Stephanie Krimou

Emma Gairin

Laura Gautrand

Jerome Sowinski

Mila Trotier

Lana Minier

Hugo Bischoff

Vincent Sturny

Tehani Maueau

Viliame Waqalevu

Fabio Bulleri

Xavier Raick

Frederic Bertucci

David Lecchini

HERBIVORY EFFECTS OF SEA URCHIN SPECIES ON A CORAL REEF (BORA-BORA, FRENCH POLYNESIA)

Keywords: Herbivorous, Macroalgae, Grazing, Tripneuste gratilla, Diadema setosum, Bora-Bora Conceptualization, Funding acquisition, Methodology, Supervision

Macroalgae are becoming dominant on coral reefs worldwide, replacing corals as key habitat formers. Incrementing consumption rates by herbivores, such as sea urchins, has been identified as a viable strategy for promoting coral recovery. By means of a 28-day experiment, we assessed the grazing effects of the sea urchins, Tripneustes gratilla and Diadema setosum, on the macroalgal assemblages of shallow reefs at Bora-Bora (French Polynesia). In particular, each urchin species was either absent or maintained, using cages ( n = 10), at a density of 5 individuals/m 2 . T. gratilla exerted stronger negative effects than D. setosum on all the examined macroalgae (i.e., Turbinaria ornata, Padina boryana, Halimeda spp, Dictyota spp). In addition, the grazing effects by T. gratilla on macroalgae emerged shortly after the start of the experiment, more rapidly than those of D. setosum. Despite an apparent lower food preference for T. ornata in D.setosum, all macroalgae were consumed by both species and the composition of the macroalgal assemblages (i.e., the proportion of each species) did not change after 28 days. Overall, our study suggests that herbivorous sea urchins, when at relatively high densities, could contribute to sustaining coral-dominance on coral reefs by keeping macroalgae under control.

Introduction

Coral reefs are among the most productive and biodiverse ecosystems on the planet, providing a range of ecosystem services on which humanity depends [START_REF] Spalding | World Atlas of Coral Reefs[END_REF]. Unfortunately, reef health is declining at a global scale, impinging on their ability to provide goods and services [START_REF] Eddy | Global decline in capacity of coral reefs to provide ecosystem services[END_REF]. A variety of anthropogenic perturbations, acting at range of spatial scales, from global to local, can promote a shift in dominance from corals to macroalgae. In the aftermath of major events of disturbance, such as heatwaves and hurricanes, causing coral die-offs, freed space is typically colonized by filamentous ephemeral algae, that are rapidly replaced by longer-lived taxa, such as those belonging to the genera Turbinaria, Sargassum, Lobophora, Dictyota and Padina (e.g., [START_REF] Brown | The dynamics of coral-algal interactions in space and time on the southern great barrier reef[END_REF][START_REF] Mumby | Fishing, trophic cascades, and the process of grazing on coral reefs[END_REF]. Chronic alteration of local environmental conditions, such as excessive nutrient loading, or the collapse of herbivores can then favour the persistence of macroalgae, reducing coral recovery. Once dominant, macroalgae can, in fact, directly or indirectly impact adult and juvenile corals, as well as the settlement of larvae, through shading, abrasion, transmission of diseases or microbes and through the release of allelochemicals (e.g., [START_REF] Hughes | Phase shifts, herbivory, and the resilience of coral reefs to climate change[END_REF][START_REF] Nugues | Algal contact as a trigger for coral disease[END_REF]Vieira et al., 2016;[START_REF] Bulleri | Erect macroalgae influence epilithic bacterial assemblages and reduce coral recruitment[END_REF]. Understanding the mechanisms that maintain the competitive balance in favour of hard corals is key to preventing phase shifts towards the algal-dominated state and to promote shifts back to the coral-dominated one.

