consequences of increased atmospheric carbon dioxide on coral reefs. Science 284: 118-20. [ Links ]
Langdon, C. 2000. Review of experimental evidence for effects of CO2 on calcification of reef builders. Proc. 9th Int. Coral Reef Symp., Bali 2: 1091-1098. [ Links ]
Langdon, C. and M.J. Atkinson. 2005. Effect of elevated pCO2 on photosynthesis and calcification of corals and interactions with seasonal change in temperature/ irradiance and nutrient enrichment. J. Geophys. Res. 110, CO9S07. [ Links ]
Langdon, C., W.S. Broecker & D.E. Hammond. 2003. Effect of elevated CO2 on the community metabolism of an experimental coral reef. Glob. Biogeochem. Cycles 17: 1101-1114. [ Links ]
Lee, S., J-H. Park, D. Kwak & K. Cho. 2010. Coral mineralization CaCO3 deposition via CO2 sequestration from the atmosphere. Cryst. Grow. Des. 10: 851-855. [ Links ]
Lopez, O., P. Zuddas & D. Faivre. 2009. The influence of temperature and seawater composition on calcite crystal growth mechanisms and kinetics: Implications for Mg incorporation in calcite lattice. Geochim. Cosmochim. Acta 73: 337–347. [ Links ]
Lopez, S., J. France, W.J. Gerrits, M.S. Dhanoa, D.J. Humphries & J. Dijkstra. 2000. A generalized Michaelis-Menten equation for the analysis of growth. J. Anim. Sci. 78: 1816-1828 [ Links ]
Mann, S. 1983. Mineralization in biological systems. Struct. Bond 54: 125-174. [ Links ]
Marsh, J.A. 1970. Primary productivity of reef-building calcareous red algae. Ecology 51: 255-263. [ Links ]
Marubini, F. & B. Thake. 1999. Bicarbonate addition promotes coral growth. Limnol. Oceanogr. 44: 716-720. [ Links ]
Marubini F., H. Barnett, C. Langdon & M.J. Atkinson. 2004. Dependence of calcification on light and carbonate ion concentration for the hermatypic coral Porites compressa. Mar. Ecol. Prog. Ser. 220: 153-162. [ Links ]
McConnaughey, T.A. 1989. 13C and 18O isotopic disequilibrium in biological carbonates. II. In vitro simulation of kinetic isotope effects. Geochim. Cosmochim. Acta 53: 163-171. [ Links ]
McConnaughey, T.A. 2000. Community and environmental influences on reef coral calcification. Limnol. Oceanogr. 45: 1667-1671. [ Links ]
Moya, A., S. Tambutté, E. Tambutté, D. Zoccola, N. Camaniti & D. Allemand. 2006. Study of calcification during a daily cycle of the coral Stylophora pistillata: implications for ‘light enhanced calcification’. J. Exp. Biol. 209: 3413-3419. [ Links ]
Muehllehner, N. & P.J. Edmunds. 2008. Effects of ocean acidification and increased temperature on skeletal growth of two scleratinian corals, Pocillopora meandrina and Porites rus. Proc. 11th Int. Coral Reef Symp. ,Ft. Lauderdale 3: 57-61. [ Links ]
Puverel, S., E. Tambutté, L. Pereira-Mouriès, D. Zoccola, D. Allemand & S. Tambutté. 2005. Soluble organic matrix of two scleratinian corals: Partial and comparative analysis. Comp. Biochem. Physiol. 141B: 480-487. [ Links ]
Rahman, M.A. & T. Oomori. 2009. In vitro regulation of CaCO3 crystal growth by the highly acidic proteins of calcitic sclerites in soft coral, Sinularia polydacta. Connect. Tiss. Res. 50: 285-293. [ Links ]
Reynaud, S., N. Leclercq, S. Romaine-Lioud, C. Ferrier-Pages, J. Jaubert & J.-P. Gattuso. 2003. Interacting effects of CO2 partial pressure and temperature on photosynthesis and calcification in a scleractinian coral. Glob. Change Biol. 9: 1660-1668. [ Links ]
