Journal of Experimental Marine and Ecology 394 (2010) 53–62

Contents lists available at ScienceDirect

Journal of Experimental and Ecology

journal homepage: www.elsevier.com/locate/jembe

Photoacclimation mechanisms of corallimorpharians on coral reefs: Photosynthetic parameters of zooxanthellae and host cellular responses to variation in irradiance

Baraka Kuguru a,b, Yair Achituv a, David F. Gruber c, Dan Tchernov b,d,e,⁎ a The Mina and Everard Goodman Faculty of Life Sciences, Bar-Ilan University, Ramat Gan 52900, Israel b The Interuniversity Institute for Marine Sciences in Eilat, P.O. Box 469, Eilat 88103, Israel c Department of Natural Sciences, City University of New York, , P.O. Box A-0506, 17 Lexington Avenue, New York, New York 10010, d Department of Evolution, Systematics and Ecology, Hebrew University of Jerusalem, Edmund Safra Campus, Givat Ram, Jerusalem 91904, Israel e Marine Biology Department, The Leon H. Charney School of Marine Sciences, University of Haifa, Mount Carmel, Haifa 31905, Israel article info abstract

Article history: Rhodactis rhodostoma and Discosoma unguja are the most common corallimorpharians on coral reefs in the Received 17 March 2010 northern Red Sea, where individuals of R. rhodostoma form large aggregations on intertidal flats and Received in revised form 7 July 2010 those of D. unguja occupy holes and crevices on the reef slope. Aside from these contrasting patterns of Accepted 8 July 2010 microhabitat, little is known concerning their mechanisms of photoacclimation to environmental conditions. We demonstrate here that different mechanisms of photoacclimation operate in both and that these Keywords: differences explain, in part, the contrasting patterns of distribution and abundance of these common Corallimorpharians corallimorpharians. Experimental exposure of the species' respective polyps to the synergistic effects of Microhabitat Photoacclimation ultraviolet and photosynthetically active radiation revealed that endosymbiotic zooxanthellae protected the Photosynthesis host R. rhodostoma from photooxidation damage. Zooxanthellae do so by reducing their chlorophyll pigment Ultraviolet radiation and cellular abundance, as well as by adjusting their efficiency of light absorption and utilization according to Zooxanthellae the level of irradiance. The host photoprotects its endosymbionts from harmful ultraviolet radiation (UVR) by synthesizing enzymatic antioxidants against radicals. In contrast, individuals of D. unguja utilize a behavioral mechanism of photoacclimation in which they physically migrate away from exposed areas and towards shaded habitats and thus avoid the damaging biological effects of UVR. We conclude that a combination of physiological and behavioral mechanisms appear to control microhabitat segregation between these corallimorpharian species on tropical reefs. These various mechanisms of local adaptation to environmental conditions may be largely responsible for the wide distributional ranges of some corallimorpharians, and may enable these common reef organisms to tolerate environments that are highly variable, both spatially and temporally. © 2010 Elsevier B.V. All rights reserved.

1. Introduction steady rise in atmospheric CO2 has led to higher sea surface (SST) (Hoegh-Guldberg, 1999; Hoegh-Guldberg et al.,

Corallimorpharians are non-calcifying, evolutionarily important 2002, 2007) and lower pH levels. Increasing atmospheric CO2 has relatives of stony corals (Medina et al., 2006), although the exact been postulated to deplete the layer (Austin et al., 1992), relationship between Corallimorpharia and the Scleractinia remains leading to an increase of ultraviolet radiation (UVR) on the oceans' under debate (Fukami et al, 2008). Increased understanding of the surfaces (Harley et al., 2006). Understanding the protective mechan- ecophysiology of corallimorpharians can provide insights into the isms used by marine organisms to mitigate the damage caused by UVR evolution of corals from Mesozoic to recent forms (Stanley and Fautin, is particularly urgent today, as the thinning of atmospheric ozone by 2001) and their ability to survive drastic climatic changes. greenhouse gases has magnified the intensity of UVR reaching the sea Coral reefs are among the most vital and biologically diverse surface in some areas (McKenzie et al., 1998). In clear tropical ecosystems on the planet. Despite their great value, both ecological seawater, UVR penetrates to ecologically important depths (Gleason and socio-economical, however, coral reefs are severely threatened by and Wellington, 1993). UVR radiation breaks down dissolved organic anthropogenic global (IPCC, 2001; IPCC, 2007). The carbon (Hader et al., 2007), which is responsible for short-wavelength absorption in the water column. In addition, oceanic warming and acidification results in faster degradation of dissolved and particulate ⁎ Corresponding author. Marine Biology Department, The Leon H. Charney School of organic carbon (DOC, POC), thereby enhancing the penetration of UVR Marine Sciences, University of Haifa, Mount Carmel, Haifa 31905, Israel. Tel.: +972 4 8288790; fax: +972 4 8282515. into the water column (Sinha and Hader, 2002). Short-term increases E-mail address: [email protected] (D. Tchernov). in UVR intensity under calm, clear water conditions may expose

