Invertebrate Biology 124(3): 185193. r 2005 American Microscopical Society, Inc. DOI: 10.1111/j.1744-7410.2005.00018.

The effect of temperature on the size and population density of dinoflagellates in larvae of the reef coral Porites astreoides
Peter J. Edmunds,1,a Ruth D. Gates,2 William Leggat,3 Ove Hoegh-Guldberg,3 and Laurie Allen-Requa1
2

Department of Biology, California State University, Northridge, CA 91330-8303, USA Hawaii Institute of Marine Biology, University of Hawaii Manoa, Kaneohe, HI 96744-1346, USA 3 Centre for Marine Sciences, University of Queensland, St. Lucia, Queensland, Australia

Abstract. Pre-settlement events play an important role in determining larval success in marine invertebrates with bentho-pelagic life histories, yet the consequences of these events typically are not well understood. The purpose of this study was to examine the pre-settlement impacts of different seawater temperatures on the size and population density of dinoagellate symbionts in brooded larvae of the Caribbean coral Porites astreoides. Larvae were collected from P. astreoides at 1420 m depth on Conch Reef (Florida) in June 2002, and incubated for 24 h at 15 temperatures spanning the range 25.1130.01C in mean increments of 0.470.11C (7SD). The most striking feature of the larval responses was the magnitude of change in both parameters across this 51C temperature range within 24 h. In general, larvae were largest and had the highest population densities of Symbiodinium sp. between 26.41 27.71C, and were smallest and had the lowest population densities at 25.81C and 28.81C. Larval size and symbiont population density were elevated slightly (relative to the minimal values) at the temperature extremes of 25.11C and 301C. These data demonstrate that coral larvae are highly sensitive to seawater temperature during their pelagic phase, and respond through changes in size and the population densities of Symbiodinium sp. to ecologically relevant temperature signals within 24 h. The extent to which these changes are biologically meaningful will depend on the duration and frequency of exposure of coral larvae to spatiotemporal variability in seawater temperature, and whether the responses have cascading effects on larval success and their entry to the post-settlement and recruitment phase.
Additional key words: scleractinia, coral, larvae, temperature, Porites

The success of the pelagic larvae of benthic marine invertebrates pivots around pre-settlement events mediated by the physical environment and/or biotic pressures (Thorson 1950; Rumrill 1990; Morgan 1995). Such events can create patterns of variability in the distribution of recruits (Reed et al. 2000; Underwood & Keough 2001) and play important roles in mediating population dynamics through post-settlement events, recruitment, and adult success (Caley et al. 1996; Gosselin & Qian 1997). The roles of postsettlement events in the population dynamics of benthic marine invertebrates are relatively well understood (Gosselin & Qian 1997; Hunt & Sheibling 1997), but progress in understanding pre-settlement

Author for correspondence. E-mail: peter.edmunds@csun.edu

events has been considerably slower (Rumrill 1990; Morgan 1995, 2001; Underwood & Keough 2001), in part because of the difculty of studying larvae under ecologically relevant conditions. In those systems where study has been possible, aspects such as the food reserves of settling larvae play a very decisive role in post-settlement success (e.g., Emlet & HoeghGuldberg 1996; Hoegh-Guldberg & Emlet 1997). Two critical aspects of the biology of pelagic larvae, maturation and dispersal, are strongly inuenced by life history strategy, the size of food reserves, and whether or not feeding is possible (Thorson 1950; Strathmann 1985; Levin & Bridges 1995). Additionally, the time required for maturation is related to the rate and magnitude of the developmental changes necessary before settlement occurs (Harrison & Wallace 1990; Levin & Bridges 1995), and the degree to which these changes are dynamically shaped by the

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Edmunds, Gates, Leggat, Hoegh-Guldberg, & Allen-Requa LAMARCK 1816. To this end, we incubated larvae across a gradient of 15 temperatures separated by 0.470.11C (7SD) between 25.1130.01C, which is ecologically relevant for the study reef and to the periods of warmer than normal seawater seen over the past 30 years (Leichter et al. 1996; Hoegh-Guldberg 1999). The responses to these temperatures were assessed by measuring larval size and the population densities of algal symbionts after 24 h. This duration was selected because it is relatively lengthy compared to both the rate of development of coral larvae (Ball et al. 2002) and the time necessary for larvae to have a high chance of encountering water at different temperatures on the study reef (Leichter et al. 1996). Our results are notable in that they demonstrate rapid changes in larval size and population density of algal symbionts in response to relatively small changes in seawater temperature.

