Pr Rodolphe Cecilia Lemée

Pr Marie-Yasmine Totti

Dr Elisa Bottein

Berdalet

Dr Jean-Pierre Gattuso

Natalie Vigier

Jean Michel

Fabrice Guillaume

Gilbers

Pour M'avoir

Benoît

Alix Audrey

Miriam Julie Sylvain

Gilles Kilian

Keywords: forêts marines, Cystoseira compressa, communautés de macroalgues, réchauffement des océans, acidification des océans, changement climatique, changements de régime, efflorescences algales nuisibles, dinoflagellés benthiques, Ostreopsis marine forests, Cystoseira compressa, macroalgal communities, ocean warming, ocean acidification, climate change, herbivory, regime shifts, HABs, benthic dinoflagellates, Ostreopsiss

Time flies and I still don't believe that I am writing these lines as it means that this great adventure is coming to an end. These three years in Nice,

IV

I would like to thank you all for having transmitted to me your scientific spirit, your enthusiasm and your passion for research. I hope to have many opportunities to work again together in the future.

Being away from home and friends has not been easy, but I am very lucky to have family and friends who make it very easy for me. Vull donar les gràcies al meu grup d'amigues, a ses nines, tenc tanta sort de tenir-vos! Sempre heu estat al meu costat, just a una cridada de distància. M'he perdut moltes festes, verbenes post-covid, despedides de soltera i baby showers, però per molt tard que fos o moltes copes que hi hagués, sempre heu trobat un moment per fer-me una videotrucada.

He de dir que no me vaig perdre el primer concert d'Antonia Font, i això va ser també gracies vosaltres i a les vostres super-estratègies 😊 El àudios de WhatsApp han sigut més terapèutics que mai. Gràcies amigues.

Merci aussi à Sara et Maurine, mes colloques, et à Marie Laure, avec qui j'ai survie une pandémie mondiale et plusieurs confinements. On a eu de temps suffisant pour vous m'apprendre le français, de célébrer les Pâques, de faire de CrossFit et de beaucoup dansé et mangé. Vous avez été comme le soleil ! Quelle chance j'ai eu de vous trouver comme colocataires, vous me manquez tellement ! Vous êtes de personnes incroyables de qui j'ai appris tant de choses différentes <3 Gràcies també a tu, Alex. I do not know if one day I will be able to thank you for everything you have done and do every day for me. You have been my biggest support during these three years, encouraging me to do better but also to keep my feet on the ground. I am not sure who is happier about submitting this thesis :P New adventures begin for both of us and I hope to live them by your side.

I am fortunate to have a family that unconditionally supported me emotionally, and financially, throughout all the way to get here. Gràcies per tot el que feis per jo cada dia. Gràcies als meus pares y al meu germà Miquel, per escoltar i intentar entendre en que consisteixen els meus experiments, per començar a comprendre que és un paper i per és tan difícil publicar-ne un, per aplaudir cada petitíssima passa que faig. Gràcies a ma mare, per les mil vegades que puc arribar a telefonar-la en un dia, i per anar a agafar mostres d'aigua a s'Arenal per jo quan deien que podrien ser d'Ostreopsis (tot i que després van resultar ser E. coli...). Gràcies a mon pare, per guardar-me tots els retalls de diari que parlen de ciència, d'ecologia, d'entrevistes a científics o novetats de l'IMEDEA o l'IEO de Palma, i per les nostres infinites conversacions amb musica de fons. Gràcies també a les meves padrines i padrins i a sa madrina, per estimar-me i recolzar-me sempre incondicionalment, i per entendre que estar enfora ha estat molt important per jo. Vos he enyorat molt, estar enfora de vosaltres ha estat la part més difícil. Gràcies per tot, em sento molt afortunada.

VII

RESUME

Les grandes algues brunes sont des espèces clés dans les écosystèmes marins des latitudes tempérées où elles forment les forêts marines, parmi les écosystèmes les plus productifs et diversifiés au monde. En Méditerranée, elles sont représentées par les espèces du genre Cystoseira sensu lato, pour la plupart endémiques et caractérisées par leur longue durée de vie et faible dispersion. Cependant, les impacts anthropogéniques causent leur régression, entrainant des changements abruptes vers des communautés moins complexes (communautés gazonnantes et déserts marins), rendant l'écosystème potentiellement plus vulnérable aux phénomènes émergents, tels que les efflorescences de microalgues benthiques nuisibles. Les efflorescences de dinoflagellés benthiques du genre Ostreopsis, se développant sur les communautés macroalgales ont augmenté au cours des dernières décennies dans les régions tempérées, y compris la Méditerranée. Elles sont connues pour leurs effets néfastes sur la santé publique, les écosystèmes et l'économie qui en dépend.

Les objectifs de cette thèse sont d'évaluer (i) les causes abiotiques (température et acidification) et biotiques (herbivorie, facilitation écologique) potentiellement à l'origine de la régression des forêts marines Méditerranéennes, et (ii)

GENERAL INTRODUCTION AND OBJECTIVES

GENERAL INTRODUCTION AND OBJECTIVES …………………………………………..………………………………

THE MARINE ENVIRONMENT

A threatened biodiversity

Oceans constitute the largest volume of life on the planet covering 70.8 % of the Earth's surface [START_REF] Boeuf | Marine biodiversity characteristics[END_REF]. They host an important number of species but also play an important role in climate regulation and provide major ecosystem services for humans [START_REF] Mcleod | A blueprint for blue carbon: toward an improved understanding of the role of vegetated coastal habitats in sequestering CO2[END_REF]. However, the increasing number of human populations ( reaching 8 billion in November 2022; United Nations Department of Economic and Social Affairs, Population Division, 2022) is changing global biodiversity at unprecedented rates [START_REF] Zalasiewicz | Stratigraphy of the Anthropocene[END_REF]. A large and rising proportion of the population lives close to the coast (three times more people than the global average is living less than 100 km from the sea; [START_REF] Small | A Global Analysis of Human Settlement in Coastal Zones[END_REF][START_REF] Todd | Towards an urban marine ecology: characterizing the drivers, patterns and processes of marine ecosystems in coastal cities[END_REF], negatively impacting the coastal environment, which constitutes an important zone with a lot of spatial heterogeneity influencing the temperature, salinity, and primary production of coastal ecosystems.

Different types of anthropogenic stressors can impact marine ecosystems, some at a global scale, such as global warming and ocean acidification; and others at a regional or local scale, such as urbanisation, sedimentation, overfishing, invasive species and water pollution. As a consequence of these impacts and their interactions, reductions in habitat structure, biodiversity and trophic complexity are expected [START_REF] Doney | Climate Change Impacts on Marine Ecosystems[END_REF][START_REF] Duarte | Global change and the future ocean: a grand challenge for marine sciences[END_REF][START_REF] Hall-Spencer | Ocean acidification impacts on coastal ecosystem services due to habitat degradation[END_REF].

Among the anthropogenic pressures, climate change is expected to be the strongest force of biodiversity change at the global scale [START_REF] Bellard | Impacts of climate change on the future of biodiversity[END_REF]. In the present, climate change is already producing changes in demographic rates and forcing the redistribution, adaptation and acclimation of species [START_REF] Mooney | Biodiversity, climate change, and ecosystem services[END_REF][START_REF] Doney | Climate Change Impacts on Marine Ecosystems[END_REF][START_REF] Doney | The Impacts of Ocean Acidification on Marine Ecosystems and Reliant Human Communities[END_REF][START_REF] Bernhardt | Resilience to Climate Change in Coastal Marine Ecosystems[END_REF].

All these impacts will probably influence the communities composition and interactions among species [START_REF] Bakker | Herbivory on freshwater and marine macrophytes: A review and perspective[END_REF][START_REF] Pagès | Contrasting effects of ocean warming on different components of plant-herbivore interactions[END_REF], producing cascade effects at the ecosystem level, with ecological and socioeconomic consequences [START_REF] Hoegh-Guldberg | The Impact of Climate Change on the World's Marine Ecosystems[END_REF][START_REF] Pecl | Biodiversity redistribution under climate change: Impacts on ecosystems and human wellbeing[END_REF][START_REF] Doney | The Impacts of Ocean Acidification on Marine Ecosystems and Reliant Human Communities[END_REF]. At present there is not a single marine ecosystem unthreatened by anthropogenic stressors (Halpern et al., 2008b, 20118), but the hard-bottom coastal areas and rocky reefs are the highest impacted [START_REF] Halpern | Evaluating and Ranking the Vulnerability of Global Marine Ecosystems to Anthropogenic Threats[END_REF][START_REF] Halpern | Recent pace of change in human impact on the world's ocean[END_REF]

The key role of macroalgae dominated communities

Macroalgae or seaweeds are key organisms in temperate coastal ecosystems functioning around the globe, largely contributing to benthic primary production (Steneck et al., 2002;[START_REF] Cheminée | Nursery value of Cystoseira forests for Mediterranean rocky reef fishes[END_REF][START_REF] Teagle | The role of kelp species as biogenic habitat formers in coastal marine ecosystems[END_REF][START_REF] Piazzi | Biodiversity in canopy-forming algae: Structure and spatial variability of the Mediterranean Cystoseira assemblages[END_REF]. Macroalgae also participate in carbon sequestration by capturing, storing, and potentially sequestering CO2 in the ocean through transportation to deep marine sediments (Krause-Jensen and [START_REF] Krause-Jensen | Substantial role of macroalgae in marine carbon sequestration[END_REF][START_REF] Krause-Jensen | Sequestration of macroalgal carbon: the elephant in the Blue Carbon room[END_REF][START_REF] Filbee-Dexter | Substantial blue carbon in overlooked Australian kelp forests[END_REF][START_REF] Filbee-Dexter | Seaweed forests are carbon sinks that can mitigate CO2 emissions[END_REF][START_REF] Wright | Climate-driven shifts in kelp forest composition reduce carbon sequestration potential[END_REF]. Furthermore, they provide other invaluable ecosystem services such as supporting fisheries and mariculture, nutrient cycling and protecting the shoreline (Bennett et al., 2015a;[START_REF] Blamey | The economic value of South African kelp forests and temperate reefs: Past, present and future[END_REF]Eger et al., 2021a).