0022-0981/$ – see front matter © 2010 Elsevier B.V. All rights reserved. doi:10.1016/j.jembe.2010.07.007 54 B. Kuguru et al. / Journal of Experimental Marine Biology and Ecology 394 (2010) 53–62 marine algae and to photophysiological effects of UVR carotenoids, xanthophyll pigments, and antioxidants [both enzymatic, stress (Gleason and Wellington, 1993). As a result, the productivity of i.e., superoxide dismutase (SOD) and catalase; and non-enzymatic, marine ecosystems may be greatly and adversely affected (Sinha and i.e., vitamin E (Lesser, 2006)]; (3) changes in tissue structure and Hader, 2002). morphology (Brown et al., 1994; Loya et al., 2001; Kuguru et al., 2007; Tropical reef-building corals and other photosynthetic marine Mass et al., 2007); and (4) sunscreens in the form of UV-absorbing invertebrates live in habitats where solar irradiance may be extremely compounds (UVAC), also known as mycosporine-like amino acids high (Shick et al., 1996; Lesser, 2000; Banaszak and Lesser, 2009). (MAAs) (Shick et al., 1999; Banaszak et al., 2000). These mechanisms Several studies have implicated exposure to elevated solar radiation, of photoacclimation vary among species of symbiotic host cnidarians, including both photosynthetically active radiation (PAR, 400–700 nm, partly because their zooxanthellae comprise a highly divergent group Hoegh-Guldberg and Smith, 1989; Lesser and Shick, 1989) and UVR of dinoflagellates (Coffroth and Santos, 2005) with a broad range of (290–400 nm, Lesser and Stochaj, 1990; Kinzie, 1993; Baruch et al., genotypic and phenotypic responses to light (Iglesias-Prieto and 2005; Lesser and Shick, 1989), as a major contributor to coral Trench, 1994, 1997; Savage et al., 2002; Robison and Warner, 2006), bleaching stress. In particular, UVB (290–320 nm) suppresses photo- which in turn influence their ecological distributions (LaJeunesse, synthesis while simultaneously increasing the risk of damage to DNA, 2002; Iglesias-Prieto et al., 2004). proteins, and membrane (Greenberg et al., 1989; Lyons et al., In addition, variation among host species in traits such as behavior 1998; Baruch et al., 2005), partly through the production of reactive [e.g., polyp retraction (Brown et al., 1994), and contraction/expansion oxygen species (ROS) (Lesser, 1989; Lesser and Lewis, 1996; Shick et (Brown et al., 2002)], gastrodermal tissue structure (Kuguru et al., al., 1996; Jokiel et al., 1997; Tchernov et al., 2004). While the 2007), skeletal structure (Mass et al., 2007), and light absorption by photophysiological effects of UVR stress in some hexacorallians fluorescent proteins (Salih et al., 2000), all potentially modulate the (scleractinians, actiniarians and zoanthids) have been studied available light and, thereby, impact the photochemical response of extensively, corallimorpharians have been almost completely over- their endosymbionts. While species-specific patterns of photoaccli- looked in this context. mation have been elucidated for marine algae (Huner et al., 1996), Like many marine invertebrates, some corallimorpharians harbor stony corals (Falkowski and Dubinsky, 1981; Warner and Berry-Lowe, endosymbiotic dinoflagellate algae, also known as zooxanthellae 2006), and actinian sea anemones (Stoletzki and Schierwater, 2005), (den-Hartog, 1980; LaJeunesse, 2002; Kuguru et al., 2007, 2008). almost nothing is known about mechanisms of response to light stress Zooxanthellae play a critical role in host nourishment via the trans- in corallimorpharians. location of photosynthates (Muscatine et al., 1981). Recent studies have The corallimorpharians R. rhodostoma and D. unguja are both demonstrated that zooxanthellae are made up of eight broad clades common on coral reefs in some areas of the Indo-Pacific, and are (designated A–H), each of which contains multiple, closely related successful recolonizers of shallow habitats following disturbances molecular types that exhibit a range of physiological responses and such as bleaching, which kill stony corals and other zooxanthellates tolerances (reviewed in Coffroth and Santos, 2005; Stat et al., 2006). The (Chadwick-Furman and Spiegel, 2000; Kuguru et al., 2004; Work et al., specific type(s) that an organism harbors may affect its 2008). They occupy contrasting microhabitats on the reef: individuals distribution and reaction to extreme environmental conditions (Robison of R. rhodostoma form large aggregations on intertidal reef flats while and Warner, 2006). In the Red Sea, the common corallimorpharians those of D. unguja occupy holes and crevices deeper on the reef slope Rhodactis rhodostoma and Discosoma unguja both host Symbiodinium (Muhando et al., 2002; Kuguru et al., 2008). Effects of UVR are type C1 in shallow waters (1–6 m), and types D1a and C1 in deeper expected to be highly pronounced in the Red Sea, which is classified as waters (18–20 m) (Kuguru et al., 2007, 2008). A recent study in an oligotrophic class II water body, representing one of the most Australia (Jones et al., 2008) indicated that both clades C1 and D1 may optically clear water bodies in the world (Stambler, 2005). Thus, confer equal thermotolerance to host corals. Both of the above understanding differences in mechanisms of UVR acclimation be- corallimorpharian species, when transplanted from their original tween these two corallimorpharians may provide insights into their habitats to either shallower or deeper water, shuffletheirsymbiont bathymetric distributional patterns and predict the extent to which types, a mechanism that may contribute to their occupation of a wide they can withstand climate change. The objective of the present study bathymetric range on coral reefs (Kuguru et al., 2008). However, was to experimentally assess the photoacclimation mechanisms of individuals of D. unguja are more susceptible to light stress than are these two corallimorpharian species in response to increased PAR and those of R. rhodostoma, and tend to occur in deeper, more shaded UVR in terms of the photosynthetic parameters of their zooxanthellae habitats (Kuguru et al., 2008). The mechanisms employed by these two density and the cellular responses of their tissues. corallimorpharians to photoacclimatize to microhabitats on the that differ in level of irradiance (UVR+PAR) are not yet understood. The success of corals and other reef invertebrates that have 2. Materials and methods essentially transparent tissues but manage to thrive in shallow water indicates that they have developed effective mechanisms for UVR 2.1. Study site and polyp collection protection. There are substantial differences between closely related species in their ability to escape the damaging effects of UVR in this This study was conducted during the period January–August 2007 high-energy waveband (Sinha and Hader, 2002). The ability of at the Interuniversity Institute for Marine Sciences (IUI), in Eilat, scleractinian corals and other reef organisms to survive environmen- Israel, in the northern Red Sea (29º30′N, 34°55′E). Polyps of the tal changes depends on their physiological mechanisms of acclima- corallimorpharians R. rhodostoma and D. unguja were collected on tization (Gates and Edmunds, 1999). Many symbiotic cnidarians coral reefs adjacent to the IUI at depths of 3–20 m, and attached to display rapid modifications in behavior, morphology, and physiology polyvinyl chloride (PVC) bases using underwater epoxy. Care was that enable them to photoacclimate to changing light conditions, thus taken to ensure that the replicate polyps were located at least 10 m demonstrating considerable biological flexibility. Shallow reef organ- apart from each other, to avoid collecting individuals that originated isms exposed to high levels of solar UVR have evolved several types of asexually from the same parent colonies. Following a one-month photoacclimation mechanisms to cope with light stress, including: (1) acclimation period in outdoor aquaria (irradiance of 300 μmol pho- behavioral avoidance, such as migrating away from intense light tons m− 2 s− 1 (PAR), equivalent to that at 18–20 m depth, flow- (Gleason et al., 2006) and into shaded microhabitats (crevices and through seawater at 120 L h− 1), the polyps were transferred to holes on the reef); (2) mechanisms to control internal cellular experimental aquaria for treatments (see below, modified after damage, such as development of free-radical quenching agents like Kuguru et al., 2007, 2008). B. Kuguru et al. / Journal of Experimental Marine Biology and Ecology 394 (2010) 53–62 55

2.2. Experimental design

We assayed three major types of characteristics related to host and algal responses to light stress (increased PAR and UVR) in these corallimorpharians: (1) physiological parameters reporting photo- synthetic activity (zooxanthella density, chlorophyll a (chl a) pigment biomass, and chl a fluorescence), to indicate whether metabolic processes (photosynthesis and oxidative phosphorylation) were subject to photoinhibition; (2) host- peroxide (LPO) levels, to indicate whether the structural integrity of host cells was challenged; and (3) host enzymatic activity, specifically copper/zinc superoxide dismutase (Cu/ZnSOD), to indicate whether there was an enzymatic response to oxidative stress. These parameters were selected because they reflect cellular physiological functions that provide evidence for identifying the stressors in terms of PAR, UVR, or both (Dunlap and Chalker, 1986; Kana, 1992; Iglesias-Prieto and Trench, 1994; Downs et Fig. 1. Irradiance spectrum of natural solar radiation over outdoor experimental aquaria al., 2002). at the Interuniversity Institute for Marine Science in Eilat, Israel, and of 290 nm and fi In this study, we performed a laboratory experiment under 400 nm cut-off lters deployed to create experimental treatments (see text for details). controlled conditions to understand the physiological acclimatization mechanisms of both photosymbiont and host cells of the two species of corallimorpharians when exposed to changes in environmental sunset using the fluorescence induction and relaxation system FIRe conditions. The setup of the experiment was based on results obtained [Satlantic Instrument, Canada (Kolber et al., 1998), see below for from our previous studies (Kuguru et al., 2007, 2008). This further explanation]. Then, at midday the following day, a small experiment was performed using twelve 9-L aquaria, each supplied amount of tissue (approximately 0.1 g) was removed from the oral with ambient (26 °C) through-flow seawater at a constant flow rate of disc of each polyp and immediately snap-frozen in liquid nitrogen to 2 L min− 1 and a mixing pump of 230 L h− 1.Afilter (100 μm) was minimize handling effects and stop enzymatic activities. Collection fitted to the 20-mm supply pipe connected to a 50-mm main seawater took place between 12:00 and 13:00 h to ensure that explants were supply pipe to filter out bivalve shells while allowing phytoplankton sampled at the highest light intensities. Samples were then stored at and to pass through. The filter was cleaned every two −80 °C. days to ensure a uniform flow rate to each unit. For biochemical analysis, frozen samples (approximately 0.1 g Every aquarium contained one polyp each of R. rhodostoma and D. fresh ) were homogenized with 2 mL of ice-cold 50 mM unguja. Six individuals of each species were photoacclimated using potassium phosphate buffer (pH 7.4). To prevent oxidation of layers of plastic-netting shade for one month in either of two outdoor samples, 10 μL 0.5 M butylated hydroxytoluene (BHT) in acetonitrile treatments: (1) high light (HL), 50% shade, with a maximum midday was added to 1 mL of tissue homogenate. For separation of irradiance of 700 μmol photons m− 2 s− 1, equivalent to irradiance at zooxanthellae and host cells, the resulting slurry from homogenized approximately 5 m water depth; and (2) low light (LL), 90% shade, tissue was centrifuged for 5 min at 6000 rpm at 4 °C. The pellet with a maximum midday irradiance of 350 μmol photons m− 2 s− 1, (zooxanthella cells) was re-suspended in 2 mL 50 mM potassium equivalent to irradiance at approximately 20 m depth. Thus, there phosphate buffer (pH 7.4), aliquoted in 1000 μL and stored at −80 °C. were six aquaria for each treatment, totalling 12 aquaria. Light (PAR; The supernatant (host cells) was placed in a new eppendorf tube and in μmol photons m− 2 s− 1) was measured using a quantum sensor (LI- further centrifuged for 10 min at 10,000 rpm at 4 °C to remove host 190SA, LiCor, USA). The experiment was designed to mimic the tissue. The clear supernatant was aliquoted in 400 μL and stored at natural light conditions during summer months on coral reefs in Eilat, −80 °C. The aliquots of zooxanthella cells were used for zooxanthella based on PAR measurements. abundance and chlorophyll a pigment assessment. Host cells aliquots After one month of pre-acclimation in LL and HL, the organisms were used for Cu/ZnSOD and LPO assessment. were covered with two different cut-off filters: PAR alone (700 nm≤ In zooxanthella-containing fractions from the corallimorpharians λ≥400 nm), hereafter called UVO; or PAR+UVR (700≤λ≥295 nm), (see above), we measured (1) chl a content (overnight extraction in hereafter called UVT; each with 90% spectral irradiance transmission 90% acetone), with absorbance readings at 630, 664, and 750 nm (Fig. 1). Thus, four PAR/UV treatments were generated: (1) high light (Jeffrey and Humphrey, 1975); and (2) zooxanthella abundance (cells with UVT (HLUVT); (2) high light without UV (HLUVO); (3) low light counted under a light microscope using a haemocytometer) after with UVT (LLUVT); and (4) low light without UV (LLUVO). There were Kuguru et al. (2007). Algal pigment and abundance were normalized three polyps of each species in each of these four treatments. per milligram of total protein. Normalizing data per wet tissue mass Irradiances (UVR+PAR; in μWcm− 2)ineachtreatmentwere (Kuguru et al., 2007) was not possible because all collected samples quantified using a radiometer (Biospherical PRR-800, San Diego, were frozen immediately in liquid nitrogen to minimize handling USA). We used a ‘preacclimatization’ stage before exposing the effects and to stop enzymatic activities. corallimorpharians to the four treatments in consultation with results To determine the synergistic effects of PAR and UVR, the activity from our previous study (Kuguru et al., 2008), which showed that when of antioxidant enzymes (Cu/ZnSOD) due to oxidative stress in host exposed to high irradiance under laboratory conditions or in field cells was measured using a spectrophotometric kit assay for transplants for about a month, members of both corallimorpharian superoxide dismutase (Cu/ZnSOD-525, Catalog number 21010 from species hosted Symbiodinium type C1. Thus, in the present study, we Oxis International, Inc., USA) according to the manufacturer's pre-acclimatized all polyps for one month to ensure that all hosted only instructions. Each assay was done in triplicate in each of the four − one Symbiodinium clade to prevent clade-based variation in their irradiance treatments. Results were expressed as Cu/ZnSOD Umg 1 tolerance/acclimatization mechanisms. soluble host protein. At the end of the above experiment on light/UV exposure (i.e., To test whether oxidative and cell damage was associated with 30 days), the stress responses of polyps of R. rhodostoma and D. lipid peroxidation (LPO) due to the synergistic effects of PAR and UVR unguja to the synergistic effects of UVR and PAR were assessed. on host cells, malondialdehyde (MDA) was assayed spectrophoto- First, a chlorophyll a fluorescence reading was taken 30 min after metrically using a lipid peroxidation assay kit (Oxford Biomedical 56 B. Kuguru et al. / Journal of Experimental Marine Biology and Ecology 394 (2010) 53–62