environment. Studies of the development of invertebrate larvae and the interactive effects of environmental signals are particularly challenging in tropical reef corals, because analyses of the early development of symbiotic corals involves teasing apart the complex phenotypic consequences of both host and symbiont genomes. However, there are substantial rewards for tackling this challenge, because the development of cnidarians provides clues to the control of morphological complexity in more derived metazoans (Miller et al. 2000; Ball et al. 2002), and potentially could provide much-needed insights into the roles of coral larvae in determining the trajectories of change in coral populations (Hughes et al. 2000). Scleractinian corals that broadcast spawn release gametes into the water column, where fertilization occurs, and thereafter, where the zygote develops until forming a settlement-competent larva (Harrison & Wallace 1990). Cleavage and larval development occur quickly in these corals, typically producing well-developed larvae in which a mouth and mesenteries appear within 4896 h (Babcock & Heyward 1986; Schwarz et al. 1999; Ball et al. 2002; Miller & Mundy 2003). These larvae can metamorphose within 90 h of fertilization (Miller & Mundy 2003), but most require a minimum of 96144 h before they are competent to settle (Harrison & Wallace 1990). Brooding species, on the other hand, release mature larvae that have the potential to settle rapidly (Harrison & Wallace 1990). Less is known about the actual development of these brooded larvae, but fertilized eggs can develop through the planula stage within 96 h of fertilization (Szmant-Froelich et al. 1985), which is similar to the development rates for broadcast-spawners. Thereafter, brooded larvae are retained 23 weeks before release (Szmant-Froelich et al. 1985; Chornesky & Peters 1987), after which they may settle within minutes (Carlon & Olsen 1993), or spend hours to upwards of 100 days in the plankton (Richmond 1987; Harii et al. 2002). Ocean temperatures have increased by as much as 1.51C since the late 19th century. These changes have led to periods in which the temperature of tropical oceans has periodically exceeded long-term maxima and has led to extensive periods of mass coral bleaching and mortality (Glynn 1993; Hoegh-Guldberg 1999). So far, studies of this phenomenon have focused almost exclusively on the adult phase of corals, and only a few have considered the potential impacts of warming seas on larval phases (Edmunds et al. 2001; Ward et al. 2001). The goal of the present study was to evaluate the short-term impacts of ecologically relevant temperatures on the biology of brooded larvae released by the reef coral Porites astreoides

Methods
Collection
Larvae were collected using demersal plankton nets (Brazeau et al. 1998; Edmunds et al. 2001) placed over 200 colonies of the brown morph of Porites astreoides, located 1420 m deep on Conch Reef, Florida Keys. Sampling was carried out to span the new moon in June 2002, a period towards the end of the spring larval release for this species (McGuire 1998). Starting on June 7 and daily thereafter, all the traps were visually inspected at B10:00 AM for the presence of larvae. When collection tubes were found to contain larvae, they were replaced with fresh tubes and those containing the larvae taken to the surface by divers. These tubes were placed in an opaque plastic barrel lled with 120 L of fresh seawater, and immediately transported to the lab on Key Largo, 13 km from Conch Reef. During transport, the larvae were kept in the dark at ambient temperature, and seawater was allowed to exchange across the 250-mmmesh windows on the side of the collection tubes. Assuming larvae of P. astreoides are released after dark (Edmunds et al. 2001) and close to 2300 h (Edmunds, unpubl. data), they were B11 h post-release when collected. On shore, all the larvae from multiple parent colonies were mixed in a plastic tub and then randomly allocated to the temperature treatments described below.