Consequently, changes in the macroalgal composition may sway coastal ecosystems [START_REF] Sala | Fishing, Trophic Cascades, and the Structure of Algal Assemblages: Evaluation of an Old but Untested Paradigm[END_REF][START_REF] Shears | Toxic cascades: Multiple anthropogenic stressors have complex and unanticipated interactive effects on temperate reefs[END_REF][START_REF] Mineur | European seaweeds under pressure: Consequences for communities and ecosystem functioning[END_REF][START_REF] Smale | Climate-driven substitution of foundation species causes breakdown of a facilitation cascade with potential implications for higher trophic levels[END_REF]. The diverse macroalgal species thriving on rocky reefs can be characterised by their sizes, shapes, structure and architecture of their thallus. Based on these factors, we can differentiate turf, erect and forest-forming or canopyforming macroalgae. Turf-forming macroalgae are represented by species with tightly packed fronds and filamentous thin axes, usually forming dense and compact mats [START_REF] Stewart | Fluctuations in the quantity of sediments trapped among algal thalli on intertidal rock platforms in southern California[END_REF]Sala et al., 2012;[START_REF] Connell | What are algal turfs? Towards a better description of turfs[END_REF][START_REF] Thiriet | Abundance and Diversity of Crypto-and Necto-Benthic Coastal Fish Are Higher in Marine Forests than in Structurally Less Complex Macroalgal Assemblages[END_REF][START_REF] Mauffrey | Seaweed functional diversity revisited: Confronting traditional groups with quantitative traits[END_REF]. Erect macroalgae are described as foliose laminar, ribbon-like, massive or fan-like thallus and erect arborescent tufts, which do not form a canopy but a shrub [START_REF] Sala | The Structure of Mediterranean Rocky Reef Ecosystems across Environmental and Human Gradients, and Conservation Implications[END_REF][START_REF] Thiriet | Abundance and Diversity of Crypto-and Necto-Benthic Coastal Fish Are Higher in Marine Forests than in Structurally Less Complex Macroalgal Assemblages[END_REF][START_REF] Bertolini | Can secondary species maintain a primary role? Consistent inter-regional effects of understory algae on diversity[END_REF].

Forest-forming macroalgae include the most structurally complex macroalgae with cylindrical axes, branched and tree-like ramifications [START_REF] Sala | The Structure of Mediterranean Rocky Reef Ecosystems across Environmental and Human Gradients, and Conservation Implications[END_REF][START_REF] Thiriet | How 3D Complexity of Macrophyte-Formed Habitats Affect the Processes Structuring Fish Assemblages Within Coastal Temperate Seascapes?[END_REF][START_REF] Bertolini | Can secondary species maintain a primary role? Consistent inter-regional effects of understory algae on diversity[END_REF][START_REF] Shelamoff | Ecosystem engineering by a canopy-forming kelp facilitates the recruitment of native oysters[END_REF][START_REF] Assis | A finetuned global distribution dataset of marine forests[END_REF]. The macroalgae from this group are considered habitatforming species able to create three-dimensional habitats with a canopy and understory, which supports distinct communities of fish, invertebrates and other plants [START_REF] Cheminée | Nursery value of Cystoseira forests for Mediterranean rocky reef fishes[END_REF][START_REF] Cheminée | Juvenile fish assemblages in temperate rocky reefs are shaped by the presence of macroalgae canopy and its three-dimensional structure[END_REF][START_REF] Thiriet | Abundance and Diversity of Crypto-and Necto-Benthic Coastal Fish Are Higher in Marine Forests than in Structurally Less Complex Macroalgal Assemblages[END_REF][START_REF] Shelamoff | Ecosystem engineering by a canopy-forming kelp facilitates the recruitment of native oysters[END_REF]Wernberg and Filbee-Dexter, 2019). Analogically to terrestrial environments, the physical traits of the dominant species are used to classify macroalgal communities into turfs, shrubs and forests, indicating the functional similarities between habitats on land and in the sea (Figure 1; Wernberg and Filbee-Dexter, 2019).

MARINE FORESTS

Macroalgal marine forests are constituted by large brown forest-forming macroalgae of the orders Laminariales, Tylopteridales, Desmarestiales and Fucales (Wernberg and Filbee-Dexter, 2019) and are considered among the most productive and biodiversity-rich ecosystems on Earth (Wernberg and Filbee-Dexter, 2019;[START_REF] Pessarrodona | Global seaweed productivity[END_REF]. They form the tallest biotic component of the seascape, altering the physical and biological environment in the understorey [START_REF] Thiriet | How 3D Complexity of Macrophyte-Formed Habitats Affect the Processes Structuring Fish Assemblages Within Coastal Temperate Seascapes?[END_REF][START_REF] Veiga | Structural complexity of macroalgae influences epifaunal assemblages associated with native and invasive species[END_REF]. These foundation species dominate intertidal and subtidal rocky shores in temperate and polar regions worldwide (Figure 2 ;[START_REF] Feldmann | Les Laminariacées de la Méditerranée et leur répartition géographique[END_REF][START_REF] Boudouresque | Where Seaweed Forests Meet Animal Forests: the Examples of Macroalgae in Coral Reefs and the Mediterranean Coralligenous Ecosystem[END_REF]Wernberg and Filbee-Dexter, 2019). They provide functions very similar to terrestrial forests such as: (i) habitat, food and shelter to a multitude of species at different life stages, (ii) supporting a high biodiversity of primary and secondary producers and decomposers, and (iii) playing an important role in the functioning and structure of the ecosystem maintaining food-webs and enhancing the secondary production [START_REF] Ballesteros | Deep-water stands of Cystoseira zosteroides C. Agardh (Fucales, Ochrophyta) in the Northwestern Mediterranean: Insights into assemblage structure and population dynamics[END_REF][START_REF] Smale | Threats and knowledge gaps for ecosystem services provided by kelp forests: a northeast Atlantic perspective[END_REF][START_REF] Teagle | The role of kelp species as biogenic habitat formers in coastal marine ecosystems[END_REF]Wernberg et al., 2019a;[START_REF] Fragkopoulou | Global biodiversity patterns of marine forests of brown macroalgae[END_REF]. The structure and productivity of marine forests are influenced by many environmental factors that drive the growth, survival, reproduction and metabolism of forest-forming macroalgae, which in turn affect the whole habitat or ecosystem [START_REF] Irving | Light, sediment, temperature, and the early life-history of the habitat-forming alga Cystoseira barbata[END_REF][START_REF] Cardona | Pulses of phytoplanktonic productivity may enhance sea urchin abundance and induce state shifts in Mediterranean rocky reefs[END_REF][START_REF] Pessarrodona | Global seaweed productivity[END_REF][START_REF] Smale | Climate-driven substitution of foundation species causes breakdown of a facilitation cascade with potential implications for higher trophic levels[END_REF][START_REF] Smith | Examining the influence of regional-scale variability in temperature and light availability on the depth distribution of subtidal kelp forests[END_REF]. A variety of anthropogenic impacts are altering the environmental parameters that drive the functioning of ecosystems [START_REF] Krumhansl | Global patterns of kelp forest change over the past half-century[END_REF][START_REF] Pessarrodona | Can ecosystem functioning be maintained despite climate-driven shifts in species composition? Insights from novel marine forests[END_REF], and as a result negatively affect marine forests that at the present are in regression worldwide [START_REF] Filbee-Dexter | Rise of Turfs: A New Battlefront for Globally Declining Kelp Forests[END_REF][START_REF] Pessarrodona | Homogenization and miniaturization of habitat structure in temperate marine forests[END_REF]. Several local and global impacts are involved in the regression of marine forests such as urbanisation, marine farming, local pollution and herbivory (Steneck et al., 2002;[START_REF] Krumhansl | Global patterns of kelp forest change over the past half-century[END_REF][START_REF] Orfanidis | Effects of Natural and Anthropogenic Stressors on Fucalean Brown Seaweeds Across Different Spatial Scales in the Mediterranean Sea[END_REF], but also ocean warming (OW) and potentially ocean acidification (OA; [START_REF] Wernberg | Seaweed Communities in Retreat from Ocean Warming[END_REF][START_REF] Connell | The other ocean acidification problem: CO2 as a resource among competitors for ecosystem dominance[END_REF][START_REF] Friedlander | Kelp forests at the end of the earth: 45 years later[END_REF][START_REF] Smale | Impacts of ocean warming on kelp forest ecosystems[END_REF]. These stressors are putting marine forests to the limit, confining them to refuge locations characterized by less unfavourable conditions [START_REF] Straub | Resistance, Extinction, and Everything in Between -The Diverse Responses of Seaweeds to Marine Heatwaves[END_REF][START_REF] Verdura | Localscale climatic refugia offer sanctuary for a habitat-forming species during a marine heatwaves[END_REF]; and making them less resilient against future impacts [START_REF] Capdevila | Warming impacts on early life stages increase the vulnerability and delay the population recovery of a long-lived habitat-forming macroalga[END_REF][START_REF] Straub | Resistance, Extinction, and Everything in Between -The Diverse Responses of Seaweeds to Marine Heatwaves[END_REF].