Research MDA-586, Catalog number 21025, from Oxford Biomedical 3. Results Research, USA) according to the manufacturer's instructions. Each assay was done in triplicate in each of the four irradiance treatments. The irradiance spectra of the UVR cut-off filters used in this study Results were expressed as μM LPO mg− 1 soluble host protein. show that these filters screened UVR effectively (Fig. 1). A comparison The level of protein in the crude extracts of samples of the irradiance quantities measured at sea and in each experimental (n=3) in all irradiance treatments was determined spectrophoto- laboratory treatment revealed that the corallimorpharian polyps metrically (Bradford, 1976) using a protein assay kit [Bovine serum under the HLUVT treatment experienced PAR and UVR (313 nm) albumin (BSA) standard set, Catalog number 500-0207 from Bio-Rad, equal to radiation experienced by polyps at approximately 2-m depth USA] according to the manufacturer's instructions. in the sea (Table 1). Polyps under LL treatment experienced PAR equal Comparison of the synergistic effects of UVR and PAR on the to approximately 23-m depth, but UVR (313 nm) in the LLUVT was photosynthetic performance of algae associated with the polyps in the equivalent to approximately 5-m depth. Polyps situated under the four treatments was performed at the end of PAR/UVR acclimation, hanging roof experienced PAR equal to approximately 40-m depth i.e., after one month of incubation. Chlorophyll fluorescence yields and UVR (313 nm) equal to approximately 10-m depth (Table 1). were measured using the fluorescence induction and relaxation Polyps of the two corallimorpharian species differed in their system (FIRe; Satlantic Instrument) as an indication of the number of physiological responses to solar radiation effects (PAR and/or UVR photosystem II (PSII) units (Kolber et al., 1998). FIRe fluorometry only). The zooxanthella abundance and chlorophyll a measures chlorophyll fluorescence transients using a controlled series of both species decreased with increasing levels of irradiance of subsaturating flashes that cumulatively saturate PSII within (HLUVTNHLUVONLLUVTNLLUVO; Fig. 2a, b). However, individuals of ~100 μs, i.e., a single photochemical turnover (Kolber and Falkowski, D. unguja had significantly higher levels of chlorophyll a concentration

1993; Kolber et al., 1998). The FIRe technique can, therefore, measure (2-way ANOVA: F1, 24=7.66, p=0.014) and lower levels of zooxan- several fluorescence parameters. Fo and Fm are minimum and thella abundance (2-way ANOVA: F1, 24 =37.65, p=0.00001) in their maximum yields of chl a fluorescence, measured in a dark-adapted tissues than did individuals of R. rhodostoma (Fig. 2a, b). Despite state. Fv is variable fluorescence (Fo −Fm), and Fv/Fm (−(Fm −Fo)/Fm) these trends, chlorophyll a pigment per zooxanthella cell did not differ is the maximum quantum yield of photochemistry in PSII as measured significantly between the two species among treatments (2-way in a dark-adapted state. In addition to these parameters, the FIRe ANOVA, F3, 24 =1.24,p=0.33; Fig. 2c). There was no significant inter- fluorometer instantaneously measures the functional absorption cross action effect between species and treatment group for chlorophyll section of PSII (σPSII), which characterizes the capacity to absorb and pigment (2-way ANOVA: F3, 24=3.06, p=0.058), zooxanthella abun- utilize visible radiation (Kolber et al., 1998) via the fluorescence dance (2-way ANOVA F3, 24=1.16, p=0.4), and chlorophyll concen- signature of the O2-evolving complex, PSII. Cross sections derived tration per algal cell (2-way ANOVA, F3, 24 =1.36, p=0.3; Fig. 2a, b, c). from conventional O2 evolution measurements were similar to σPSII Measurements of chlorophyll a fluorescence parameters (Fv/Fm, (Falkowski et al., 1988). Using a pulse amplitude-modulated fluo- σPSII,andQm) revealed that higher irradiance levels reduced rometer (Diving PAM, Walz, Germany), light or excitation photosynthetic processes for both species of corallimorpharians pressure on photosystem II (Qm) was determined according to Kuguru (Fig. 3). The impact was most vivid in the HLUVT treatment, where et al. (2007). Differences in Fv/Fm, σPSII, and excitation pressure on PS high irradiance levels reduced Fv/Fm (2-way ANOVA: F1, 24 =21.03, II between symbionts within some corals provide a predictive p=0.0003; σ PSI, 2-way ANOVA: F1, 24 =4.9, p=0.04), and also measure of how coral species and algal symbionts respond to natural caused a significant increase in Qm (2-way ANOVA: F1, 24 =16.93, thermal stress (Falkowski and Chen, 2003). p=0.001) levels in polyps of D. unguja compared to R. rhodostoma A separate laboratory experiment was conducted to assess the (Fig. 3). There was a significant interaction effect between species and behavioral responses of both corallimorpharian species to UVR stress. treatment group for Fv/Fm (2-way ANOVA: F3, 24 =8.22, p=0.002), An outdoor 100-L flow-through aquarium supplied with running σPSII (2-way ANOVA: F3, 24 =5.48, p=0.01), and Qm (2-way ANOVA: −1 seawater at a constant flow rate of 120 L h was set up under an F3, 24 =9.28, p=0.001; Fig. 3). Student-Newman-Keuls (SNK) post- overhanging roof (not exposed to direct sunlight), with a maximum hoc tests at Pb0.05 revealed that polyps of D. unguja exposed to − 2 − 1 midday irradiance of approximately 85% μmol photons m s HLUVT irradiance levels had significantly lower values of Fv/Fm and (equivalent to irradiance at approximately 20–30-m depth). This σPSII than did polyps of D. unguja exposed to the other three irradiance treatment mimicked irradiance levels at the deeper end of the depth range of these two species (Chadwick-Furman and Spiegel, 2000; Kuguru et al., 2007). Five polyps of each species were affixed to a PVC Table 1 base, and then the bases were attached to a 4×30-cm plastic plate. Irradiance (UVR+PAR) levels from radiometer (Biospherical PRR-800, San Diego, USA) fi Two plates, each with five polyps of either species of corallimorphar- measurements under eld and laboratory conditions in the present study. ian, were acclimated in the aquarium for one year. Observations were A. Irradiance (μWcm− 2) attenuation in the field on coral reefs at Eilat, northern made on the behavioral responses of the corallimorpharians in terms Red Sea. of their positions on the bases. Depth (m) UVB (313 nm) UVA (340 nm) PAR (400-700 nm)