Temperature treatments
Fifteen temperature treatments were created over a stable thermal gradient established using an

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Temperature and size of coral larvae aluminum block (65 7 9 cm). When designing the experiment, we reasoned that 15 treatment conditions would facilitate statistical analysis with linear regression, an assertion that did not anticipate correctly the curvilinear response of the larvae (that precluded regression analysis). One end of the block was cooled with 24.01C water and the other warmed with 32.21C water. The cool and warm water supplies were constantly re-circulated and their temperature regulated with a laboratory chiller and a heater, respectively. Glass test tubes containing seawater were inserted into 15 pairs of holes spaced evenly across the length of the block to create replicate temperature treatments separated by B0.470.11C (7SD) as follows: 25.11, 25.41, 25.81, 26.01, 26.41, 26.81, 27.21, 27.41, 27.71, 28.11, 28.51, 28.81, 29.21, 29.51, and 30.01C. Incubations were conducted under the uorescent lights in the lab (o10 mmol photons s1 m2) that were switched on for B12 h each day. Thirty test tubes were pre-lled with 5 ml of airsaturated, ltered seawater (0.45 mm, FSW), freshly collected from 10 km offshore on the day of each experiment. Prior to starting the larval incubations, the test tubes were placed in the aluminum block and equilibrated to the incubation temperatures that corresponded to their positions in the block. Two trials were completed, one with larvae collected on June 9, and a second with larvae collected on June 10. For each trial, 5 actively swimming larvae were randomly selected from the pooled larval tub and placed in each of the 30 test tubes. The tubes were left uncovered during the incubation period to facilitate gas exchange across the air-water interface; even without this source of oxygen, the oxygen content of 5 ml of air-saturated seawater is more than enough to meet the 24 h respiratory demand of 5 larvae, assuming their respiration rate is B0.1 nmol O2 min1 larva1 (Edmunds et al. 2001). Each experiment lasted 24 h, and ended by anaesthetizing the larvae with 0.36 M MgCl2 followed by xation in 5% formalin in seawater. The preserved larvae were shipped to California and analyzed for larval size and the population densities of algal symbionts (Symbiodinium sp.). Larval length and diameter were measured using a dissecting microscope (Wild) tted with an eyepiece micrometer. Larval size was expressed as volume and determined geometrically based on the assumption that each larva was an oblate ellipsoid of revolution, or the shape of a watermelon. The population density of algal symbionts per larva was determined by homogenizing each larva with a small Teon tissue grinder in a microfuge tube, pelleting the algal symbionts by centrifugation (14,000 rpm), re-suspending the pellet in 30 mL of FSW, and counting the algal

187 symbionts using a hemocytometer. Because of the small amounts of tissue involved, only 4 replicate counts of algal symbionts were possible for each larva. These counts were averaged to express symbiont population density as algal symbionts larva1.

Statistical analyses
Larval size and algal symbionts larva1 were analyzed with a 3-way, nested ANOVA where trial and temperature were xed factors, replicate incubation tubes were nested within trial and temperature, and individual larvae were treated as replicates. Trial (i.e., time) and temperature were treated as xed factors because the initial goal of the study was to analyze their effects on larvae, an endeavor that was somewhat hindered by the short supply of larvae (see results below for details). Population densities of algal symbionts were log-transformed prior to analysis, and the normality and homoscedasticity assumptions of ANOVA tested with graphical analyses of the residuals. All statistical analyses were completed using Systat 9.0 software. The empirical relationships between temperature and each of larval size and symbionts larva1 were described for the results pooled between trials, using polynomial functions that were tted to the data using a least squares routine in Cricket Graph III software.

Results
Of the 200 colonies of Porites astreoides enclosed in larval traps and checked daily from June 818, only 20 colonies released larvae in varying amounts, and all had stopped spawning by June 11. The arrest in larval release coincided with a change in weather from clear and calm until June 10, to thick clouds, rough seas, and torrential rain thereafter. The deteriorating weather caused the waters over Conch Reef to become dark and turbid. Saturation divers stationed in the Aquarius habitat at 15-m depth reported large pressure disturbances indicating the depth to which the effects of the rough seas penetrated. Of the 300 larvae used in the two trials, only 15 (5%) were not recovered in the preserved samples, and were assumed to have died during the incubation. These putative mortalities were scattered randomly across the temperature treatments and these deaths, together with experimental errors during analysis, resulted in some samples sizes of o5 larvae per incubation tube. Of the larvae recovered at the end of the 24-h incubation period, none appeared to