Global change effects on marine forests can vary according to the location, the population characteristics and the species [START_REF] Krumhansl | Global patterns of kelp forest change over the past half-century[END_REF][START_REF] Hollarsmith | Varying reproductive success under ocean warming and acidification across giant kelp (Macrocystis pyrifera) populations[END_REF][START_REF] Verdura | Localscale climatic refugia offer sanctuary for a habitat-forming species during a marine heatwaves[END_REF]. There is evidence that early-life stages of these species are more vulnerable than adults and this, together with a very high natural mortality rate during early-life stages [START_REF] Ang | Natural dynamics of a Fucus distichus (Phaeophyceae, Fucales) population: reproduction and recruitment[END_REF][START_REF] Vadas | Recruitment and mortality of early postsettlement stages of benthic algae[END_REF][START_REF] Capdevila | Recruitment patterns in the Mediterranean deep-water alga Cystoseira zosteroides[END_REF], could lead, in the long term, to their loss [START_REF] Coelho | Impacts of anthropogenic stresses on the early development stages of seaweeds[END_REF][START_REF] Schiel | The Population Biology of Large Brown Seaweeds: Ecological Consequences of Multiphase Life Histories in Dynamic Coastal Environments[END_REF][START_REF] De Caralt | Differential effects of pollution on adult and recruits of a canopy-forming alga: implications for population viability under low pollutant levels[END_REF].

MEDITERRANEAN MARINE FORESTS

The Mediterranean Sea is the largest (more than 2 500 000 km 2 ) and deepest (average and maximum depth of 1 500 m and 5 267 m, respectively) enclosed sea on Earth [START_REF] Coll | The biodiversity of the Mediterranean Sea: estimates, patterns, and threats[END_REF]. Nowadays the Mediterranean coasts support a high density of inhabitants and represent one of the first touristic destinations in the world, with the consequent environmental cost that it entails [START_REF] Segreto | Europe at the Seaside: The Economic History of Mass Tourism in the Mediterranean[END_REF][START_REF] Coll | The biodiversity of the Mediterranean Sea: estimates, patterns, and threats[END_REF]. The Mediterranean Sea, characterized by small tides, oligotrophic waters and high salinity and mean water temperature [START_REF] Ros | Diving in blue water. The benthos[END_REF], is one of the major hotspots of biodiversity, especially on coastal areas [START_REF] Coll | The biodiversity of the Mediterranean Sea: estimates, patterns, and threats[END_REF]. Some of these unique communities are the meadows of the endemic phanerogam Posidonia oceanica, the coralligenous community built up by crustose coralline algae, the vermetid platforms, the Litophyllum brissoides algal reef and the Fucales forests [START_REF] Ros | Diving in blue water. The benthos[END_REF][START_REF] Boudouresque | Marine biodiversity in the Mediterranean: status of species, populations and communities[END_REF].

Macroalgal marine forests in the Mediterranean Sea are represented mainly by species belonging to the orders Fucales; in particular to the genera Cystoseira, Gongolaria and Ericaria, from now referred as Cystoseira sensu lato species (Box 1; Wernberg and Filbee-Dexter, 2019;[START_REF] Molinari -Novoa | Reinstatement of the genera Gongolaria Boehmer and Ericaria Stackhouse (Sargassaceae, Phaeophyceae)[END_REF]. There are more than thirty different species, most of them endemic to the Mediterranean Sea (Molinari -Novoa and Guiry, 2020), they are the dominant forest-forming species in intertidal and sublittoral rocky bottoms and despite showing smaller average size have functional traits comparable to those of larger kelps (Wernberg and Filbee-Dexter, 2019). Only two species of Laminariales are found in the Mediterranean Sea forming marine forests: Laminaria ochroeluca, in the immediacies of the Strait of Gibraltar, western coast of Africa and the Strait of Messina [START_REF] Drew | An ecological study of Laminaria ochroleuca Pyl. growing below 50 metres in the Straits of Messina[END_REF][START_REF] Flores-Moya | Warm Temperate Seaweed Communities: A Case Study of Deep Water Kelp Forests from the Alboran Sea (SW Mediterranean Sea) and the Strait of Gibraltar[END_REF]; and the critically endangered and endemic deep-water Laminaria rodriguezii [START_REF] Feldmann | Les Laminariacées de la Méditerranée et leur répartition géographique[END_REF][START_REF] Bo | Characteristics of the Mesophotic Megabenthic Assemblages of the Vercelli Seamount (North Tyrrhenian Sea)[END_REF].

BOX 1: Cystoseira sensu lato species

Cystoseira s.l. spp. have a high morphological plasticity that makes their identification complex using morphological traits only [START_REF] Rožić | Molecular identification of the brown algae, Cystoseira spp. (Phaeophycae, Fucales) from the Adriatic Seapreliminary results[END_REF][START_REF] Orellana | Diversity of Cystoseira sensu lato (Fucales, Phaeophyceae) in the eastern Atlantic and Mediterranean based on morphological and DNA evidence, including Carpodesmia gen. emend. and Treptacantha gen. emend[END_REF]. Analysis of DNA demonstrated the multiple phylogenic origin of the Mediterranean genus Cystoseira [START_REF] Draisma | DNA sequence data demonstrate the polyphyly of the genus Cystoseira and other Sargassaceae genera (Phaeophyceae)[END_REF] leading to its division in three clades [START_REF] Bruno De Sousa | Improved phylogeny of brown algae Cystoseira (Fucales) from the Atlantic-Mediterranean region based on mitochondrial sequences[END_REF]. A few years ago, [START_REF] Orellana | Diversity of Cystoseira sensu lato (Fucales, Phaeophyceae) in the eastern Atlantic and Mediterranean based on morphological and DNA evidence, including Carpodesmia gen. emend. and Treptacantha gen. emend[END_REF] [START_REF] Giaccone | Écologie et chorologie des Cystoseira de Méditerranée[END_REF]Ballesteros, 1990a[START_REF] Ballesteros | Structure and dynamics of the Cystoseira caespitosa Sauvageau (Fucales, Phaeophyceae) community in the North-Western Mediterranean. Estructura y dinámica de la comunidad de Cystoseira caespitosa Sauvageau (Fucales, Phaeophyceae) en el Mediterráneo noroccidental[END_REF][START_REF] Sala | The Structure of Mediterranean Rocky Reef Ecosystems across Environmental and Human Gradients, and Conservation Implications[END_REF]. Cystoseira s.l. spp. are considered habitat-forming as they generate a three-dimensional structure from which it is possible to differentiate different strata: an arboreal layer, on the upper part of the canopy made by the forest-forming species (i.e. Cystoseira s.l.) and the epiphytes; a middle shrubby layer on the middle canopy, formed by erect shrub-forming macroalgae; a turf layer, formed by turf-forming calcareous and sciaphilic macroalgae; and a basal layer, mainly formed by crustose calcifying algae [START_REF] Ros | Diving in blue water. The benthos[END_REF]. Cystoseira s.l. forests are one of the most productive habitats in the Mediterranean Sea (Figure 3;Ballesteros, 1989a;[START_REF] Sales | Seasonal dynamics and annual production of Cystoseira crinita (Fucales: Ochrophyta)-dominated assemblages from the northwestern Mediterranean[END_REF][START_REF] Duarte | Global estimates of the extent and production of macroalgal forests[END_REF][START_REF] Pessarrodona | Global seaweed productivity[END_REF] and provide habitat to a large number of associated species, supporting trophic networks [START_REF] Ballesteros | Els vegetals i la zonació litoral: espècies, comunitats i factors que influeixen en la seva distribució[END_REF][START_REF] Ballesteros | Deep-water stands of Cystoseira zosteroides C. Agardh (Fucales, Ochrophyta) in the Northwestern Mediterranean: Insights into assemblage structure and population dynamics[END_REF][START_REF] Thiriet | Abundance and Diversity of Crypto-and Necto-Benthic Coastal Fish Are Higher in Marine Forests than in Structurally Less Complex Macroalgal Assemblages[END_REF][START_REF] Piazzi | Biodiversity in canopy-forming algae: Structure and spatial variability of the Mediterranean Cystoseira assemblages[END_REF].

In function of their distribution and vertical zonation (that is influenced by many factors such as depth, light intensity, hydrodynamics, resistance to breaking, temperature, nutrient availability and herbivory pressure ;[START_REF] Feldmann | Les Laminariacées de la Méditerranée et leur répartition géographique[END_REF][START_REF] Vergés | Role of fish herbivory in structuring the vertical distribution of canopy algae Cystoseira spp. in the Mediterranean Sea[END_REF]Sant andBallesteros, 2020a, 2021), the forests are dominated by different Cystoseira s.l. spp. Some species are widely distributed along the Mediterranean Sea and others restricted to very specific locations, being most of the species found on shallow sheltered or exposed locations [START_REF] Ribera | Check-list of Mediterranean Seaweeds[END_REF].