Interactions between corals and macroalgae are often interpreted in terms of a competitive superiority of macroalgae under reduced herbivory or increased nutrient availability [START_REF] Mccook | Competition between corals and algae on coral reefs: a review of evidence and mechanisms[END_REF]. In nondegraded reefs, the combination of bottom-up (e.g., limited nutrient supply) and topdown controls (e.g., high consumer pressure) can limit macroalgal proliferation (Valentine and Edgar, 2010). In addition to competitive inhibition by corals [START_REF] Mccook | Competition between corals and algae on coral reefs: a review of evidence and mechanisms[END_REF], herbivory is one of the critical factors that control macroalgal abundance in terms of biomass per unit area (Viviani et al., 2019). For instance, browsing fishes are often critical for maintaining coral-dominated reefs through their consumption of macroalgae (e.g., [START_REF] Fong | Coral reef algae[END_REF][START_REF] Hughes | Phase shifts, herbivory, and the resilience of coral reefs to climate change[END_REF][START_REF] Mumby | Fishing, trophic cascades, and the process of grazing on coral reefs[END_REF]. In particular, parrotfish, surgeonfish, rabbitfish and batfish are dominant herbivores in coral ecosystems [START_REF] Bellwood | Sleeping functional group drives coralreef recovery[END_REF][START_REF] Tebbett | The functioning of future coral reefs: fishes, sediments and productivity[END_REF] and thus play a pivotal role in the resilience of these highly threatened habitats [START_REF] Bellwood | Sleeping functional group drives coralreef recovery[END_REF][START_REF] Bonaldo | The ecosystem roles of parrotfishes on tropical reefs[END_REF]Viviani et al., 2019). Sea urchins have been also shown to play an important role in controlling macroalgal biomass on coral reefs. For example, in the Caribbean, the recovery of Diadema antillarum has resulted in the reduction of macroalgae and in the recovery of coral cover [START_REF] Edmunds | Recovery of Diadema antillarum reduces macroalgal cover and increases abundance of juvenile corals on a Caribbean reef[END_REF]. Likewise, some promising trials were performed with Tripneustes gratilla in Hawai'i, demonstrating that biocontrol through manual removal combined with hatchery raised urchins can be an effective management approach in controlling invasive macroalgae, with their cover reduced by 85% [START_REF] Neilson | Herbivore biocontrol and manual removal successfully reduce invasive macroalgae on coral reefs[END_REF].

Nonetheless, our understanding of their role in some tropical regions, such as the South Pacific remains limited. A recent study by [START_REF] Bulleri | Compounded effects of sea urchin grazing and physical disturbance on macroalgal canopies in the lagoon of Moorea, French Polynesia[END_REF], performed in the lagoon of Moorea, has shown that diadematid urchins (i.e., Diadema savignyi, Echinothrix diadema, E. calamaris) had a limited ability to control macroalgal recovery when at relatively low densities (i.e., 1 individual/m 2 ). The present study aimed to experimentally assess the ability of another diadematid species, Diadema setosum (Leske 1778), to reduce the cover of erect macroalgae in the lagoon of Bora-Bora, when maintained at higher densities. The grazing effects of D. setosum was compared to those of Tripneustes gratilla (L. 1767), a species previously suggested to be an effective consumer of brown seaweeds in the Indian Ocean [START_REF] De | Food selectivity of Tripneustes gratilla (L.) (Echinodermata: Echinoidea) in oligotrophic and nutrient-enriched coral reefs at La Reunion (Indian Ocean)[END_REF].

Material and methods

AREA OF INVESTIGATION AND SAMPLE COLLECTION

Bora-Bora (16°30' S, 151°45' W), often called the 'Pearl of the Pacific', is a high volcanic island in French Polynesia [START_REF] Gabrie | Study of the coral reefs of Bora-Bora (society archipelago, French Polynesia) for the development of a conservation and management plan[END_REF]. Coral reefs surrounding Bora-Bora have an area of approximately 70 km 2 , with four main geomorphological units (from the coast to ocean): fringing reef, channel, barrier reef, and outer slope [START_REF] Lecchini | Assessment of ecological status of the lagoon of Bora-Bora Island (French Polynesia)[END_REF]. The present study was performed from April to July 2022 on a fringing reef at the Marine Education Area (MEA) of Bora-Bora High School (Fig. 1). The site is dominated by dead coral reefs mainly colonized by four macroalgal species (i.e., Turbinaria ornata, Padina boryana, Halimeda spp, Dictyota spp).