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Edmunds, Gates, Leggat, Hoegh-Guldberg, & Allen-Requa (Fig. 1C,F). These curves highlight the curvilinear responses and demonstrate that larvae were largest and contained the highest symbiont population densities at intermediate temperatures of 26.8127.41C, 1.510.91C (respectively) lower than the ambient seawater temperature at the time of larval release (28.31C). The smallest larvae with the lowest population densities of algal symbionts were found at temperatures in between these intermediate temperatures and the temperature extremes. The larvae also tended to be slightly bigger and contain slightly more algal symbionts (relative to the smallest sizes) at the extreme temperatures (25.11 and 30.01C). To gain further insight into the potential biological significance of the changes in larval size and symbiont density, we statistically compared the 3 extreme values (i.e., 1 high and 2 lows) for each variable using ANOVA. These analyses reveal that larval size varies

have settled or started metamorphosis, and all were actively swimming and appeared healthy. Graphical analyses of larval size and population densities of algal symbionts reveal large differences across very narrow temperature ranges (Fig. 1). Although these responses for both traits were broadly similar in shape (i.e., with regards to the relative position on the temperature scale for the maxima and minima in response variables, Fig. 1), they were more strongly developed in Trial 2 than in Trial 1, as indicated by the significant effect of trial for content of algal symbionts (Table 1). To provide a summary of the overall response of larvae of Porites astreoides to the temperature treatments, the data for each trait were pooled between trials, and the larval size and symbiont population densities as a function of temperature were tted with 5th order (r2 5 0.86), or 4th order (r2 5 0.63) polynomial curves, respectively

Fig. 1. AC. Volume (i.e., size), DF. population density, of symbionts of larvae of Porites astreoides exposed to 15 temperatures for 24 h. Values are presented as means 7 SE (sample sizes range 610 per trial and 1520 for the pooled results), and were calculated from original measures for larval volume, and from log-transformed data followed by backtransformation of the descriptive statistics for symbiont population density per larva; the asymmetric error bars in F are a consequence of these transformations. The bars in panels A, B, D, and E are arranged with their midpoints aligned with the treatment temperatures on the abscissa in order to illustrate the absolute difference between treatments, which is not constant. Polynomial functions were tted to mean values in C and F, and r2 values display the coefcients of determination for each curve. A 5th order curve was tted to the larval size data [(C), y 5 0.006x510.882x448.678x31 1342.396x218487.042x1101715.792] and a 4th order curve was tted to the algal symbiont data [(F), y 5 160.0x4 17609x31725697x213272767x190908254]. Dashed vertical line in C and F indicate ambient seawater temperature (28.31C) when the larvae were released.

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Temperature and size of coral larvae

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Table 1. Results of 3-way, nested ANOVAs comparing the size and symbiont population density of larvae of Porites astreoides among temperatures and trials (both xed factors), and tube (a random factor nested within temperature and trial). The normality and homoscedasticity assumptions of ANOVA were tested through graphical analyses of the residuals. See text for further details and Fig. 1 for graphical presentation of the results. Variable Larval size (volume) Source Temp Trial Trial Temp Tube (Trial Temp) Error Temp Trial Trial Temp Tube (Trial Temp) Error SS 0.777 0.029 0.327 1.057 5.525 11.045 10.745 10.230 28.962 64.224 d.f. 14 1 14 30 221 14 1 14 30 204 MS 0.056 0.029 0.023 0.035 0.025 0.789 10.745 0.731 0.965 0.315 F 1.576 0.818 0.663 1.409 0.817 11.130 0.757 3.067 P 0.144 0.373 0.790 0.086 0.646 0.002 0.704 o.001

Algal symbionts/larva

significantly between the maximum (26.81C) and the two minima (28.81C and 25.81C) (F 5 8.089, df 5 2,52, P 5 .001), and the population densities of algal symbionts differ significantly between the low points of the minima (25.81C and 29.21C), and the high point of the maximum (27.41C) (F 5 7.140, df 5 2,53, P 5 0.002). Although the result of these analyses must be interpreted cautiously because of the explicit choice of values for comparison, the significant outcomes lend support to a biological basis (rather than an artifact of random variability) for the curvilinear response (i.e., Fig. 1).