Threats and status of Cystoseira sensu lato forests

Many anthropogenic impacts are pushing Mediterranean marine ecosystems to the edge [START_REF] Coll | The biodiversity of the Mediterranean Sea: estimates, patterns, and threats[END_REF]. The overexploitation and habitat loss (e.g. urbanisation) have been affecting the ecosystems for centuries and at present, together with fishing impacts, pollution, eutrophication and climate change, represent the most important threats to almost all species, including Cystoseira s.l. forests (Mangialajo et al., 2008b;[START_REF] Coll | The biodiversity of the Mediterranean Sea: estimates, patterns, and threats[END_REF][START_REF] Giakoumi | Relationships between fish, sea urchins and macroalgae: The structure of shallow rocky sublittoral communities in the Cyclades, Eastern Mediterranean[END_REF][START_REF] Orfanidis | Effects of Natural and Anthropogenic Stressors on Fucalean Brown Seaweeds Across Different Spatial Scales in the Mediterranean Sea[END_REF]. The Mediterranean Sea is very sensitive to climate change [START_REF] Belkin | Rapid warming of Large Marine Ecosystems[END_REF][START_REF] Lejeusne | Climate change effects on a miniature ocean: the highly diverse, highly impacted Mediterranean Sea[END_REF][START_REF] Tuel | Why Is the Mediterranean a Climate Change Hot Spot?[END_REF] and the seawater temperature is already increasing [START_REF] Lionello | The relation between climate change in the Mediterranean region and global warming[END_REF] as well as the intensity and frequency of marine heatwaves [START_REF] Oliver | Longer and more frequent marine heatwaves over the past century[END_REF][START_REF] Oliver | Projected Marine Heatwaves in the 21st Century and the Potential for Ecological Impact[END_REF]. In addition, global biodiversity scenarios also predict that invasive species will be an increasing problem in the Mediterranean Sea, which seems to be more threatened than other ecosystems [START_REF] Sala | Global Biodiversity Scenarios for the Year 2100[END_REF]. Therefore the introduction and the establishment of alien species of herbivores, macroalgae and microalgae can represent an additional stressor for marine forest (Vergés et al., 2014a(Vergés et al., , 2014b;;[START_REF] Marampouti | Mediterranean alien harmful algal blooms: origins and impacts[END_REF][START_REF] Iveša | Increased ammonium levels occurring during benthic algal blooms are potentially toxic to fucalean algae[END_REF].

As many other foundation species, Cystoseira s.l. spp. are very sensitive to both local and large scale variations on environmental conditions [START_REF] Irving | Light, sediment, temperature, and the early life-history of the habitat-forming alga Cystoseira barbata[END_REF][START_REF] Cardona | Pulses of phytoplanktonic productivity may enhance sea urchin abundance and induce state shifts in Mediterranean rocky reefs[END_REF]Mancuso et al., 2019;[START_REF] De Caralt | Differential effects of pollution on adult and recruits of a canopy-forming alga: implications for population viability under low pollutant levels[END_REF] and, in some cases, the unfavourable conditions are restricting Cystoseira s.l. populations to refuge locations where requisites for their survival are ensured [START_REF] Verdura | Localscale climatic refugia offer sanctuary for a habitat-forming species during a marine heatwaves[END_REF]. Because of the high sensitivity to human disturbances (Mangialajo et al., 2008b;[START_REF] Sales | Shallow Cystoseira (Fucales: Ochrophyta) assemblages thriving in sheltered areas from Menorca (NW Mediterranean): Relationships with environmental factors and anthropogenic pressures[END_REF] these species are used in many protocols with the objective of characterising coastal habitats [START_REF] De La Fuente | Ecological status improvement over a decade along the Ligurian coast according to a macroalgae based index (CARLIT)[END_REF][START_REF] Bahbah | Cartography of littoral rocky-shore communities to assess the ecological status of water bodies through the application of CARLIT method in Algeria (South-Western Mediterranean Sea)[END_REF]. Since decades, declines of Cystoseira s.l. forests have been reported all along the Mediterranean Sea [START_REF] Cormaci | Changes of the benthic algal flora of the Tremiti Islands (southern Adriatic) Italy[END_REF][START_REF] Thibaut | Long-term decline of the populations of Fucales (Cystoseira spp. and Sargassum spp.) in the Albères coast (France, Northwestern Mediterranean)[END_REF][START_REF] Thibaut | Decline and local extinction of Fucales in French Riviera: the harbinger of future extinctions?[END_REF][START_REF] Bianchi | Thirty years after -dramatic change in the coastal marine habitats of Kos Island (Greece), 1981-2013[END_REF][START_REF] Buonomo | Predicted extinction of unique genetic diversity in marine forests of Cystoseira spp[END_REF][START_REF] Mariani | Past and present of Fucales from shallow and sheltered shores in Catalonia[END_REF]. Consequently, most of the Cystoseira s.l. spp. are included in the Annex II of the Barcelona Convention (UNEP/MAP, 2013) and considered threatened or endangered and in need GENERAL INTRODUCTION AND OBJECTIVES …………………………………………..……………………………… of protection measures [START_REF] Verlaque | Mediterranean seaweeds listed as threatened under the Barcelona Convention: A critical analysis[END_REF]. The only Cystoseira s.l. spp. not included in the annex is the shallow widely distributed Cystoseira compressa [START_REF] Verlaque | Mediterranean seaweeds listed as threatened under the Barcelona Convention: A critical analysis[END_REF].

Declines in marine forests are attributed mainly to anthropogenic factors, local stressors seem to have a strong direct and indirect impact [START_REF] Russell | Synergistic effects of climate change and local stressors: CO2 and nutrient-driven change in subtidal rocky habitats[END_REF][START_REF] Leal | Copper pollution exacerbates the effects of ocean acidification and warming on kelp microscopic early life stages[END_REF][START_REF] De Caralt | Differential effects of pollution on adult and recruits of a canopy-forming alga: implications for population viability under low pollutant levels[END_REF][START_REF] Gissi | A review of the combined effects of climate change and other local human stressors on the marine environment[END_REF], but global stressors such as OW and OA are also affecting these ecosystems [START_REF] Celis-Plá | Macroalgal responses to ocean acidification depend on nutrient and light levels[END_REF][START_REF] Fernández | Nitrogen sufficiency enhances thermal tolerance in habitat-forming kelp: implications for acclimation under thermal stress[END_REF][START_REF] Smale | Impacts of ocean warming on kelp forest ecosystems[END_REF][START_REF] Verdura | Localscale climatic refugia offer sanctuary for a habitat-forming species during a marine heatwaves[END_REF]. Locally, the proliferation of herbivores is believed to be one of the most important biological factors controlling the mortality of early-life stages of Cystoseira s.l. spp. and responsible for the degradation and maintenance of post-regime shifts communities, such as turfs or barren grounds [START_REF] Giakoumi | Relationships between fish, sea urchins and macroalgae: The structure of shallow rocky sublittoral communities in the Cyclades, Eastern Mediterranean[END_REF]Vergés et al., 2014a;[START_REF] Gianni | Threats to large brown algal forests in temperate seas: the overlooked role of native herbivorous fish[END_REF]. Alongside, eutrophication and pollution affect the biomass, growth and photosynthetic yield of Cystoseira s.l. spp. and have been responsible for the regression of the populations in many locations [START_REF] Thibaut | Long-term decline of the populations of Fucales (Cystoseira spp. and Sargassum spp.) in the Albères coast (France, Northwestern Mediterranean)[END_REF][START_REF] Pinedo | Long-term changes in sublittoral macroalgal assemblages related to water quality improvement[END_REF][START_REF] Blanfuné | The ups and downs of a canopy-forming seaweed over a span of more than one century[END_REF][START_REF] De Caralt | Differential effects of pollution on adult and recruits of a canopy-forming alga: implications for population viability under low pollutant levels[END_REF]. Globally, OW has already been signalled as a thread for the survival of both adults and recruits of Cystoseira s.l. spp., affecting their phenology, recruitment and growth (Celis-Plá et al., 2017a;[START_REF] Bevilacqua | Climatic anomalies may create a long-lasting ecological phase shift by altering the reproduction of a foundation species[END_REF][START_REF] Falace | Is the South-Mediterranean Canopy-Forming Ericaria giacconei (= Cystoseira hyblaea) a Loser From Ocean Warming? Front[END_REF][START_REF] Verdura | Localscale climatic refugia offer sanctuary for a habitat-forming species during a marine heatwaves[END_REF][START_REF] Bennett | Thermal Performance of Seaweeds and Seagrasses Across a Regional Climate Gradient[END_REF][START_REF] Orlando-Bonaca | Diversity of Molluscan Assemblage in Relation to Biotic and Abiotic Variables in Brown Algal Forests[END_REF]. Instead, the effects of OA are expected to favour the growth of Cystoseira s.l. spp. when other environmental conditions, such as temperature and nutrients, are optimal [START_REF] Celis-Plá | Macroalgal responses to ocean acidification depend on nutrient and light levels[END_REF], 2017a). However, the interactive effects of local and global stressors, need to be further investigated to better understand the causes of such declines [START_REF] Hepburn | Diversity of carbon use strategies in a kelp forest community: implications for a high CO2 ocean[END_REF][START_REF] Connell | The other ocean acidification problem: CO2 as a resource among competitors for ecosystem dominance[END_REF]Celis-Plá et al., 2017a;[START_REF] Leal | Ocean acidification and kelp development: Reduced pH has no negative effects on meiospore germination and gametophyte development of Macrocystis pyrifera and Undaria pinnatifida[END_REF].