Tripneustes gratilla (Fig. 2a) is one of the four sea urchin species of the genus Tripneustes [START_REF] Kroh | World Echinoidea Database. Tripneustes gratilla[END_REF]. This species is globally distributed on coral reefs [START_REF] Kroh | World Echinoidea Database. Tripneustes gratilla[END_REF], it feeds mainly at night and is primarily an opportunistic omnivore displaying a range of diets in wild and aquaculture environments. While the diet of juveniles of T. gratilla is mainly composed by sessile diatoms, adult individuals feed on macroalgae (reviewed in [START_REF] Lawrence | Tripneustes[END_REF]. In particular, adult T. gratilla forage close to the substratum, ingesting leaves of Thalassia hemprichii, Halimeda sp., Syringodium isoetifolium and coral rubble. Diadema setosum (Fig. 2b) is a species of Diadematidae, a common urchin family in tropical seas. In some regions, such as the Caribbean, Taiwan and Easter Island they have been shown to play a key role in controlling macroalgal development and, hence, in sustaining high coral cover (e.g., [START_REF] Ruyter | Changes in abundance of coral-reef bottom components related to mass mortality of the sea urchin Diadema antillarum[END_REF][START_REF] Myhre | Recovery of sea urchin Diadema antillarum populations is correlated to increased coral and reduced macroalgal cover[END_REF]. Species belonging to this family have also been reported as important agents of coral bioerosion [START_REF] Carreiro-Silva | Echinoid bioerosion and herbivory on Kenyan coral reefs: the role of protection from fishing[END_REF][START_REF] Glynn | Bioerosion and coral-reef growth: A dynamic balance[END_REF].

The densities of both sea urchin were low at the study site, i.e., < 0.001 individual/m2 [START_REF] Lecchini | Assessment of ecological status of the lagoon of Bora-Bora Island (French Polynesia)[END_REF].

EXPERIMENTAL DESIGN AND SAMPLING PROTOCOL

Adult sea urchins (T. gratilla: n = 94, size = 7 ± 1 cm -mean ± SD; D. setosum: n = 60, test diameter = 6 ± 1 cm) were collected by snorkelling on the fringing reefs of Bora-Bora, temporarily stored in coolers containing seawater at ambient temperature and relocated within a few hours in mono-specific enclosures made of wire mesh (5 cm) and metal stakes (40 to 70 cm) on the fringing reef of the Marine Published in : Journal of Experimental Marine Biology and Ecology (2023), vol. 564, 151900 DOI: 10.1016/j.jembe.2023.151900 Status : Postprint (Author's version) Educative Area of Bora-Bora, where they acclimated for 24 h. They were then kept for another 24 h in similar types of enclosures, but built on sand and, hence, without macroalgae. Thirty experimental enclosures were deployed over dead coral patches of 1 m 2 to 4 m 2 that, at the start of the experiment, had an overall macroalgal cover above 80%. The cages were spaced at least three meters apart. Ten replicate enclosures were randomly assigned to each of the following urchin treatments: 1) presence of T. gratilla; presence of D. setosum; 3) no sea urchins (i.e., Control). Each urchin species was kept at a density of 5 individuals/m 2 . Previous studies performed in Jamaica showed that a high density (e.g., D. setosum: 5 ind/m 2 ) was necessary to generate positive effects on coral cover through macroalgal control [START_REF] Idjadi | Recovery of the sea urchin Diadema antillarum promotes scleractinian coral growth and survivorship on shallow Jamaican reefs[END_REF][START_REF] Sammarco | Diadema and its relationship to coral spat mortality: grazing, competition, and biological disturbance[END_REF].

The total percentage of algal cover inside enclosures (Fig. 3) and the percentage of each algal species (T. ornata, P. boryana, Halimeda spp., Dictyota spp.) were recorded before the start of the experiment (Day 0) and, thereafter, every four days over a period of 28 days, for a total of 9 sampling events. The same observer (S. Krimou) positioned herself in front of each enclosure and visually quantified the percentage of macroalgae, by choosing 10 random visual intercept points on each of the four sides and on the top of the coral patches present at different points (method adapted from [START_REF] Dethier | Visual versus random-point percent cover estimations: 'objective' is not always better[END_REF]. Moreover, the observer had trained before the beginning of the experiment by comparing her visual estimates with data obtained from photo-quadrats and Image J processing.

All urchins survived the experiment and were released close to collection areas after 28 days.