Discussion
This study tests the response of larvae of Porites astreoides to conditions that they are likely to encounter on a reef where seawater temperatures uctuate on a scale of hours to days (Leichter et al. 1996), or when they are carried, for example, into a warm bay or backreef location. They also represent the types of seawater temperatures that are likely within a few decades due to global climate change. Our results are significant in demonstrating that both the animal (reected largely in the changes in larval size) and the algal fractions (reected in the change in the population densities of algal symbionts) display similar, non-linear responses to temperature treatments lasting only 24 h. Given the half-century since Thorson (1950) underscored the important effects of temperature on pelagic larvae, and the numerous studies that since have quantied such effects (e.g., Pechenik 1984; Hoegh-Guldberg & Pearse 1995; Miller & Emlet 1999; Drent 2002), it is not surprising that temperature affects coral larvae and perhaps mediates their development (discussed below). However,

the speed and complexity with which organismic traits of coral larvae (i.e., their size and symbiont content) were modied by temperature was unexpected. The rapidity of these responses draws attention to the potential importance of environmental signals in modifying the molecular mechanisms involved in the development of coral larvae (e.g., Ball et al. 2002), and the possibility for rapid biochemical responses (for example, occurring on a time scale of seconds [Gorbunov et al. 2001]) to have cascading effects that manifest at the organismic level on a very short time scale. Moreover, the similarity in responses to temperature of the size of the larvae and the density of algal symbionts suggests that the functions of the animal host and the algal symbiont are integrated closely, even in early life stages of reef corals. Ecologically, the importance of these ndings lie in understanding how the pre-settlement success of coral larvae is affected by physical factors including (but not limited to) exposure to increased seawater temperatures associated with global climate change (Hoegh-Guldberg 1999; IPCC 2001). Unfortunately, we cannot be sure to what extent the changes in larvae of Porites astreoides reect growth and/or shrinkage, as we do not know their size and content of algal symbionts when they rst were collected. This omission was driven by differences between our planned experiment and the reality of what was experimentally possible given the weak spawning of P. astreoides in June 2002. We feel it is unlikely that the trends (Fig. 1) are procedural artifacts because larvae were pooled among colonies and allocated randomly among treatments to prevent confounding effects of genotype, and any preservation artifacts (e.g., caused by post-xation shrinkage) would uniformly affect larvae regardless of temperature.

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Edmunds, Gates, Leggat, Hoegh-Guldberg, & Allen-Requa et al. 1983). Clearly, these calculations provide only weak inferential support for the role played by cell division of algal symbionts in generating the present results. Nevertheless, there is sufcient evidence to conclude that both cell loss and division probably were responsible for the dynamic responses of larvae of P. astreoides to temperature. Such changes could evoke a positive feedback effect on larvae size, because changes in symbiont population density would alter the capacity for the symbionts to support photoautotrophic nutrition of the larvae (Richmond 1987), thereby mediating differences in larval size through differential supply of carbon. In the present study, however, such effects are unlikely to have played an important role in determining larvae size, because the incubations were completed under low light levels (o10 mmol photons s1 m2) that would curtail photosynthesis. Where daytime light levels would be considerably higher, the size of the symbiont population in coral larvae could play a substantial role in determining larval size. The changes in the size of larvae of Porites astreoides could result from one or more mechanisms mediating either growth and/or shrinkage. Of the potentially important mechanismsvariation in the rates of cell division (or its arrest), osmotic effects, and/or anabolic and catabolic processescurrently we favor changes in overall rates of development (i.e., growth and cell division) and perhaps osmotic effects for their ability to account for the observed results. While aspects of the present results probably reect the effects of temperature on the development of invertebrate larvae (Thorson 1950), there are few quantitative data regarding the potential stimulation of larval development by temperature within the scleractinian corals. However, the relationship has been reported anecdotally (Ball et al. 2002), and there is inferential support for this effect from a variety of other situations. For example, the settlement of scleractinian larvae is related positively to temperature over a range of values in the laboratory (Jokiel & Guinther 1978; Coles 1985), and ecological data from the Virgin Islands suggest similar effects occur in hospite (Edmunds 2004). For the deep-water scleractinian coral Oculina varicose, off the Florida coast, temperature positively affects larval swimming speed (Brooke & Young 2003), and for the soft coral Heteroxenia fuscescens in Eilat (Red Sea), the rate of larval metamorphosis is high in warmer compared to cooler months of the year (Ben David-Zaslow & Beneyahu 1996). Changes in larval size through the osmotic ux of water also could contribute to the changes in the size of larvae of P. astreoides. To effect water balance,