The substitution of marine forests by structurally less complex macroalgal communities such as shrubs and turfs, and even barren grounds or deserts, has been observed and studied in the Mediterranean Sea [START_REF] Chemello | Regime shifts and alternative stable states in intertidal rocky habitats: State of the art and new trends of research[END_REF][START_REF] Benedetti-Cecchi | Species Interactions and Regime Shifts in Intertidal and Subtidal Rocky Reefs of the Mediterranean Sea[END_REF][START_REF] Álvarez-Losada | A regime shift in intertidal assemblages triggered by loss of algal canopies: A multidecadal survey[END_REF] and the miniaturisation of the habitats is a common event on temperate areas worldwide [START_REF] Filbee-Dexter | Rise of Turfs: A New Battlefront for Globally Declining Kelp Forests[END_REF][START_REF] Pessarrodona | Homogenization and miniaturization of habitat structure in temperate marine forests[END_REF]. The loss of these habitat forming species involves a series of cascade effects with profound changes on the ecosystem function together with the loss of ecosystem services [START_REF] Airoldi | The Gray Zone: Relationships between habitat loss and marine diversity and their applications in conservation[END_REF][START_REF] Hall-Spencer | Ocean acidification impacts on coastal ecosystem services due to habitat degradation[END_REF][START_REF] Pessarrodona | Can ecosystem functioning be maintained despite climate-driven shifts in species composition? Insights from novel marine forests[END_REF][START_REF] Smale | Climate-driven substitution of foundation species causes breakdown of a facilitation cascade with potential implications for higher trophic levels[END_REF].

The characteristics of Cystoseria s.l. spp., long-lived species with a short dispersal capacity (because of the large size of their zygotes, 100 -120 µm), make very difficult the natural recovery of the forests, especially considering that populations are in many cases fragmented [START_REF] Clayton | The adaptive significance of life history characters in selectedorders of marine brown maeroalgae[END_REF][START_REF] Capdevila | Effective dispersal and density-dependence in mesophotic macroalgal forests: Insights from the Mediterranean species Cystoseira zosteroides[END_REF][START_REF] Riquet | Highly restricted dispersal in habitat-forming seaweed may impede natural recovery of disturbed populations[END_REF]. Even if the improvement of environmental conditions has been attained in some locations (e.g. improvement in water quality) allowing the regeneration of some populations (Thibaut et al., 2005, 200;[START_REF] Perkol-Finkel | Loss and recovery potential of marine habitats: an experimental study of factors maintaining resilience in subtidal algal forests at the Adriatic sea[END_REF][START_REF] Blanfuné | The ups and downs of a canopy-forming seaweed over a span of more than one century[END_REF], natural recovery is rare [START_REF] Iveša | Long-term fluctuations in Cystoseira populations along the west Istrian Coast (Croatia) related to eutrophication patterns in the northern Adriatic Sea[END_REF]. In most cases, the only solution for restoring locally extinct or highly endangered populations is the active restoration [START_REF] Gianni | Conservation and restoration of marine forests in the Mediterranean Sea and the potential role of Marine Protected Areas[END_REF]Eger et al., 2022). Recently, great efforts have been done to explore innovative restoration techniques for recovering or improving the status of endangered populations [START_REF] Falace | Ex situ cultivation protocol for Cystoseira amentacea var. stricta (Fucales, Phaeophyceae) from a restoration perspective[END_REF][START_REF] De La Fuente | First ex situ outplanting of the habitat-forming seaweed Cystoseira amentacea var. stricta from a restoration perspective[END_REF][START_REF] Cebrian | A Roadmap for the Restoration of Mediterranean Macroalgal Forests[END_REF]. So far, successful restoration actions of Cystoseira s.l. forests in the Mediterranean Sea are scarce [START_REF] Falace | Ex situ cultivation protocol for Cystoseira amentacea var. stricta (Fucales, Phaeophyceae) from a restoration perspective[END_REF][START_REF] Verdura | Restoration of a Canopy-Forming Alga Based on Recruitment Enhancement: Methods and Long-Term Success Assessment[END_REF][START_REF] De La Fuente | First ex situ outplanting of the habitat-forming seaweed Cystoseira amentacea var. stricta from a restoration perspective[END_REF][START_REF] Savonitto | Addressing reproductive stochasticity and grazing impacts in the restoration of a canopyforming brown alga by implementing mitigation solutions[END_REF]. The next main issues that need to be addressed and further studied for the accomplishment of restoration actions are herbivory and climate change [START_REF] Gianni | Conservation and restoration of marine forests in the Mediterranean Sea and the potential role of Marine Protected Areas[END_REF][START_REF] Tamburello | Enhanced nutrient loading and herbivory do not depress the resilience of subtidal canopy forests in Mediterranean oligotrophic waters[END_REF][START_REF] Medrano | From marine deserts to algal beds: Treptacantha elegans revegetation to reverse stable degraded ecosystems inside and outside a No-Take marine reserve[END_REF].

BENTHIC HARMFUL ALGAL BLOOMS PROLIFERATION

The same factors that are driving Cystoseira s.l. forests to the edge (e.g. degradation of natural habitats due to pollution, urbanisation, overfishing and climate change) and are promoting regime shifts could also make ecosystems more vulnerable to new biological threats. Recently, an increase in harmful algal blooms (HABs) has been recorded in temperate locations around the globe including the Mediterranean Sea [START_REF] Anderson | Scaling up from regional case studies to a global harmful algal bloom observing system[END_REF]. Anthropogenic activities and climate change are considered the main contributors of alien invasions and main enablers of HAB events [START_REF] Marampouti | Mediterranean alien harmful algal blooms: origins and impacts[END_REF]. In the global change context, ocean warming is expected to promote the expansion and growth of tropical and sub-tropical harmful algal dinoflagellates, including the genera Ostreopsis, Gambierdiscus and Fukoya [START_REF] Tester | Climate change and harmful benthic microalgae[END_REF]. A significant proportion of studies concerning benthic HABs are focused on tropical benthic dinoflagellate ecology, primarily as a result of the incidence of ciguatera fish poisoning (CFP) in tropical and subtropical areas [START_REF] Litaker | Taxonomy of Gambierdiscus including four new species, Gambierdiscus caribaeus, Gambierdiscus carolinianus, Gambierdiscus carpenteri and Gambierdiscus ruetzleri (Gonyaulacales, Dinophyceae)[END_REF] and the expansion of toxic species in temperate waters [START_REF] Rhodes | World-wide occurrence of the toxic dinoflagellate genus Ostreopsis Schmidt[END_REF]. To the present, no CFP episodes have been reported in the Mediterranean Sea, but the recent findings of species of Gambierdiscus in Crete (Aligizaki and Nikolaidis, 2008) and in Balearic Islands [START_REF] Laza-Martínez | Characterization of a Strain of Fukuyoa paulensis (Dinophyceae) from the Western Mediterranean Sea[END_REF][START_REF] Tudó | First report of Gambierdiscus in the Western Mediterranean Sea (Balearic Islands)[END_REF]Tudó et al., , 2020)), are rising the alert. Still, Ostreopsis spp. (Box 2) are already widely distributed in the Mediterranean Sea, where their blooms are frequent and important in magnitude (Mangialajo et al., 2011;Accoroni et al., 2012;Accoroni and Totti, 2016;Açaf et al., 2020). The increasing rate of Ostreopsis spp. blooms and the hazardous impacts that they have on the environment, economy, and human health make it important to have accurate knowledge about its development [START_REF] Berdalet | Marine harmful algal blooms, human health and wellbeing: challenges and opportunities in the 21st century[END_REF][START_REF] Berdalet | Environmental, human health and socioeconomic impacts of Ostreopsis spp. Blooms in the NW Mediterranean[END_REF]. The increased seawater temperatures in the Mediterranean Sea, predicted to become the norm in the mid-21 st century, should stimulate more intense Ostreopsis spp. blooms in the future [START_REF] Tester | Climate change and harmful benthic microalgae[END_REF] and both Gambierdiscus and Ostreopsis populations are expected to increase, especially in warm locations (Açaf et al., 2020;[START_REF] Tester | Climate change and harmful benthic microalgae[END_REF].

GENERAL INTRODUCTION AND OBJECTIVES …………………………………………..………………………………

Proliferation of Ostreopsis species BOX 2: Ostreopsis Johs.Schmidt, 1901

The genus Ostreopsis, belonging to the family of Ostreopsidaceae (Gonyaulacales, Dinophyceae, Dinozoa) includes several species largely distributed from tropical to temperate marine coastal areas worldwide.

Ostreopsis is a benthic dinoflagellate that develop in close relationship with a biotic or abiotic substrate (Totti et al., 2010). Ostreopsis spp. are usually epiphytic on macrophytes [START_REF] Rhodes | World-wide occurrence of the toxic dinoflagellate genus Ostreopsis Schmidt[END_REF], but can also be found on dead corals, sediments or rocks, constituting the stock of cells (A; [START_REF] Bomber | Genetic variability in toxin potencies among seventeen clones of Gambierdiscus toxicus (dinophyceae). Genet. Var. Toxin Potencies Seventeen Clones Gamb[END_REF]Vila et al., 2001;Shears and Ross, 2009;Totti et al., 2010). Also, due to a combination of physical and biological processes Ostreopsis spp. cells can be found in the water column (B) or forming aggregates on the water surface (C; Mangialajo et al., 2011;[START_REF] Pavaux | Daily variations of Ostreopsis cf. ovata abundances in NW Mediterranean Sea[END_REF][START_REF] Berdalet | Environmental, human health and socioeconomic impacts of Ostreopsis spp. Blooms in the NW Mediterranean[END_REF].