STATISTICAL ANALYSIS

Shapiro-Wilk tests were used to check for the normal distribution of the total percentage cover of macroalgae in T. gratilla, D. setosum and control enclosures (T. gratilla, W = 0.95, p-value = 0.006; D. setosum, W = 0.95, p-value = 0.003; Control, W = 0.95, p-value = 0.004). Parametric test assumptions were not met, so non-parametric tests were used. A Kruskal-Wallis test showed no differences in the total percentage of macroalgae among the three treatments at Day 0 (Kruskal-Wallis, χ 2 = 1.74, df = 2, p-value = 0.42). Thus, for each condition, the total percentage of macroalgae was compared between the first and last day (D0 vs. D28) using Wilcoxon -rank tests. The grazing rate was then calculated as the slope of the linear regression between macroalgal cover and time (i.e., from D0 to D28). A Kruskal-Wallis test, followed by a Dunn post hoc test, was then performed to compare the slopes of the linear regression across the three treatments. Finally, the percentage cover of each macroalgal species (T. ornata, P. boryana, Halimeda spp, Dictyota spp) at D0 versus D28 were compared for each of the three treatments (T. gratilla, D. setosum and Control) with Pearson's χ 2 tests. All the statistics were performed with R version 4.1.1 (2021).

Results

TEMPORAL DYNAMICS OF MACROALGAL GRAZING BY SEA URCHINS

In T. gratilla enclosures, macroalgal cover (Fig. 4) significantly decreased from 88.2 ± 3.3% (mean ± SD) at D0 to 26.1 ± 3.5% at D28 (Wilcoxon signed-rank test; V = 55, p = 0.006). There was also a significant decrease of macroalgal cover in D. setosum enclosures (V = 55, p = 0.006), which was, however, smaller (i.e., from 93.1 ± 2.5% at D0 to 68.9 ± 6.1% at D28). In control enclosures, the change in macroalgal cover, although significant (V = 34.5, p = 0.025) was moderate (i.e., from 91.3 ± 20% at D0 to 83.9 ± 3.2% at D28).

The grazing rate (quantified as the change of macroalgal cover through time; Fig. 4) differed significantly among treatments (Kruskal-Wallis; χ 2 = 22.2, df = 2, p < 0.0001). There was a significant difference in the rate of macroalgal grazing between T. gratilla and the control (Dunn's post-hoc test: Z = 4.64, p < 0.0001) and between T. gratilla and D. setosum (Z = 2.91, p = 0.0054). By contrast, the grazing rate did not differ between D. setosum and the control (Z = 1.90, p = 0.057).

MACROALGAL CONSUMPTION

Changes in the percentage cover of each of the macroalgae species between the beginning and end of the experiment (D0 vs. D28) showed that T. ornata was consumed more by T. gratilla (4 ± 2% consumed after 1 day to 82 ± 19% consumed after 28 days) than by D. setosum (2 ± 1% at D1 to 19 ± 14% at D28) (Fig. 5). In control enclosures, T. ornata cover decreased by 7 ± 1% after 28 days. The three other macroalgal species (P. boryana, Halimeda spp, and Dictyota spp) were also more consumed by T. gratilla than D. setosum, while their covers changed little in the absence of sea urchins (Fig. 5).

None of the three treatments showed a significant difference in macroalgal composition between D0 and D28 (Pearson's tests, T. gratilla: χ 2 = 5.76, df = 3, p = 0.12; D. setosum: χ 2 = 0.97, df = 3, p = 0.81; Control: χ 2 = 0.17, df = 3, p = 0.98) (Fig. 6).

Published in : Journal of Experimental Marine Biology and Ecology (2023), vol. 564, 151900 DOI: 10.1016/j.jembe.2023.151900 Status : Postprint (Author's version)

Figure. 4. Total percentage (mean ± SD) of macroalgal cover for all replicate enclosures with

Tripneustes gratilla, Diadema setosum and the no sea urchins control from D0 to D28. Linear regression equations = T. gratilla : y = -2,17x + 84,28; D. setosum : y = -0,71x + 89,46; Control: y = -0,25x + 91,39.

Figure. 5. Consumption percentage of each macroalgae species (mean ± SD) over time in the three conditions (T. gratilla, D. setosum, control enclosures).