Moreover, the broadly similar shape of the response curves between trials suggests that the causal basis is systematic rather than artifactual, with the significant differences between trials potentially caused by differences in the larvae depending on the time of release (Edmunds et al. 2001; Isomura & Nishihira 2001). The best estimate of the characteristics of larvae freshly collected from Porites astreoides comes from our studies of this species at the same site in June 1999 (Edmunds et al. 2001). This study showed that freshly collected larvae contain a mean of 5,2387452 algal symbionts per larvae (7SE) and vary in length B0.51.0 mm (although length was not recorded systematically in our earlier study). Each larva in this case had 5,238 algal symbionts, which compares well to the mean population densities recorded here (2,1157,548 algal symbionts per larva in Trials 1 and 2, and 2,4284,994 per larva overall, i.e., pooled by trial; Fig. 1). The range of larval lengths is also similar to the mean size recorded here (0.9270.01 mm [7SE]). In terms of discerning whether the larvae grew or shrank (in terms of size and/or symbiont content) in our experiments, the outcome of the comparison with the larvae from 1999 is equivocal. The larvae in some tubes contained more, while others fewer, algal symbionts, and overall (i.e., pooled by trial), the mean population density of algal symbionts was lower (i.e., o5,238). Thus, overall the comparison supports the hypothesis that the responses (Fig. 1) are a result of symbiont losses, or perhaps digestion (Titlyanov et al. 1998), but it is noteworthy that there were no conspicuous signs (i.e., brown deposits or turbidity) of algal symbionts in the incubation water at the conclusion of the experiment. Moreover, we found a few dividing symbionts in most of the larvae when they were analysed (Edmunds, unpubl. data). Thus, in the case of the present study, there is no direct evidence that algal symbionts were expelled from the larvae, and it is unknown whether (or to what extent) the freshly collected larvae in 1999 are representative of those obtained in 2002. Conversely, there is evidence that the algal symbionts were proliferating in the 2002 larvae, and such proliferation could account for a sizeable change in symbiont population density, independent of cell loss/degradation. For example, based on larvae of Stylophora pistillata, in which r8% of the algal symbionts are in division at any one point in time (Titlyanov et al. 1998), larvae of P. astreoides containing 3,762 algal symbionts (the median value in Fig. 1F) could boost their algal symbiont population to 4,386 in 24 h (i.e., a 17% increase) with two divisions per cell per day (Wilkerson