Ostreopsis spp. have been reported for a long time in tropical ciguatera endemic areas and, since some decades, have become common in temperate areas as well [START_REF] Litaker | Taxonomy of Gambierdiscus including four new species, Gambierdiscus caribaeus, Gambierdiscus carolinianus, Gambierdiscus carpenteri and Gambierdiscus ruetzleri (Gonyaulacales, Dinophyceae)[END_REF][START_REF] Rhodes | World-wide occurrence of the toxic dinoflagellate genus Ostreopsis Schmidt[END_REF]. Recurrent Ostreopsis spp. blooms have been recorded throughout the globe (Chang et al., 2000;[START_REF] Rhodes | Toxic marine epiphytic dinoflagellates, Ostreopsis siamensis and Coolia monotis (Dinophyceae)[END_REF]Lenoir et al., 2004;Shears and Ross, 2009;Nascimento et al., 2012b;[START_REF] Yamaguchi | Effects of temperature, salinity and their interaction on growth of the benthic dinoflagellate Ostreopsis cf. ovata (Dinophyceae) from Japanese coastal waters[END_REF]Gomaa et al., 2018;Tibirica et al., 2019;[START_REF] Solino | Toxin Profile of Ostreopsis cf. ovata from Continental Portuguese Coast and Selvagens Islands (Madeira, Portugal)[END_REF]Zou et al., 2020) and in the Mediterranean Sea (Vila et al., 2001;[START_REF] Penna | Characterization of Ostreopsis and Coolia (Dinophyceae) isolates in the western Mediterranean Sea based on morphology, toxicity and internal transcribed spacer 5.8s rDNA sequences[END_REF]Chiantore et al., 2008;Ismael and Halim, 2012;Açaf et al., 2020;Gémin et al., 2020).

Within the eleven species of Ostreopsis identified (Fukuyo, 1981;[START_REF] Norris | Benthic dinoflagellates associated with ciguatera from the Florida Keys. 1. Ostreopsis heptagona sp[END_REF]Quod, 1994;Faust and Morton, 1995;Faust, 1999), at least five produce palytoxin-like compounds, known for being among the most toxic marine compounds (Usami et al., 1995;[START_REF] Ukena | Structure elucidation of ostreocin D, a palytoxin analog isolated from the dinoflagellate Ostreopsis siamensis[END_REF][START_REF] Ciminiello | Complex palytoxin-like profile of Ostreopsis ovata. identification of four new ovatoxins by high-resolution liquid chromatography/mass spectrometry[END_REF][START_REF] Suzuki | LC-MS/MS analysis of novel ovatoxin isomers in several Ostreopsis strains collected in Japan[END_REF]Varela et al., 2021). Humans can be affected by Ostreopsis spp. blooms by the inhalation of marine aerosols which can lead to respiratory disease [START_REF] Tichadou | Health impact of unicellular algae of the Ostreopsis genus blooms in the Mediterranean Sea: Experience of the French Mediterranean coast surveillance network from 2006 to 2009[END_REF][START_REF] Tubaro | Case definitions for human poisonings postulated to palytoxins exposure[END_REF]Vila et al., 2016). The most important human intoxications in temperate locations were recorded in the Mediterranean Sea, in Italy in 2001 and 2005 [START_REF] Brescianini | Ostreopsis ovata algal blooms affecting human health in Genova, Italy, 2005 and 2006[END_REF]) and in Spain in 2004and 2006[START_REF] Barroso García | An epidemic outbreak with respiratory symptoms in the province of Almeria [Spain] due to toxic microalgae exposure [Brote con síntomas respiratorios en la provincia de Almería por una posible exposición a microalgas tóxicas[END_REF], affecting hundreds of locals swimmers and beach goers, mainly affected by respiratory intoxications caused by the inhalation of seawater aerosols containing toxins (Mangialajo et al., 2011). Recurrent events of symptoms on humans due to Ostreopsis spp. blooms have been observed in several parts in the Mediterranean Sea [START_REF] Tubaro | Case definitions for human poisonings postulated to palytoxins exposure[END_REF]Illoul et al., 2012) and in 2021 an important event was recorded for first time in the Atlantic French coast (in Biarritz; [START_REF] Drouet | Current distribution and potential expansion of the harmful benthic dinoflagellate Ostreopsis cf. siamensis towards the warming waters of the Bay of Biscay, North-East Atlantic[END_REF][START_REF] Chomérat | First Characterization of Ostreopsis cf. ovata (Dinophyceae) and Detection of Ovatoxins during a Multispecific and Toxic Ostreopsis Bloom on French Atlantic Coast[END_REF]. The whole ecosystem is also affected by Ostreopsis spp. blooms [START_REF] Turner | Marine invertebrate interactions with Harmful Algal Blooms -implications for One Health[END_REF], and potentially macroalgae [START_REF] Iveša | Increased ammonium levels occurring during benthic algal blooms are potentially toxic to fucalean algae[END_REF]. Most intense blooms of Ostreopsis spp. can result in mass mortalities of invertebrates such as bivalves, gastropods, crustaceans, and echinoderms (Shears and Ross, 2009;[START_REF] Ramos | Palytoxin and analogs: Biological and ecological effects[END_REF]Guidi-Guilvard et al., 2012;[START_REF] Parsons | Gambierdiscus and Ostreopsis: Reassessment of the state of knowledge of their taxonomy, geography, ecophysiology, and toxicology[END_REF]Accoroni and Totti, 2016;Migliaccio et al., 2016;Neves et al., 2018).

Original illustration from Alberto Molina Serrano, http://www.albertomolina.es/.

Surface seawater temperature is thought to be one of the cardinal factors affecting the development of Ostreopsis spp. blooms, while the role played by other abiotic factors such as hydrodynamics, salinity and nutrients is still unclear (Accoroni and Totti, 2016). This lack of knowledge highlights that, as reported in [START_REF] Pavaux | Chemical Ecology of the Benthic Dinoflagellate Genus Ostreopsis : Review of Progress and Future Directions[END_REF], only a small part of the studies on Ostreopsis spp. focus on its ecological aspects. Nevertheless, it seems that the synergic effect of more than one of these factors could influence the development of blooms (Accoroni and Totti, 2016). On one hand, most studies relate larger abundances of Ostreopsis spp. to sheltered or low hydrodynamic conditions (e.g. Chiantore et al., 2008;Shears and Ross, 2009;Richlen and Lobel, 2011;Mohammad-Noor et al., 2016;Asnaghi et al., 2017;Boisnoir et al., 2018;Hachani et al., 2018), while others suggest larger abundances of Ostreopsis spp. in higher or slightly higher levels of water motion (Vila et al., 2001;Selina et al., 2014). The relationship between blooms of Ostreopsis spp. and hydrodynamics could be related to the dense mucilaginous aggregations formed by cells of Ostreopsis spp. covering different substrates (e.g. macrophytes), that can be released due to wave action and suspended in the water column or on the sea surface (Vila et al., 2001;Aligizaki and Nikolaidis, 2006;Chiantore et al., 2008;Shears and Ross, 2009;Totti et al., 2010;[START_REF] Tester | Climate change and harmful benthic microalgae[END_REF].

On the other hand, the inshore occurrence of Ostreopsis spp. has raised the question concerning their association with nutrient enrichments due to human activities (Faust et al., 1996), but there are mixed conclusions on how nutrient concentrations influence population dynamics of Ostreopsis spp. (Ungano et al., 2010;Cohu et al., 2011b;Accoroni et al., 2012). In some studies dinoflagellate abundances were positively correlated with several nutrient parameters, such as nitrates, nitrites, phosphates and silicates (Delgado et al., 2006;Parsons and Preskitt, 2007;Skinner et al., 2013;Accoroni et al., 2020), while other studies found no significant relationship (Okolodkov et al., 2007;Asnaghi et al., 2012;Nascimento et al., 2012a). Nevertheless, abundances of Ostreopsis spp. decrease with depth (Richlen and Lobel, 2011;Cohu and Lemee, 2012), maybe because of the light availability, even if, again, there are controversial results between light intensity and the growth of Ostreopsis spp. in experimental conditions (Accoroni and Totti, 2016).

Preferences for Ostreopsis species proliferation

Due to the benthic nature of Ostreopsis spp. is expected that the substrate plays a major role in the development of blooms. However, substrate preferences for Ostreopsis spp. are still ambiguous sometimes giving discordant patterns (Monti et al., 2007;Totti et al., 2010;Cohu et al., 2013;[START_REF] Blanfuné | Distribution and abundance of Ostreopsis spp. and associated species (Dinophyceae) in the northwestern Mediterranean: the region and the macroalgal substrate matter[END_REF]Moncer et al., 2017;Meroni et al., 2018;Gémin et al., 2020;Ternon et al., 2020). The abundances of Ostreopsis on different substrates are highly variable and linked to several assessment limitations that make comparisons difficult. The comparisons usually depend on the type and characteristics of the substrate (e.g. surface for rocks and fresh weight for macrophytes), ideally not allowing direct comparisons (Mangialajo et al., 2017;[START_REF] Tester | Using artificial substrates to quantify Gambierdiscus and other toxic benthic dinoflagellates for monitoring purposes[END_REF].

In order to avoid the technical issues linked to the measurements different techniques have been proposed such as the use of artificial substrates (Tester et al., 2014;Jauzein et al., 2016;Yong et al., 2018;Fernandez-Zabala et al., 2019;[START_REF] Carreira-Flores | Artificial substrates as sampling devices for marine epibenthic fauna: A quest for standardization[END_REF], the syringe method [START_REF] Abbate | A new method for sampling potentially toxic benthic dinoflagellates[END_REF] and the benthic dinoflagellate integrator device (BEDI; Mangialajo et al., 2017) to obtain comparable estimates of number of cells at multiple sites. However, these techniques are not yet widely used. The high variability on abundances of Ostreopsis spp. among substrates and the discordant patterns, could be due to several biotic and abiotic factors acting at a larger scale than the substrate itself, such as the morphology, palatability, and microbial community associated to the substrate and the herbivory and allelopathic interactions due to the production of secondary metabolites [START_REF] Cruz-Rivera | Macroalgal palatability and the flux of ciguatera toxins through marine food webs[END_REF]Totti et al., 2010;Accoroni et al., 2015;Pavaux et al., 2020;Ternon et al., 2020).