0 17.5 56.6 155.5 1 9.3 34.5 94.2 2.3. Statistical analyses 3 2.9 16.2 47.0 5 0.7 6.4 29.8 The combined effects of PAR and UVR on zooxanthella abundance, 15 0.1 2.4 19.4 20 0.0 0.8 11.5 chlorophyll-pigment concentration per cell, Fv/Fm, Qm, Sigma, SOD, 23 0.0 0.5 9.0 and LPO were examined using two-way ANOVA. The raw data were − used following examination of the homogeneity of the variances B. Irradiance levels (μWcm 2) within each laboratory treatment. (Levene's test) and the normality of the data (normal probability Irradiance UVB (313 nm) UVA (340 nm) PAR plots). Proportional data were arcsine-transformed prior to statistical treatment analysis. Significant differences between levels within the factors HL UVT 6.7 23.9 89.8 were examined post-hoc with Student-Newman-Keuls (SNK) tests. UVO 0.0 0.0 89.5 The behavioral response (habitat preference) by corallimorpharians LL UVT 0.7 2.4 9.0 UVO 0.0 0.0 9.0 was evaluated using the Strauss (1979) selectivity index (L). B. Kuguru et al. / Journal of Experimental Marine Biology and Ecology 394 (2010) 53–62 57

Fig. 2. Variation in zooxanthella and chlorophyll parameters among laboratory treatments in polyps of the corallimorpharians Rhodactis rhodostoma and Discosoma unguja. Data shown are after 30 days of exposure to four laboratory treatments: high light (daily maximum 700 μmol photons m− 2 s− 1) with UV radiation (HLUVT), high light with no UV radiation (HLUVO), low light (daily maximum 350 μmol photons m− 2 s− 1) with UV radiation (LLUVT), and low light with no UV radiation (LLUVO). Data are means±SE, n=3 polyps per treatment. Treatments marked with different superscript letters are significantly different at Pb0.05 (Student-Newman-Keuls post-hoc tests, see text for details). treatment levels (HLUVO, LLUVT/-UVO). In contrast, polyps of R. four irradiance treatment levels, indicating that UVR did not induce rhodostoma demonstrated equal values of Fv/Fm (2-way ANOVA: F3, 24 = stress responses in the algal symbionts of this host species (Fig. 3a, b). 1.86, p=0.2) and σPSII (2-way ANOVA: F3, 24 =10.85, p=0.5) at all HL irradiance treatments significantly increased Qm values in the

Fig. 3. Variation in chlorophyll a fluorescence parameters among four laboratory treatments in polyps of the corallimorpharians Rhodactis rhodostoma and Discosoma unguja. Data shown are after 30 days of exposure to four laboratory treatments: high light (daily maximum 700 μmol photons m− 2 s− 1) with UV radiation (HLUVT), high light with no UV radiation (HLUVO), low light (daily maximum 350 μmol photons m− 2 s− 1) with UV radiation (LLUVT), and low light with no UV radiation (LLUVO). Data are means±SE, n=3 polyps per treatment. Treatments marked with different superscript letters are significantly different at Pb0.05 (Student-Newman-Keuls post-hoc tests, see text for details). 58 B. Kuguru et al. / Journal of Experimental Marine Biology and Ecology 394 (2010) 53–62 zooxanthellae of both host species as compared to LL irradiance Table 2 Proportions of corallimorpharian polyps maintaining their position on plastic bases (R), treatments (2-way ANOVA: F3, 24 =2.73, p=0.08). In contrast, D. the relative contribution of polyps of both corallimorpharian species (two plates unguja was significantly more sensitive to UVR than was R. pooled) (P), and the Strauss (1979) Linear Selectivity Index (L). NB: Selectivity Index rhodostoma (2-way ANOVA: F1, 24 =16.93, p=0.001; Fig. 3c) in that (L) values range from −1 to 1, with positive values indicating preference and negative the zooxanthellae of D. unguja had significantly higher values of Qm values indicating avoidance; **=L very significant from 0. when exposed to UVT versus UVO treatments, while those of R. RPL=R−PL=0 rhodostoma did not (SNK post-hoc tests at Pb0.05; Fig. 3c.) Exposure to high levels of irradiance induced increased host Rhodactis rhodostoma 1.00 0.50 0.50 ** Discosoma unguja 0.00 0.50 −0.50 ** cellular (enzymatic) Cu/ZnSOD and LPO activity in both R. rhodostoma and D. unguja, but the magnitude of these responses differed between the host species (2-way ANOVA: F1, 12 =16.13, p=0.004; Fig. 4a), and LPO (2-way ANOVA, F1, 24 =24.83, p=0.0001; Fig. 4b). For the HLUVT 4. Discussion treatment, higher irradiance levels caused significantly lower Cu/

ZnSOD activities (2-way ANOVA: F 3, 12 =351.05, p=0.0001; Fig. 4a) We demonstrate here that two common corallimorpharians on and higher LPO (2-way ANOVA: F1, 24 =24.83, p=0.000; Fig. 4b) Indo-Pacific coral reefs utilize a variety of contrasting mechanisms of (host cellular degradation) in D. unguja as compared to R. rhodostoma. photoacclimation to mitigate the damaging effects of high solar In addition, all treatments with UVT, but not UVO, caused significantly irradiation. Their defensive mechanisms, when experimentally ex- higher host cellular (protective) Cu/ZnSOD enzymes (2-way ANOVA: posed to the synergistic effects of UVR+PAR, included: (1) reductions

F3, 12 =351.05, p=0.0001; Fig. 4a). In both LL Cu/ZnSOD treatments, in the abundance of zooxanthellae and concentration of chlorophyll a samples were inadvertently mistreated and, therefore, no data were (Fig. 2); (2) quenching of the light excitation energy, as revealed by obtained. variations in Qm and σPSII (Fig. 3b, c); (3) host-mediated synthesis of The behavioral response experiment revealed D. unguja to be light- enzymatic antioxidants (Fig. 4a, b); and (4) behavioral responses to sensitive [Strauss (1979) selectivity index L=−0.5; Table 2]. All five high irradiance via polyp migration to shaded microhabitats (Fig. 5). polyps of D. unguja moved from the upper exposed surfaces to the Our observations that both chlorophyll concentration and zoo- more shaded sides of their PVC bases within 6–8 weeks (Fig. 5b). In xanthella abundance in these corallimorpharians decrease with level contrast, the polyps of R. rhodostoma did not relocate at all during the of irradiance (Fig. 2a, b) are similar to patterns observed in other 12-month period [Strauss (1979) selectivity index L=0.5; Table 2], cnidarians with variations in depth, irradiance, and and remained on top of their bases, fully exposed to ambient irradiance (Fagoonee et al., 1999; Fitt et al., 2000; Kuguru et al., 2007; Mass et al., (Fig. 5a). 2007). These changes apparently prevent host cellular damage from the oxidative stress induced by elevated sea temperature, PAR, and/or UVR (Baruch et al., 2005; Lesser and Shick, 1989), and also allow the host to adjust its photosynthate supply to nutritional demands. The amount of chl a per symbiont cell in these corallimorpharians was not affected by light regime, in contrast to the stony coral Stylophora pistillata, which increases its pigment per symbiont cell with decreasing light (Falkowski and Dubinsky, 1981). However, patterns in the corallimorpharians are consistent with the effects of ambient UVR on the stony coral Montipora verrucosa zooxanthellae, in which equal values of chl a occur per algal cell at both low and high irradiance levels (Kinzie, 1993). Gleason and Wellington (1993) concluded that coral bleaching induced by UVR results from reductions in zooxanthella densities rather than the combined effects of zooxanthella expulsion and decreases in chlorophyll concentration per algal cell, as has been noted for temperature-related bleaching (Glynn and Dcroz, 1990). We did not quantify levels of accessory pigments other than chlorophyll in the zooxanthellae, but these also may have responded differently to the irradiance treatments.