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Temperature and size of coral larvae osmolytes potentially could be provided by glycerol created through the mobilization of the lipid reserves of coral larvae (Stimson 1987; Gates & Edmunds 1999), or the temperature-mediated increases in intracellular calcium concentrations (Fang et al. 1997). However, these putative osmotic effects cannot fully account for the changes in larval size, because the significant and positive correlation between size and symbiont population density (r 5 0.348, df 5 262, Po.001) shows that cell proliferation contributes to the changes (i.e., the size of the algal symbiont populations are increasing to match, in part, the increase in larval size). Given the low irradiances in the incubation tubes, the autotrophic capacity of larvae of P. astreoides (Edmunds et al. 2001) would be unlikely to fuel the increases in size through anabolic processes (described above). Likewise, catabolism probably does not play a major role in larval shrinkage, because with a metabolic rate of 0.13 nmol O2 min1 larva1 (Edmunds et al. 2001) and an energy content of 270 mJ larva1 (as in Porites porites [Edmunds & Spencer Davies 1986]), the daily metabolic costs would utilize only B0.03% of the energy reserves. With such a discrepancy between energy reserves and metabolic rate (see also Spencer Davies 1991), it is unlikely that larval shrinkage could be driven by the stimulatory effect of temperature on respiration (i.e., a Q10 effect). In evaluating the experimental effects of temperature on coral larvae, care must be taken to consider the range of temperatures applied, exposure times, and the species studied, all of which strongly affect the outcome. Thus, extreme temperatures (e.g., 4301C) increase mortality, reduce motility, and promote development aberrations in larvae of Diploria strigosa (Bassim et al. 2002; Bassim & Sammarco 2003), yet in Porites astreoides they increase mortality without affecting motility, and also increase the rate of metamorphosis (Edmunds et al. 2001). These, as well as other effects of elevated temperature on scleractinian corals, will vary depending on the duration of exposure to the thermal stress (Fitt et al. 2001; Bassim et al. 2002; Bassim & Sammarco 2003). In the case of the early research of Jokiel & Guinther (1978) and Coles (1985), the positive effects of temperature on coral settlement (described above) reversed at higher temperatures, and indeed the shape of the settlement-temperature responses are evocative of the complex curvilinear responses reported here. Of particular note is the decrease in settlement at more extreme temperatures, followed by slightly increased settlement at still more extreme temperatures (Coles 1985: gure 5; Jokiel & Coles 1978: gure 1), which may be similar to the upturns in size and symbiont population density of larvae of P. astreoides ex-

191 posed to o25.81C or 29.01C (Fig. 1). It is possible that these responses are examples of hormesis known initially as the Arndt Schulz Lawwhich refers to over-stimulation of physiological activity (often associated with growth) upon exposure to a sub-lethal dose of a harmful agent (Stebbing 1982; Calabrese & Baldwin 2001), and a response previously documented for the photosynthesis of corals exposed to low doses of the water-soluble fraction of crude oil (Rinkevich & Loya 1983). The biological significance of these potential hormesis responses is unclear, but they warrant further investigation in light of their ramications for coral settlement under extreme conditions, and the suggestion that hormetic effects are evolutionary-based adaptive responses to environmental stimuli (Calabrase & Baldwin 2001). Finally, it is important to note that both traits investigated in the present study are functionally important for coral larvae. For example, size would affect the magnitude of the lipid reserves, and symbiont population density would affect the ability to feed autotrophically, both of which inuence the ability to meet the metabolic costs of planktonic dispersal, growth, metamorphosis, and post-settlement success (Richmond 1987; Isomura & Nishihira 2001). As a result, larval size and symbiont population density probably have tness consequences, as indicated by the inverse relationship between larval size and survivorship for 3 coral species in Okinawa (Isomura & Nishihira 2001). While the selective value of larval size and symbiont population density clearly would be context specific (for example, depending on the necessity of long-distance dispersal), these putative benets nevertheless demonstrate that the patterns recorded in the present study could have selective value with regards to larval success. Thus, the shape of the response curves and their position relative to ambient seawater temperature (Fig. 1) could reect an unknown benecial strategy for survival. Clearly, a pressing research goal is to characterize these potential selective values, and elucidate the causal basis of the changes in larval size and symbiont population density. To accomplish these tasks, multidisciplinary approaches spanning scales of investigation ranging from larval ecology to the molecular control of embryology will be required. Acknowledgments. This research was supported by the National Undersea Research Center (NURC) at the University of North Carolina at Wilmington, NOAA Grant #NA36RU-0132, and was completed under permits from the Florida Keys National Marine Sanctuary (FKNMS-2002-021), and the Florida Fish and Wildlife Conservation Commission (02R-691). We

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Edmunds, Gates, Leggat, Hoegh-Guldberg, & Allen-Requa

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thank S. Miller and the staff at NURC Key Largo for facilitating our Aquarius mission. This project would not have been possible without the commitment of our saturation team: J. A. Idjadi, S. Lee, J. Talacek, and M. S. Hulsbeck, and our surface support team: S. Saucedo and C. Terhorst. We thank R. C. Carpenter for the loan of equipment, and Professor L. Muscatine for the donation of the aluminum block that made this work possible. R. C. Carpenter generously provided comments that improved an earlier draft of this paper. This is contribution number 122 of the CSUN Marine Biology Program, and 1190 of the Hawaii Institute of Marine Biology.

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