Recent studies focusing on CFP are suggesting that the sea bottom complexity and heterogeneity could affect the development of benthic harmful dinoflagellates (Meroni et al., 2018;Yong et al., 2018;Boisnoir et al., 2019;Bravo et al., 2020;Lee et al., 2020). Some studies suggest that Ostreopsis spp. prefer habitats with high coral cover (Yong et al., 2018) while others associate

Gambierdiscus and Ostreopsis spp. with turf algal communities, suggesting an effect of the architecture of the thallus of the macrophyte on hosting benthic dinoflagellates (Totti et al., 2010;[START_REF] Catania | The influence of macroalgae on the proliferation and regulation of the benthic dinoflagellate Ostreopsis cf[END_REF][START_REF] Mustapa | Growth and epiphytic behavior of three Gambierdiscus species (Dinophyceae) associated with various macroalgal substrates[END_REF]Bravo et al., 2020;Lee et al., 2020). Furthermore, lower abundances of Ostreopsis spp. could be related to Cystoseira s.l. forests [START_REF] Catania | The influence of macroalgae on the proliferation and regulation of the benthic dinoflagellate Ostreopsis cf[END_REF]Meroni et al., 2018). These results are consistent with observations of higher abundances of Ostreopsis spp.

at sites heavily impacted by humans where healthy macroalgal communities are replaced by opportunistic and fast-growing turf macroalgae [START_REF] Rhodes | World-wide occurrence of the toxic dinoflagellate genus Ostreopsis Schmidt[END_REF][START_REF] Fraga | Review of the main ecological features affecting benthic dinoflagellate blooms[END_REF]Meroni et al., 2018;[START_REF] Roselli | Harmful algae and pressure-impact relationship: Noxious blooms and toxic microalgae occurrence from coastal waters of the Apulia region (Adriatic and Ionian Seas, Mediterranean)[END_REF]. According to some authors, the continued and extensive destruction of natural habitats and large metropolitan areas could promote Ostreopsis spp. and Gambierdiscus spp. blooms [START_REF] Rhodes | World-wide occurrence of the toxic dinoflagellate genus Ostreopsis Schmidt[END_REF][START_REF] Fraga | Review of the main ecological features affecting benthic dinoflagellate blooms[END_REF]Meroni et al., 2018;[START_REF] Mustapa | Growth and epiphytic behavior of three Gambierdiscus species (Dinophyceae) associated with various macroalgal substrates[END_REF]Tester et al., 2020;[START_REF] Roselli | Harmful algae and pressure-impact relationship: Noxious blooms and toxic microalgae occurrence from coastal waters of the Apulia region (Adriatic and Ionian Seas, Mediterranean)[END_REF]. The artificialisation of the coast and the miniaturisation and degradation of macroalgal communities due to regime shifts could, therefore, favour the expansion of Ostreopsis spp. [START_REF] Fraga | Review of the main ecological features affecting benthic dinoflagellate blooms[END_REF]Mangialajo et al., 2017;Meroni et al., 2018;Lee et al., 2020;[START_REF] Tester | Climate change and harmful benthic microalgae[END_REF][START_REF] Roselli | Harmful algae and pressure-impact relationship: Noxious blooms and toxic microalgae occurrence from coastal waters of the Apulia region (Adriatic and Ionian Seas, Mediterranean)[END_REF].

OBJECTIVES

The main objective of this thesis is to assess the causes and some potential effects of marine forests loss in the Mediterranean Sea. As causes of loss we assessed the effects of abiotic (climate change)

and biotic (species facilitation and herbivory) factors. As effects of loss, we investigated for the first time the relationship between the present regression of marine forests and the proliferation of benthic harmful algal blooms. [START_REF] Garreta | Flora phycologica ibérica: Fucales[END_REF][START_REF] Rodriguez-Prieto | Guía de las Macroalgas y Fanerógamas Marinas del Mediterráneo Occidental[END_REF].

Image modified from [START_REF] Garreta | Flora phycologica ibérica: Fucales[END_REF].

A B C D

The disappearance of Mediterranean marine forests and their substitution by less structurally complex macroalgal communities could lead to profound changes on ecosystems, facilitating emergent phenomena such as harmful algal blooms. Thus, as an effect of Mediterranean marine forests loss, we selected the emergent phenomena of benthic harmful algal blooms proliferation of the genus Ostreopsis. Benthic harmful algal blooms are causing growing concerns in the Mediterranean Sea given their catastrophic impacts on coastal ecosystems and humans, and their economic consequences (tourism, fishing, etc.; [START_REF] Berdalet | Environmental, human health and socioeconomic impacts of Ostreopsis spp. Blooms in the NW Mediterranean[END_REF]. While Ostreopsis spp. are well studied, not much is known about their ecology [START_REF] Pavaux | Chemical Ecology of the Benthic Dinoflagellate Genus Ostreopsis : Review of Progress and Future Directions[END_REF] and since they settle on benthic surfaces, a better understanding of their relationship with marine vegetation could improve the risk assessment and management of toxic blooms. Therefore, the second objective of this thesis is to study the proliferation of toxic benthic dinoflagellates from the genus Ostreopsis as potential consequence of Cystoseira s.l. forests loss.

The chapters presented hereafter combine field surveys, experiments in the field, controlled experiments in the laboratory and a systematic review. The specific objectives of each chapter are detailed below:

- -Chapter 2. Grazing effects on recruits of the forest-forming fucoid Cystoseira compressa.

Here we examine the effects of different herbivorous species on recruits of C. compressa through (i) field surveys, to characterise the herbivores present in our experimental locations; (ii) experiments in the field, to study the effect of the in-situ herbivory on recruits obtained using different restoration techniques; and (iii) in the laboratory, specifically testing the potential of some selected species to graze on recruits of C. compressa.

-Chapter 3. The role of habitat in the facilitation of Ostreopsis spp. blooms. This chapter is the first step to collect information and better understand the relationship between blooms of Ostreopsis spp. and marine vegetation. In this literature review, we collect information on Ostreopsis spp. blooms at the substrate, habitat, and ecosystem scale, in order to assess eventual preferences and trends for the development of blooms of Ostreopsis spp.

-Chapter 4. Ostreopsis spp. blooms in relation to macroalgal communities. In this chapter we collect information on abundances of Ostreopsis spp. at the substrate, habitat and ecosystem scale during an annual bloom in Rochambeau (Villefranche-sur-Mer, France)

and Vernazzola (Genova, Italy), to assess the preferences and trends of Ostreopsis spp. on different types of substrates (macroalgal and artificial substrates) at different spatial scales.

INTRODUCTION

The ocean plays an important role in the regulation of climate and offers numerous ecosystem services for humans. However, the ocean is affected by multiple anthropogenic impacts including climate change (IPCC, 2022). Ocean warming (OW) and ocean acidification (OA) are expected to affect most marine ecosystems with consequences to humans [START_REF] Doney | Climate Change Impacts on Marine Ecosystems[END_REF][START_REF] Gattuso | Contrasting futures for ocean and society from different anthropogenic CO2 emissions scenarios[END_REF][START_REF] Hall-Spencer | Ocean acidification impacts on coastal ecosystem services due to habitat degradation[END_REF]. Ecosystems all around the globe are expected to experience reductions in habitat structure, biodiversity and trophic complexity as sea temperature rises [START_REF] Straub | Resistance, Extinction, and Everything in Between -The Diverse Responses of Seaweeds to Marine Heatwaves[END_REF] and oceanic pH decreases [START_REF] Kroeker | Divergent ecosystem responses within a benthic marine community to ocean acidification[END_REF][START_REF] Connell | The other ocean acidification problem: CO2 as a resource among competitors for ecosystem dominance[END_REF]. At the same time, an expansion of opportunistic and turf-forming species is foreseen, with the consequent loss of ecosystem services [START_REF] Wernberg | Climate-driven regime shift of a temperate marine ecosystem[END_REF][START_REF] Sunday | Ocean acidification can mediate biodiversity shifts by changing biogenic habitat[END_REF][START_REF] Harvey | Ocean acidification locks algal communities in a species-poor early successional stage[END_REF].