Our measurements of σPSII revealed that the mechanisms of physiological acclimation differed significantly between R. rhodostoma

and D. unguja (Fig. 3b). σPSII measures the efficiency of excitation energy transfer from the antenna pigments to the PSII reaction center and represents the amount of light energy usable for photochemical reactions (Kolber et al., 1998). It strongly depends on environmental

conditions such as Qm/light pressure (Kolber et al., 1990). Qm values showed higher excitation energy in D. unguja zooxanthellae than in

those of R. rhodostoma (Fig. 3c).Inaddition,asignificant decline in σPSII values during HLUVT treatment in D. unguja zooxanthellae (Fig. 3b) suggests that they were highly susceptible to the synergistic effects of high irradiance and UVR. Lower values of both σ and F /F in HL Fig. 4. Variation in levels of superoxide dismutase (Cu/ZnSOD) and lipid peroxidation PSII v m (LPO) of the corallimorpharians Rhodactis rhodostoma and Discosoma unguja among compared to LL treatments in D. unguja (Fig. 3a, b) most likely resulted laboratory treatments. Data shown are after 30 days of exposure to four laboratory from a combination of photodamage and photoprotective processes treatments: high light (daily maximum 700 μmol photons m− 2 s− 1) with UV radiation (Warner et al., 2002) and correlated with long-term photoinhibition (qI) (HLUVT), high light with no UV radiation (HLUVO), low light (daily maximum values (Brown et al., 1999; Falkowski and Chen, 2003; Gorbunov et al., 350 μmol photons m− 2 s− 1) with UV radiation (LLUVT), and low light with no UV 2001). In contrast, both σ and F /F values did not change in R. radiation (LLUVO). Data are means±SE, n=3 polyps per treatment. Treatments PSII v m marked with different superscript letters are significantly different at Pb0.05 (Student- rhodostoma zooxanthellae regardless of the irradiance treatment, Newman-Keuls post-hoc tests, see text for details). suggesting that these symbionts were not physiologically compromised, B. Kuguru et al. / Journal of Experimental Marine Biology and Ecology 394 (2010) 53–62 59

Fig. 5. Photographs of representative polyps of the corallimorpharians Rhodactis rhodostoma and Discosoma unguja, showing their behavioral responses to irradiance under: (a, b) laboratory conditions and (c, d) field conditions, on coral reefs in the Red Sea. possibly because they possess effective mechanisms for the dissipation UVR versus either PAR or UVR alone (Fig. 4). Host cellular responses of excess light excitation energy. The cellular mechanisms underlying (Fig. 4a) indicate that individuals of D. unguja have higher levels of ROS these physiological responses may involve differential gene expression than do those of R. rhodostoma. In other cnidarians, high levels of ROS induced by changes in irradiance, leading to antenna systems with correlate with exposure to UVR radiation, which causes inhibition in different σPSII values (Mortainbertrand et al., 1990; Smith et al., 2005). the photosynthetic machinery of PSII (Iwanzik et al., 1983)by This resilience to photooxidative stress can also result from other degrading the D1/32 kDa protein complex (Melis et al., 1992; Malanga underlying processes including sustained PSII repair, compensatory et al., 1997). UVR radiation also causes ROS and high levels of SOD in electron flow through remaining functional reaction centers (Behrenfeld green algae and diatoms (Lesser, 2006). Generation of photosynthesis- et al., 1998) and structural differences in thylakoid membranes mediated ROS occurs in the chloroplast through several mechanisms (Tchernov et al., 2004). associated with photosystem I- and photosystem II-catalyzed electron Under laboratory conditions at low light irradiance, R. rhodostoma transfer, the most notable being the Mehler reaction and the harbors symbiont type C1 while D. unguja harbors symbiont type D1a. generation of hydrogen peroxide by the oxygen-evolving complex

However, at high irradiance, both R. rhodostoma and D. unguja harbor (Kana, 1992; Downs et al., 2002). H2O2 may diffuse out of the Symbiodinium type C1 (Kuguru et al., 2008). A recent study in chloroplasts through the algal symbiont into the host cytoplasm, Australia (Jones et al., 2008) indicates that both clades C1 and D1 may where it can be neutralized by either an enzymatic or non-enzymatic confer equal thermo-tolerance to host corals. Hence, the differences in antioxidant pathway or catalyzed through the Fenton reaction to zooxanthella photosynthetic performance observed here were most hydroxyl (OH) radical (Downs et al., 2002). Above a threshold level, likely due to microenvironment effects (Coffroth and Santos, 2005) the presence of OH will cause the host to either expel or destroy the provided by the host rather than to genetic differences among their zooxanthellae as a defense mechanism against oxidative stress (Lesser zooxanthellae. Host-mediated photoacclimation mechanisms (Lesser and Shick, 1989). Perez and Weis (2006) later suggested that ROS and Shick, 1989; Ambarsari et al., 1997; Salih et al., 2000; Brown et al., released from heat/UV-stressed photosystems in algal cells might 2002) may enhance photosynthetic rates in resident zooxanthellae induce nitric-oxide (NO) production in host cells. They specifically (Gates and Edmunds, 1999). Furthermore, inconsistencies between hypothesized that NO converts to cytotoxic peroxynitrite upon the phylogenetic and physiological attributes of zooxanthellae interaction with superoxide in the host cell, leading to cell death and (Savage et al., 2002; Tchernov et al., 2004; Robison and Warner, bleaching. In our case, the significantly higher Cu/ZnSOD and LPO 2006; Kuguru et al., 2008), and the physiological responses of isolated levels (Fig. 4a, b) in D. unguja host cells in the HLUVT treatment (Fig. 4) versus in hospite Symbiodinium (Bhagooli and Hidaka, 2003; Goulet et indicate that the enzymatic and non-enzymatic oxidants within these al., 2005), suggest that the host could be the determinant factor for the cells were unable to compensate for and ameliorate the destructive fitness of the holobiont. capacity of the ROS. On the other hand, low values of LPO and Cu/ Levels of oxidative stress in the host were exacerbated when the ZnSOD in the host cells of R. rhodostoma demonstrate that they more corallimorpharians were exposed to the synergistic effects of PAR+ effectively suppress the ROS produced by zooxanthellae. 60 B. Kuguru et al. / Journal of Experimental Marine Biology and Ecology 394 (2010) 53–62