Published in : Journal of Experimental Marine Biology and Ecology (2023), vol. 564, 151900 DOI: 10.1016/j.jembe.2023.151900 Status : Postprint (Author's version) Figure. 6. Average percentage stacked bar plot of each macroalgae species present in the three experimental enclosures at day 0 and after 28 days of grazing.

Discussion

Herbivory has long been recognized as a major determinant of plant community organization in both terrestrial [START_REF] Harper | The role of predation in vegetational diversity[END_REF] and marine environments [START_REF] Crisp | Grazing in Terrestrial and Marine Environments: A Symposium of the British Ecological Society[END_REF]. Echinoids are among the most important groups of benthic grazers in temperate and tropical marine ecosystems [START_REF] Lawrence | Effects of feeding on the environment: Echinoidea[END_REF]. Our results showed that the efficiency and rate of grazing of T. gratilla were significantly greater than those of D. setosum (Figs. 4,5), despite the specimen of both species that were used were adults. Indeed, in our enclosures, T. gratilla caused a decrease in total macroalgal cover from 88% to 26% in just 28 days. Our results are in contrast with those of [START_REF] Carpenter | Invertebrate predators and grazers[END_REF], who found that the effects of grazing of T. gratilla were weaker in comparison to those of different Diadema spp. On the other hand, other in situ experiments have brought evidence that T. gratilla could markedly reduce macroalgal biomass within cages (e.g., [START_REF] Chon | Optimizing biocontrol: Assessing benthic community changes with different sizes and densities of Tripneustes gratilla on patch reef dominated by invasive Kappaphycus spp[END_REF][START_REF] Conklin | Abundance and spread of the invasive red algae, Kappaphycus spp., in Kane'ohe bay, Hawai'I and an experimental assessment of management options[END_REF]Westbrook et al., 2015). At La Reunion Island, Lison de Loma et al. ( 2002) found that T. gratilla actively selects food items, exhibiting a marked preference for T. ornata. We found no significant preference of T. gratilla for a specific macroalgal species since the proportions of macroalgal species did not change between the start and the end of the study (see Fig. 6). T. gratilla may benefit from a mixed diet that would provide a more balanced nutrition and, potentially, a dilution of toxic secondary metabolites produced by some of the macroalgal species, such as Dyctiotales [START_REF] Bernays | Dietary mixing in a generalist herbivore: tests of two hypotheses[END_REF][START_REF] Hay | Marine chemical ecology: what's known and what's next?[END_REF].

In D. setosum enclosures, the magnitude of the decrease in total macroalgal cover, although significant, was smaller. Weaker effects of D. setosum in comparison with T. gratilla could be a result of an active avoidance of T. ornata, as previously documented [START_REF] Coppard | Grazing preferences of diadematid echinoids in Fiji[END_REF][START_REF] Dajka | Investigating sea urchin densities critical to macroalgal control on degraded coral reefs[END_REF]. T. ornata is, in fact, mechanically and chemically defended against herbivores [START_REF] Sakanishi | Characterization of thallus mechanical and physiological traits of tropical fucoids: a preliminary study[END_REF][START_REF] Stiger | Phenolic contents of two brown algae, Turbinaria ornata and Sargassum mangarevense on Tahiti (French Polynesia): interspecific, ontogenic and spatiotemporal variations[END_REF]. Although the composition of the macroalgal assemblage at the end of the study, in terms of the proportion of each species, was not significantly different from the beginning, D. setosum appeared to consume P. boryana, Halimeda spp, and Dictyota spp more than T. ornata. This could be explained by T. ornata providing refuge against grazing for palatable macroalgae, as suggested by previous studies [START_REF] Bittick | Turbinaria ornate as an herbivory refuge for associate algae[END_REF][START_REF] Bulleri | Compounded effects of sea urchin grazing and physical disturbance on macroalgal canopies in the lagoon of Moorea, French Polynesia[END_REF][START_REF] Davis | Associational refuge facilitates phase shifts to macroalgae in a coral reef ecosystem[END_REF].