Ridgway, R. L. & D. F. Moffett, 1986. Regional differences in the histochemical localization of carbonic anhydrase in the midgut of tobacco hornworm (Manduca sexta). J. Exp. Zool. 237: 407-412. [ Links ]
Rodolfo-Metalpa, R., S. Martin, C. Ferrier-Pagés & J-P. Gattuso. 2010. Response of the temperate coral Cladocera caespitosa to midand long term exposure to pCO2 and temperature levels projected for the year 2100. Biogeosciences 7:289-300. [ Links ]
Rodolfo-Metalpa, R., F. Houlbréque, É. Tambutte, F. Boisson, C. Baggini, F.P. Patti, R. Jeffree, M. Fine, A. Foggo, J.P. Gattuso & J.M. Hall-Spencer. 2011. Coral and mollusc resistance to ocean acidification adversely affected by warming. Nature Clim. Change 1: 308-312. [ Links ]
Sandeman, I.M. 2008a. Fine banding in the septa of corals. Proc. 11th. Int. Coral Reef Symp., Ft. Lauderdale 3: 57-61. [ Links ]
Sandeman, I.M. 2008b. Light driven lipid peroxidation of coral membranes and a suggested role in calcification. Rev. Biol. Trop. 56 (Suppl. 1): 1-9. [ Links ]
Smith, S.V. & G.S. Key. 1975. Carbon dioxide and metabolism in marine environments. Limnol. Oceanogr. 20: 493-495. [ Links ]
Tambutté, S., É. Tambutté, D. Zoccola, N. Camaniti, S. Lotto, A. Moya, D. Allemand & J. Adkins. 2007. Characterization and role of carbonic anhydrase in the calcification process of the azooxanthellate coral Tubastrea aurea. Mar. Biol. 151: 71-83. [ Links ]
Villegas-Jiménez, A. Mucci & J. Paquette. 2009. Proton/ calcium ion exchange behavior of calcite. Phys. Chem. Chem. Phys. 11: 8895-8912. [ Links ]
Wainwright, S.A. 1963. Skeletal organization in the coral Pocillopora damicornis. Quart. J. Micros. Sci. 104: 164-183. [ Links ]
Yii, S.-H., A. Munafi, A. Bolong, T.-T. Yang & H.-C. Liew. 2009. Effect of elevated carbon dioxide on two scleractinian corals: Porites cylindrica (Dana, 1846) and Galaxea fascicularis (Linnaeus, 1767). J. Mar. Biol. 2009: doi:10.1155/2009/215196. [ Links ]
*Correspondencia: Ian M. Sandeman: Dept. of Biology, Trent University, Peterborough, Ontario, K9J 7B8, Canada; isandem@pipcom.com or isandeman@trentu.ca
1. Dept. of Biology, Trent University, Peterborough, Ontario, K9J 7B8, Canada; isandem@pipcom.com or isandeman@trentu.ca ]]>
Received 23-VI-2011. Corrected 14-XII-2011. Accepted 20-XII-2011 ]]>2003671129-1143200726531-5319982012001-200920043453-4672011119-1501999839-4319415278-912010115200354151-1872003422350-23651988248253-258200521659-772005a261-732005b150319-331200623126-371989101389-3952011751039-1056201130321-3282008201120111165-16919644991-11320002033445-3457199939160-183200520159-1741996158246-30920112198787B507-51219785529-5422010161632-1644196268149-1532010744988-50011999284118-2020002Bali 1091-10982005110CO9S072003171101-1114201010851-855200973337-3472000781816-1828198354125-174197051255-263199944716-7202004220153-162198953163-1712000451667-167120062093413-341920083Ft. Lauderdale 57-612005141B-487200950285-293200391660-16681986237407-41220111294-29520107289-30020111308-3122008a3Ft. Lauderdale 57-612008b56^s1^s111-9197520493-495200715171-832009118895-89121963104164-18320092009