Both our laboratory (Fig. 5a, b) and field observations (Fig. 5c, d) rhodostoma polyps of efficient mechanisms for quenching excitation indicate that individuals of D. unguja exhibit behavioral mechanisms energy in their photosynthetic apparatus, and amelioration of ROS in to avoid the damaging effects of high solar radiation (UVB). symbiont and host cells, all cause members of this species to be less Corallimorpharians locomote at slow rates of b10 mm/month due to susceptible to high irradiance. their lack of basilar muscles on the pedal disc (Chadwick and Adams, We conclude that exposure to UVR is a major factor controlling the 1991). Thus, we observed polyps changing their positions over a depth-related distributional patterns of these two common reef period of weeks rather than hours or days. In shallow water on coral cnidarians. Further studies will be needed to fully understand the reefs, D. unguja occupies shaded areas such as holes, crevices, and the underlying cellular mechanisms and biochemical pathways used by branches of corals (Fig. 5c). In the foliose coral Turbinaria mesenteria, corallimorpharians during adjustment of their photosystems (PSI and the vertical orientation of the plates leads to self-shading of N50% and PSII) and photoprotective pigments in response to irradiance on coral is an effective mechanism of photoprotection to avoid photoinhibition reefs. in shallow areas (Hoogenboom et al., 2008). Accordingly, individuals of D. unguja that live among the branches of T. mesenteria (Fig. 5c) Acknowledgements protect themselves from high irradiance. Furthermore, polyps of D. unguja rapidly withdraw into coral and rock crevices when physically We gratefully acknowledge the staff and students of Dan Tchernov disturbed, and also expand their oral discs upon exposure to low and Maoz Fine of the Interuniversity Institute for Marine Sciences in levels of irradiance but contract when encountering excessive Eilat for technical assistance in various aspects of field and laboratory irradiance (B. L. Kuguru, personal observation). This is similar to the studies. We also thank Gal Dishon for spectral irradiance measure- behavioral reactions of some stony corals (Levy et al., 2006). Many ments and Beverly Goodman for assistance in manuscript preparation. types of host cnidarians adopt behavioral photoprotective mechan- This research was supported by funds from the Bar-Ilan University, isms under high-light conditions (Brown et al., 1994, 2002). Tissue Auburn University, City University of New York, Baruch College, the retraction also has been reported in the Anthopleura Interuniversity Institute for Marine Sciences in Eilat and Israel Science elegantissima as a response to intense sunlight, which not only limits Foundation grant (# 981/05). We also wish to thank the US National exposure to high irradiance but also reduces the production of Science Foundation grant (# 0920572). This research was submitted harmful oxygen radicals (Shick and Dykens, 1984). Marked retraction in partial fulfilment of the requirements for a doctoral degree by B.K. when exposed to high irradiance may be triggered by high levels of − at Bar-Ilan University. Experiments performed in this study complied photosynthetically generated O2 in actinian sea anemones (Shick et with the laws of Israel. [SS] al., 1991) and by photodynamic processes in the tissues of corals (Dykens and Shick, 1984). Such avoidance behavior in anemones (Shick and Dykens, 1984), corals (Brown et al., 1994), and References corallimorpharians (Kuguru et al., 2007) may be likened to the movement of zooxanthellae away from the mesoglea in the Ambarsari, I., Brown, B.E., Barlow, R.G., Britton, G., Cummings, D., 1997. Fluctuations in algal chlorophyll and carotenoid pigments during solar bleaching in the coral corallimorpharian R. rhodostoma (Kuguru et al., 2007), as well as Goniastrea aspera at Phuket, Thailand. Mar. Ecol. Prog. Ser. 159, 303–307. chloroplast movements in plants (Brugnoli and Bjorkqman, 1992) and Austin, J., Butchart, N., Shine, K.P., 1992. Possibility of an Arctic ozone hole in a doubled- seagrasses (Sharon and Beer, 2008), where pronounced reductions in CO2 climate. Nature 360, 221–225. fl Banaszak, A.T., Lesser, M.P., 2009. Effects of solar ultraviolet radiation on coral reef chlorophyll uorescence have been noted during leaf-folding chloro- organisms. Photochem. Photobiol. Sci. 8, 1276–1294. plast re-orientation at high irradiance. In-situ experimental studies on Banaszak, A.T., LaJeunesse, T.C., Trench, R.K., 2000. The synthesis of mycosporine-like the effects of UVR on the behavior and recruitment of the stony coral amino acids (MAAs) by cultured, symbiotic dinoflagellates. J. Exp. Mar. Biol. Ecol. 249, 219–233. Porites asteoides showed that many larvae swam and settled in Baruch, R., Avishai, N., Rabinowitz, C., 2005. UV incites diverse levels of DNA breaks in habitats where UVR was reduced (Gleason and Wellington, 1995; different cellular compartments of a branching coral species. J. Exp. Biol. 208, Gleason et al., 2006). Thus, behavioral avoidance of habitats with 843–848. biologically damaging levels of UVR may be a major factor Behrenfeld, M., Prasil, O., Kolber, Z., Babin, M., Falkowski, P.G., 1998. Compensatory changes in photosystem II electron turnover rates protect photosynthesis from contributing to the successful recruitment of coral larvae. As a driving photoinhibition. Photosynth. Res. 58, 259–268. for extensive behavioral, physiological, and cellular adaptations Bhagooli, R., Hidaka, M., 2003. Comparison of stress susceptibility of in hospite and fi in benthic marine invertebrates, UVR may be a more important factor isolated zooxanthellae among ve coral species. J. Exp. Mar. Biol. Ecol. 291, 181–197. than previously recognized in determining community structure in Bradford, M.M., 1976. Rapid and sensitive method for quantitation of microgram aquatic ecosystems (Hader et al., 2007). It appears that the two quantities of protein utilizing principle of protein-dye binding. Anal. Biochem. 72, corallimorpharians examined here respond to UVR by exerting major 248–254. Brown, B.E., Le Tissier, M.D.A., Dunne, R.P., 1994. Tissue retraction in the scleractinian control over their patterns of microhabitat use and depth distributions coral Coeloseris mayeri, its effect upon coral pigmentation, and preliminary on reefs. implications for heat balance. Mar. Ecol. Prog. Ser. 105, 209–218. Some of the differences that we observed in photosynthetic Brown, B.E., Ambarsari, I., Warner, M.E., Fitt, W.K., Dunne, R.P., Gibb, S.W., Cummings, D. G., 1999. Diurnal changes in photochemical efficiency and xanthophyll concentra- performance between these corallimorpharians may be due to the tions in shallow water reef corals: evidence for photoinhibition and photoprotec- differences in the mechanisms to adjust photoprotective versus non- tion. Coral Reefs 18, 99–105. photosynthetic pigments in their light-harvesting antennae (Iglesias- Brown, B.E., Downs, C.A., Dunne, R.P., Gibb, S.W., 2002. Preliminary evidence for tissue retraction as a factor in photoprotection of corals incapable of xanthophyll cycling. Prieto and Trench, 1997). The ratio of light-protective pigments in the J. Exp. Mar. Biol. Ecol. 277, 129–144. xanthophyll cycle, diadinoxanthin (DD), to the sum of DD and Brugnoli, E., Bjorkqman, O., 1992. Chloroplast movements in leaves: influence on fl diatoxanthin (Dt), shows linear correspondence with σPSII, i.e., as DD chlorophyll uorescence and measurements of light-induced absorbence changes – de-epoxidizes (forms a quencher) to DT, the σ decreases (Kolber related to delta-pH and zeaxanthin formation. Photosynth. Res. 32, 23 35. PSII Chadwick, N.E., Adams, C., 1991. Locomotion, and killing of corals and Falkowski, 1993). Expression of low values of σPSII and high Qm at by the corallimorpharian Corynactis californica. Hydrobiologia 216 (217), 263–269. HLUVT suggests that D. unguja zooxanthellae exhibited a deficiency in Chadwick-Furman, N.E., Spiegel, M., 2000. Abundance and clonal replication in the – either de-epoxidation or epoxidation of their xanthophyll pigments tropical corallimorpharian Rhodactis rhodostoma. Invertebr. Biol. 119, 351 360. Coffroth, M.A., Santos, S.R., 2005. Genetic diversity of symbiotic dinoflagellates in the (Niyogi et al., 1997). The inability of D. unguja zooxanthellae to Symbiodinium. Protist 156, 19–34. dissipate excessive light energy apparently caused a reduction in their den-Hartog, J., 1980. Caribbean shallow water Corallimorpharia. Zool Verh. 176, 1–82. Fv/Fm at HLUVT, and higher values of host LPO and ROS at HL/LLUVT, Downs, C.A., Fauth, J.E., Halas, J.C., Dustan, P., Bemiss, J., Woodley, C.M., 2002. Oxidative stress and seasonal coral bleaching. Free Radic. Biol. Med. 33, 533–543. indicating that UVR, especially UVB, caused oxidative damage to both Dunlap, W.C., Chalker, B.E., 1986. Identification and quantitation of near-UV absorbing the photosynthetic apparatus and host cells. The presence in R. compounds (S-320) in a hermatypic scleractinian. Coral Reefs 5, 155–159. B. Kuguru et al. / Journal of Experimental Marine Biology and Ecology 394 (2010) 53–62 61