Our results are in accordance with a recent study [START_REF] Bulleri | Compounded effects of sea urchin grazing and physical disturbance on macroalgal canopies in the lagoon of Moorea, French Polynesia[END_REF], conducted at Moorea (another island in French Polynesia), revealing no effect of diadematids (Diadema savignyi, Echinothrix diadema and E. calamaris) on the covers of canopies formed by T. ornata and Sargassum pacificum these urchins are present at low densities (i.e., 1 ind/m 2 ). Stronger effects of diadematids on macroalgae have emerged at higher densities (e.g., [START_REF] Dajka | Investigating sea urchin densities critical to macroalgal control on degraded coral reefs[END_REF][START_REF] Dang | Sea urchins play an increasingly important role for coral resilience across reefs in Taiwan[END_REF] than those examined in our study (5 ind/m 2 ). Thus, the effects of diadematids on erect macroalgae appear density-dependent and there could be thresholds below which their grazing effects are negligible. The slight reduction in macroalgal cover at a D. setosum density of 5 ind/m 2 does not align with the findings of previous studies [START_REF] Idjadi | Recovery of the sea urchin Diadema antillarum promotes scleractinian coral growth and survivorship on shallow Jamaican reefs[END_REF]Sam-marco, 1980), indicating that 3-4 individuals/m 2 would be necessary to generate effects on macroalgal control. Our results rather support the postulation that the ability of diadematids to reduce macroalgal cover would be dependent upon the maintenance of higher densities of individuals and the composition of macroalgal assemblages [START_REF] Bulleri | Compounded effects of sea urchin grazing and physical disturbance on macroalgal canopies in the lagoon of Moorea, French Polynesia[END_REF][START_REF] Carpenter | Local and regional scale recovery of Diadema promotes recruitment of scleractinian corals[END_REF][START_REF] Dang | Sea urchins play an increasingly important role for coral resilience across reefs in Taiwan[END_REF]. In the present study area, it may therefore be appropriate to consider sea urchins cage culture in order to use urchins to assure an efficient grazing impact.

Conclusions

In summary, our study suggests that the strength of the effects of grazing by sea urchins are markedly species-specific. Increasing the abundance of T. gratilla appears functional for the reduction of the canopy stands formed by T. ornata, a species that, by virtue of its low palatability, is consumed by a reduced group of browsing fishes, namely Naso Iituratus and Calotomus carolinus [START_REF] Schmitt | Evaluating the precariousness of coral recovery when coral and macroalgae are alternative basins of attraction[END_REF]. Reducing the dominance of T. ornata may facilitate the grazing activity of other grazers, including diadematids, on less defended macroalgae. Stronger negative effects of diadematids on macroalgae can be expected at high densities, but these could come at a cost to corals, due to their bioerosive activity. Additional studies with a higher density of sea urchins could be pursued to see the effect of group and species behavior in the grazing of macroalgae.

CRediT authorship contribution statement

Stephanie Krimou: Investigation, Formal analysis, Writing -original draft, Writing -review & editing. Emma Gairin: Investigation, Formal analysis, Writing -original draft, Writing -review & editing. Laura Gautrand: Investigation. Jerome Sowinski: Investigation, Resources, Supervision. Mila Trotier: Investigation. Lana Minier: Investigation. Hugo Bischoff: Investigation. Vincent Sturny: Investigation,

Figure 1 .Figure 2 .

 12 Figure 1. Map of Bora Bora (A) and location of the study site in the Marine Educative Area (B).

Figure 3 .

 3 Figure 3. Location of macroalgae on a coral patch and pictures of the most abundant macroalgae species in the study area (MEA): Turbinaria ornata (A), Padina boryana (B), Dictyota spp. (C), Halimeda spp. (D). (Picture by S. Krimou).

Acknowledgements

We would like to thank the staff of Espace Bleu, Polynésienne des Eaux, Ia Vai Ma Noa Bora-Bora and the Commune de Bora-Bora for their help. This work has received several grants: Bloody Mary's, Fondation de France (2019-08602), Office Français de la Biodiversite (AFB/2019/ 385 -OFB.20.0888), Polynesienne des Eaux, ANR-19-CE34-0006-Man-ini and ANR-19-CE14-0010-SENSO, ANRT grant (CIFRE 2021/1268).

Declaration of Competing Interest

The authors declare the following financial interests/personal relationships which may be considered as potential competing interests:

Stephanie Krimou reports financial support was provided by Bloody Mary. David