Dykens, J.A., Shick, J.M., 1984. Photobiology of the symbiotic sea-anemone, Anthopleura Jeffrey, S.W., Humphrey, G.F., 1975. New spectrophotometric equations for determining elegantissima defenses against photodynamic effects, and seasonal photoacclima- chlorophylls A, B, C1 and C2 in higher-plants, algae and natural phytoplankton. tization. Biol. Bull. 167, 683–697. Biochem. Physiol. Pfl 167, 191–194. Fagoonee, I., Wilson, H.B., Hassell, M.P., Turner, J.R., 1999. The dynamics of Jokiel, P.L., Lesser, M.P., Ondrusek, M.E., 1997. UV-absorbing compounds in the coral zooxanthellae populations: a long-term study in the field. Science 283, 843–845. Pocillopora damicornis: interactive effects of UV radiation, photosynthetically active Falkowski, P.F., Chen, Y. (Eds.), 2003. Photoacclimation of light harvesting systems in radiation, and water flow. Limnol. Oceanogr. 42, 1468–1473. eukaryotic algae, Vol. Kluwer academic publishers. Jones, A.M., Berkelmans, R., van Oppen, M.J.H., Mieog, J.C., Sinclair, W., 2008. A Falkowski, P.G., Dubinsky, Z., 1981. Light-shade adaptation of Stylophora pistillata,a community change in the algal endosymbionts of a scleractinian coral following a hermatypic coral from the Gulf of Eilat. Nature 289, 172–174. natural bleaching event: field evidence of acclimatization. Proc. R. Soc. B. Biol. Sci. Falkowski, P.G., Kolber, Z., Fujita, Y., 1988. Effect of redox state on the dynamics 275, 1359–1365. photosystem-Ii during steady-state photosynthesis in eukaryotic algae. Biochim Kana, T.M., 1992. Relationship between photosynthetic oxygen cycling and carbon Biophys Acta 933, 432–443. assimilation in Synechococcus Wh7803 (Cyanophyta). J. Phycol. 28, 304–308. Fitt, W.K., McFarland, F.K., Warner, M.E., Chilcoat, G.C., 2000. Seasonal patterns of tissue Kinzie III, R.A., 1993. Effects of ambient levels of solar ultraviolet radiation on biomass and densities of symbiotic dinoflagellates in reef corals and relation to zooxanthellae and photosynthesis of the reef coral Montipora verrucosa. Mar. Biol. coral bleaching. Limnol. Oceanogr. 45, 677–685. 116, 319–327. Fukami, H., Chen, C.A., Budd, A.F., Collins, A., Wallace, C., Chuang, Y.-Y., Chen, C., Dai, C.-F., Kolber, Z., Falkowski, P.G., 1993. Use of active fluorescence to estimate phytoplankton Iwao, K., Sheppard, C., Knowlton, N., 2008. Mitochondrial and nuclear genes suggest photosynthesis in-situ. Limnol. Oceanogr. 38, 1646–1665. that stony corals are monophyletic but most families of stony corals are not (Order Kolber, Z., Wyman, K.D., Falkowski, P.G., 1990. Natural variability in photosynthetic Scleractinia, Class , Phylum ). PLoS ONE 3 (9), 1–9. energy conversion efficiency: a field study in the Gulf of Maine. Limnol. Oceanogr. Gates, R.D., Edmunds, P.J., 1999. The physiological mechanisms of acclimitization in 35, 72–79. tropical reef corals. Am. Zool. 39, 30–43. Kolber, Z.S., Prasil, O., Falkowski, P.G., 1998. Measurements of variable chlorophyll Gleason, D.F., Wellington, G.M., 1993. Ultraviolet-radiation and coral bleaching. Nature fluorescence using fast repetition rate techniques: defining methodology and 365, 836–838. experimental protocols. BBA Bioenerg. 1367, 88–106. Gleason, D.F., Wellington, G.M., 1995. Variation in UVB sensitivity of planula larvae of Kuguru, B.L., Mgaya, Y.D., Ohman, M.C., Wagner, G.M., 2004. The reef environment and the coral Agaricia agaricites along a depth gradient. Mar. Biol. 123, 693–703. competitive success in the Corallimorpharia. Mar. Biol. 145, 875–884. Gleason, D.F., Edmunds, P.J., Gates, R.D., 2006. Ultraviolet radiation effects on the Kuguru, B., Winters, G., Beer, S., Santos, S., Chadwick, N., 2007. Adaptation strategies of behavior and recruitment of larvae from the reef coral Porites astreoides. Mar. Biol. the corallimorpharian Rhodactis rhodostoma to irradiance and temperature. Mar. 148, 503–512. Biol. 151, 1287. Glynn, P.W., Dcroz, L., 1990. Experimental evidence for high-temperature stress as the Kuguru, B., Chadwick, N.E., Achituv, Y., Zendbank, K., Tchernov, D., 2008. Mechanisms of cause of El-Nino-coincident coral mortality. Coral Reefs 8, 181–191. habitat segregation between corallimorpharians: photosynthetic parameters and Gorbunov, M.Y., Kolber, Z.S., Lesser, M.P., Falkowski, P.G., 2001. Photosynthesis and Symbiodinium types. Mar. Ecol. Prog. Ser. 369, 115–129. photoprotection in symbiotic corals. Limnol. Oceanogr. 46, 75–85. LaJeunesse, T.C., 2002. Diversity and community structure of symbiotic dinoflagellates Goulet, T.L., Cook, C.B., Goulet, D., 2005. Effect of short-term exposure to elevated from Caribbean coral reefs. Mar. Biol. 141, 387–400. temperatures and light levels on photosynthesis of different host-symbiont Lesser, M.P., 1989. Cu–Zn superoxide dismutase in photoautotrophic combinations in the Aiptasia pallidal Symbiodinium . Limnol. Oceanogr. symbioses: photoadaptation and an evolutionary perspective on the origin of the 50, 1490–1498. eukaryotic chloroplast. Am. Zool. 29, A82. Greenberg, B.M., Gaba, V., Canaani, O., Malkin, S., Mattoo, A.K., Edelman, M., 1989. Lesser, M.P., 2000. Depth-dependent photoacclimatization to solar ultraviolet radiation Separate photosensitizers mediate degradation of the 32-kDa photosystem II in the Caribbean coral Montastraea faveolata. Mar. Ecol. Prog. Ser. 192, 137–151. reaction center protein in the visible and UV spectral regions. Proc. Natl Acad. Sci. Lesser, M.P., 2006. Oxidative stress in marine environments: biochemistry and USA 86, 6617–6620. physiological ecology. Ann. Rev. Physiol. 68, 253–278. Hader, D.P., Kumar, H.D., Smith, R.C., Worrest, R.C., 2007. Effects of solar UV radiation on Lesser, M.P., Lewis, S., 1996. Action spectrum for the effects of U.V. radiation on photosynthesis aquatic ecosystems and interactions with climate change. Photochem. Photobiol. in the hermatypic coral Pocillopora damicornis. Mar. Ecol. Prog. Ser. 134, 171–177. Sci. 6, 267–285. Lesser, M.P., Shick, J.M., 1989. Effects of irradiance and ultraviolet radiation on Harley, C.D.G., Hughes, R.A., Hultgren, K.M., Miner, B.G., Sorte, C.J.B., Thornber, C.S., photoadaptation in the zooxanthellae of Aiptasia pallida: primary production, Rodriguez, L.F., Tomanek, L., Williams, S.L., 2006. The impacts of climate change in photoinhibition, and enzymic defenses against . Mar. Biol. 102, coastal marine systems. Ecol. Lett. 9, 228–241. 243–255. Hoegh-Guldberg, O., 1999. Climate change, coral bleaching and the future of the world's Lesser, M.P., Stochaj, W.R., 1990. Photoadaptation and protection against active forms of coral reefs. Mar. Freshwater Res. 50, 839–866. oxygen in the symbiotic prokaryote Prochloron sp and its ascidian host. Appl. Hoegh-Guldberg, O., Smith, G.J., 1989. The effect of sudden changes in temperature, Environ. Microb. 56, 1530–1535. light and salinity on the population density and export of zooxanthellae from the Levy, O., Achituv, Y., Yacobi, Y.Z., Stambler, N., Dubinsky, Z., 2006. The impact of spectral reef corals Stylophora pistillata Esper and Seriatopora hystrix Dana. J. Exp. Mar. Biol. composition and light periodicity on the activity of two antioxidant enzymes (SOD Ecol. 129, 279–303. and CAT) in the coral Favia favus. J. Exp. Mar. Biol. Ecol. 328, 35–46. Hoegh-Guldberg, O., Jones, R.J., Ward, S., Loh, W.K., 2002. Ecology—is coral bleaching Loya, Y., Sakai, K., Yamazato, K., Nakano, Y., Sambali, H., Van Woesik, R., 2001. Coral really adaptive? Nature 415, 601–602. bleaching: the winners and the losers. Ecol. Lett. 4, 122–131. Hoegh-Guldberg, O., Mumby, P.J., Hooten, A.J., Steneck, R.S., Greenfield, P., Gomez, E., Lyons, M.M., Aas, P., Pakulski, J.D., Van Waasbergen, L., Miller, R.V., Mitchell, D.L., Jeffrey, Harvell, C.D., Sale, P.F., Edwards, A.J., Caldeira, K., Knowlton, N., Eakin, C.M., Iglesias- W.H., 1998. DNA damage induced by ultraviolet radiation in coral reef microbial Prieto, R., Muthiga, N., Bradbury, R.H., Dubi, A., Hatziolos, M.E., 2007. Coral reefs communities. Mar. Biol. 130, 537–543. under rapid climate change and ocean acidification. Science 318, 1737–1742. Malanga, G., Calmanovici, G., Puntarulo, S., 1997. Oxidative damage to chloroplasts from Hoogenboom, M.O., Sean, R.C., Kenneth, R.A., 2008. Interactions between morpholog- Chlorella vulgaris exposed to ultraviolet-B radiation. Physiol. Plant. 101, 455–462. ical and physiological plasticity optimize energy acquisition in corals. Ecology 89, Mass, T., Einbinder, S., Brokovich, E., Shashar, N., Vago, R., Erez, J., Dubinsky, Z., 2007. 1144–1154. Photoacclimation of Stylophora pistillata to light extremes: and Huner, N.P.A., Maxwell, D.P., Gray, G.R., Savitch, L.V., Krol, M., Ivanov, A.G., Falk, S., 1996. calcification. Mar. Ecol. Prog. Ser. 334, 93–102. Sensing environmental temperature change through imbalances between energy McKenzie, R.L., Paulin, K.J., Madronich, S., 1998. Effects of show cover on UV irradiance supply and energy consumption: redox state of photosystem II. Physiol. Plant. 98, and surface albedo: a case study. J. Geophys. Res. Atmos. 103, 28785–28792. 358–364. Medina, M., Collins, A.G., Takaoka, T.L., Kuehl, J.V., Boore, J.L., 2006. Naked corals: Iglesias-Prieto, R., Trench, R.K., 1994. Acclimation and adaptation to irradiance in skeleton loss in scleractinia. Proc. Natl Acad. Sci. USA 103, 9096–9100. symbiotic dinoflagellates: responses of the photosynthetic unit to changes in Melis, A., Nemson, J.A., Harrison, M.A., 1992. Damage to functional components and photon flux density. Mar. Ecol. Prog. Ser. 113, 163–175. partial degradation of photosystem-II reaction center proteins upon chloroplast Iglesias-Prieto, R., Trench, R.K., 1997. Acclimation and adaptation to irradiance in exposure to ultraviolet-B radiation. Biochim. Biophys. Acta 1100, 312–320. symbiotic dinoflagellates. II. Response of chlorophyll–protein complexes to Mortainbertrand, A., Bennett, J., Falkowski, P.G., 1990. Photoregulation of the light- different photon-flux densities. Mar. Biol. 130, 23–33. harvesting chlorophyll protein complex associated with photosystem-II in Iglesias-Prieto, R., Beltran, V.H., LaJeunesse, T.C., Reyes-Bonilla, H., Thome, P.E., 2004. Dunaliella tertiolecta: evidence that apoprotein abundance but not stability requires Different algal symbionts explain the vertical distribution of dominant reef corals in chlorophyll synthesis. Plant Physiol. 94, 304–311. the eastern Pacific. Proc. R. Soc. London, Ser. B. Biol. 271, 1757–1763. Muhando, C.A., Kuguru, B.L., Wagner, G.M., Mbije, N.E., Ohman, M.C., 2002. IPCC, 2001. Climate Change 2001: Synthesis report. Contribution of Working Groups I, II Environmental effects on the distribution of corallimorpharians in Tanzania. and III to the Third Assessment Report of the Intergovernmental Panel on Climate Ambio 31, 558–561. Change. In: Watson, R.T., the Core Writing Team (Eds.), Cambridge University Press, Muscatine, L., McCloskey, L.R., Marian, R.E., 1981. Estimating the daily contribution of Cambridge. carbon from zooxanthellae to coral . Limnol. Oceanogr. 26, IPCC, 2007. Summary for Policymakers. In: Solomon, S., Qin, D., Manning, M., Chen, Z., 601–611. Marquis, M., Averyt, K.B., Tignor, M., Miller, H.L. (Eds.), Climate Change 2007: The Niyogi, K.K., Bjorkman, O., Grossman, A.R., 1997. The roles of specific xanthophylls in Physical Science Basis. Contribution of Working Group I to the Fourth Assessment photoprotection. Proc. Natl Acad. Sci. USA 94, 14162–14167. Report of the Intergovernmental Panel on Climate Change. Cambridge University Perez, S., Weis, V., 2006. Nitric oxide and cnidarian bleaching: an eviction notice Press, Cambridge, United Kingdom and New York, NY, USA. mediates breakdown of a symbiosis. J. Exp. Biol. 209, 2804–2810. Iwanzik, W., Tevini, M., Dohnt, G., Voss, M., Weiss, W., Graber, P., Renger, G., 1983. Robison, J.D., Warner, M.E., 2006. Differential impacts of photoacclimation and thermal Action of UV-B radiation on photosynthetic primary reactions in spinach- stress on the photobiology of four different phylotypes of Symbiodinium chloroplasts. Physiol. Plant. 58, 401–407. (Pyrrhophyta). J. Phycol. 42, 568–579. 62 B. Kuguru et al. / Journal of Experimental Marine Biology and Ecology 394 (2010) 53–62

Salih, A., Larkum, A., Cox, G., Kuhl, M., Hoegh-Guldberg, O., 2000. Fluorescent pigments Stambler, N., 2005. Bio-optical properties of the northern Red Sea and the Gulf of Eilat in corals are photoprotective. Nature 408, 850–853. (Aqaba) during winter 1999. J. Sea Res. 54, 186–203. Savage, A.M., Trapido-Rosenthal, H., Douglas, A.E., 2002. On the functional significance Stanley, G.D., Fautin, D.G., 2001. The Origins of Modern Corals. Science 291, 1913–1914. of molecular variation in Symbiodinium, the symbiotic algae of Cnidaria: Stat, M., Carter, D., Hoegh-Guldberg, O., 2006. The evolutionary history of Symbiodi- photosynthetic response to irradiance. Mar. Ecol. Prog. Ser. 244, 27–37. nium and scleractinian hosts: symbiosis, diversity, and the effect of climate change. Sharon, Y., Beer, S., 2008. Diurnal movements of chloroplasts in Halophila stipulacea and Perspect. Plant Ecol. Evol. Syst. 8, 23–43. their effect on PAM fluorometric measurements of photosynthetic rates. Aquat. Bot. Stoletzki, N., Schierwater, B., 2005. Genetic and color morph differentiation in the 88, 273–276. Caribbean sea anemone Condylactis gigantea. Mar. Biol. 147, 747–754. Shick, J.M., Dykens, J.A., 1984. Photobiology of the symbiotic sea anemone Anthopleura Strauss, R.E., 1979. Reliability estimates for Ivlev's electivity index, the forage ratio, and elegantissima: photosynthesis, respiration, and behavior under intertidal condi- a proposed linear index of food selection. Trans. Am. Fish Soc. 108, 344–352. tions. Biol. Bull. 166, 608–619. Tchernov, D., Gorbunov, M.Y., de Vargas, C., Yadav, S.N., Milligan, A.J., Haggblom, M., Shick, J.M., Lesser, M.P., Stochaj, W.R., 1991. Ultraviolet radiation and photooxidative Falkowski, P.G., 2004. Membrane lipids of symbiotic algae are diagnostic of stress in zooxanthellate anthozoa the sea anemone Phyllodiscus semoni and the sensitivity to thermal bleaching in corals. Proc. Natl Acad. Sci. USA 101, octocoral Clavularia Sp. Symbiosis 10, 145–173. 13531–13535. Shick, J.M., Lesser, M.P., Jokiel, P.L., 1996. Effects of ultraviolet radiation on corals and Warner, M.E., Berry-Lowe, S., 2006. Differential xanthophyll cycling and photochemical other coral reef organisms. Global Change Biol. 2, 527–545. activity in symbiotic dinoflagellates in multiple locations of three species of Shick, J.M., Romaine-Lioud, S., Ferrier-Pages, C., Gattuso, J.P., 1999. Inhibition of the Caribbean coral. J. Exp. Mar. Biol. Ecol. 339, 86–95. shikimate pathway blocks UVB radiation-stimulated accumulation of mycospor- Warner, M.E., Chilcoat, G.C., McFarland, F.K., Fitt, W.K., 2002. Seasonal fluctuations in ine-like amino acids (MAAs) in the coral Stylophora pistillata.Photochem. the photosynthetic capacity of photosystem II in symbiotic dinoflagellates in the Photobiol. 69, 93S. Caribbean reef-building coral Montastraea. Mar. Biol. 141, 31–38. Sinha, R.P., Hader, D.P., 2002. Life under solar UV radiation in aquatic organisms. Adv. Work, T.M., Aeby, G.S., Maragos, J.E., 2008. Phase shift from a coral to a corallimorph- Space Res. 30, 1547–1556. dominated reef associated with a shipwreck on Palmyra Atoll. PLoS ONE 3 (8), 1–5. Smith, D.J., Suggett, D.J., Baker, N.R., 2005. Is photoinhibition of zooxanthellae photosynthesis the primary cause of thermal bleaching in corals? Glob Change Biol. 11, 1–11.