Large brown forest-forming macroalgae (which include the orders Laminariales, Tylopteridales, Desmarestiales, and Fucales) are dominant foundation species on intertidal and subtidal rocky shores in temperate and cold regions (Steneck et al., 2002). They form what is known as marine forests, which [START_REF] Schiel | The Population Biology of Large Brown Seaweeds: Ecological Consequences of Multiphase Life Histories in Dynamic Coastal Environments[END_REF]Wernberg and Filbee-Dexter, 2019) provide important ecosystem functions [START_REF] Cheminée | Nursery value of Cystoseira forests for Mediterranean rocky reef fishes[END_REF][START_REF] Smale | Threats and knowledge gaps for ecosystem services provided by kelp forests: a northeast Atlantic perspective[END_REF][START_REF] Carbajal | Different kelp species support unique macroinvertebrate assemblages, suggesting the potential community-wide impacts of kelp harvesting along the Humboldt Current System[END_REF]. However, CHAPTER 1 ……….…………………….. 22 marine forests are shrinking globally [START_REF] Filbee-Dexter | Rise of Turfs: A New Battlefront for Globally Declining Kelp Forests[END_REF][START_REF] Pessarrodona | Homogenization and miniaturization of habitat structure in temperate marine forests[END_REF] due to several impacts such as urbanisation, marine farming, local pollution and herbivory (Steneck et al., 2002;[START_REF] Krumhansl | Global patterns of kelp forest change over the past half-century[END_REF][START_REF] Orfanidis | Effects of Natural and Anthropogenic Stressors on Fucalean Brown Seaweeds Across Different Spatial Scales in the Mediterranean Sea[END_REF]; making these habitats more sensitive to global change [START_REF] Capdevila | Warming impacts on early life stages increase the vulnerability and delay the population recovery of a long-lived habitat-forming macroalga[END_REF][START_REF] Straub | Resistance, Extinction, and Everything in Between -The Diverse Responses of Seaweeds to Marine Heatwaves[END_REF]. The structure and productivity of marine forests are influenced by many environmental factors that drive the growth, survival, reproduction and metabolism of the organisms, which in turn affect the whole habitat or ecosystem [START_REF] Irving | Light, sediment, temperature, and the early life-history of the habitat-forming alga Cystoseira barbata[END_REF][START_REF] Smale | Climate-driven substitution of foundation species causes breakdown of a facilitation cascade with potential implications for higher trophic levels[END_REF][START_REF] Smith | Examining the influence of regional-scale variability in temperature and light availability on the depth distribution of subtidal kelp forests[END_REF]. Global change effects on marine forests can vary according to the location, the population characteristics and the species [START_REF] Krumhansl | Global patterns of kelp forest change over the past half-century[END_REF][START_REF] Hollarsmith | Varying reproductive success under ocean warming and acidification across giant kelp (Macrocystis pyrifera) populations[END_REF][START_REF] Verdura | Localscale climatic refugia offer sanctuary for a habitat-forming species during a marine heatwaves[END_REF]. As a result, in several cases, marine forests are constrained to locations with the most favourable conditions which could act as a refuge [START_REF] Mariani | Past and present of Fucales from shallow and sheltered shores in Catalonia[END_REF][START_REF] Smale | Impacts of ocean warming on kelp forest ecosystems[END_REF][START_REF] Verdura | Localscale climatic refugia offer sanctuary for a habitat-forming species during a marine heatwaves[END_REF]. There is evidence that early-life stages of these species are more vulnerable than adults which could lead, in the long term, to the loss of marine forests [START_REF] Coelho | Impacts of anthropogenic stresses on the early development stages of seaweeds[END_REF][START_REF] Schiel | The Population Biology of Large Brown Seaweeds: Ecological Consequences of Multiphase Life Histories in Dynamic Coastal Environments[END_REF][START_REF] De Caralt | Differential effects of pollution on adult and recruits of a canopy-forming alga: implications for population viability under low pollutant levels[END_REF]. A high mortality rate is naturally observed during the early stages and the resilience of a population to future impacts can be largely dependent on efficient recruitment and development of juveniles [START_REF] Vadas | Recruitment and mortality of early postsettlement stages of benthic algae[END_REF][START_REF] Capdevila | Recruitment patterns in the Mediterranean deep-water alga Cystoseira zosteroides[END_REF].

The shift in carbonate chemistry associated with OA causes an increase in dissolved CO2 that could favour photosynthesis and then the growth of photosynthetic organisms [START_REF] Connell | The other ocean acidification problem: CO2 as a resource among competitors for ecosystem dominance[END_REF][START_REF] Koch | Climate change and ocean acidification effects on seagrasses and marine macroalgae[END_REF]. The increase in CO2 modifies the dissolved CO2 to O2 ratio at the RuBisCO active site, i.e., the key enzyme in carbon fixation metabolism. Because the latter emerged in an oxygen poor environment, it is characterised by a higher affinity for O2 than CO2 [START_REF] Shih | Biochemical characterization of predicted Precambrian RuBisCO[END_REF].

Therefore, the current increase in dissolved CO2 to O2 ratio favours RuBisCO carbon fixation efficiency and then can favour the growth of photosynthetic organisms. To increase the RuBisCO carbon fixation efficiency, many algae also developed carbon concentration mechanisms (CCM), that increase the CO2 to O2 ratio in front of the RuBisCO fixation site [START_REF] Connell | The other ocean acidification problem: CO2 as a resource among competitors for ecosystem dominance[END_REF]Cornwall et al., 2017a). Still, despite having or not having CCM it is not clear if most algae respond positively to an increase of CO2 [START_REF] Hepburn | Diversity of carbon use strategies in a kelp forest community: implications for a high CO2 ocean[END_REF][START_REF] Connell | The other ocean acidification problem: CO2 as a resource among competitors for ecosystem dominance[END_REF]Cornwall et al., 2017a). OA might, therefore, have beneficial effects for some species like large brown forest-forming macroalgae, that are thought to thrive at high CO2 concentrations [START_REF] Hepburn | Diversity of carbon use strategies in a kelp forest community: implications for a high CO2 ocean[END_REF][START_REF] Porzio | Effects of ocean acidification on macroalgal communities[END_REF][START_REF] Connell | The other ocean acidification problem: CO2 as a resource among competitors for ecosystem dominance[END_REF]. However, calcifying organisms (e.g. foundation species like corals and coralline algae) are expected to be particularly affected by OA (Kroeker et al., 2013a). In particular, crustose coralline algae, which are important components of the understory of marine forests, are among the organisms potentially the most susceptible to OA [START_REF] Kroeker | Meta-analysis reveals negative yet variable effects of ocean acidification on marine organisms: Biological responses to ocean acidification[END_REF][START_REF] Rindi | Coralline Algae in a Changing Mediterranean Sea: How Can We Predict Their Future, if We Do Not Know Their Present?[END_REF]. This species can be directly and indirectly impacted by OA due to reduced calcification rates and increasing competition with algae which benefit from elevated CO2 [START_REF] James | Growth response of an early successional assemblage of coralline algae and benthic diatoms to ocean acidification[END_REF][START_REF] Comeau | Contrasting Effects of Ocean Acidification on Coral Reef 'Animal Forests' Versus Seaweed 'Kelp Forests[END_REF]Cornwall et al., 2017a). Crustose coralline algae are among the first colonizers CHAPTER 1 ……….…………………….. of bare rock on euphotic marine habitats and are quickly overgrown by later successional species such as the more structurally complex large brown forest-forming macroalgae [START_REF] Airoldi | EFFECTS OF DISTURBANCE, LIFE HISTORIES[END_REF][START_REF] Asnaghi | Colonisation processes and the role of coralline algae in rocky shore community dynamics[END_REF]. Some authors (Bulleri et al., 2002b) report that crustose coralline algae could help in the maintenance of alternative habitat states by preventing the recruitment of later colonizers (e.g. large brown macroalgae), even if this could be species-specific [START_REF] Villas Bôas | Are anti-fouling effects in coralline algae species specific?[END_REF]. While other studies suggest that crustose coralline algae could, in contrast, enhance biodiversity by facilitating the settlement of later colonists, including invertebrates (Bulleri et al., 2002a;[START_REF] Maggi | Connell and Slatyer's models of succession in the biodiversity era[END_REF][START_REF] Asnaghi | Colonisation processes and the role of coralline algae in rocky shore community dynamics[END_REF] and by creating a positive association with the forest-forming macroalgae [START_REF] Melville | Experimental effects of kelp canopies on subtidal coralline algae[END_REF][START_REF] Irving | Effects of kelp canopies on bleaching and photosynthetic activity of encrusting coralline algae[END_REF][START_REF] Irving | The response of encrusting coralline algae to canopy loss: an independent test of predictions on an Antarctic coast[END_REF]. Thus, a reduction of crustose coralline algae cover, because of climate change, may affect the recruitment of forest-forming macroalgae and therefore the maintenance of marine forest habitats [START_REF] Breitburg | Residual Effects of Grazing: Inhibition of Competitor Recruitment by Encrusting Coralline Algae[END_REF][START_REF] Bulleri | Facilitation and the niche: implications for coexistence, range shifts and ecosystem functioning[END_REF] and their resistance against climate change [START_REF] Comeau | Contrasting Effects of Ocean Acidification on Coral Reef 'Animal Forests' Versus Seaweed 'Kelp Forests[END_REF][START_REF] Van Der Heide | Facultative mutualisms: A double-edged sword for foundation species in the face of anthropogenic global change[END_REF].

In Mediterranean rocky bottoms, Cystoseira sensu lato species (including the genera Cystoseira, Ericaria and Gongolaria, hereafter referred to as Cystoseira) are the main representatives of marine forests (Wernberg and Filbee-Dexter, 2019;[START_REF] Molinari -Novoa | Reinstatement of the genera Gongolaria Boehmer and Ericaria Stackhouse (Sargassaceae, Phaeophyceae)[END_REF]. However, only a few studies have investigated the effect of climate change on this taxon and even fewer have focused on their early stages (Celis-Plá et al., 2017a;[START_REF] Hernández | Macroalgal response to a warmer ocean with higher CO2 concentration[END_REF][START_REF] Capdevila | Warming impacts on early life stages increase the vulnerability and delay the population recovery of a long-lived habitat-forming macroalga[END_REF][START_REF] Falace | Is the South-Mediterranean Canopy-Forming Ericaria giacconei (= Cystoseira hyblaea) a Loser From Ocean Warming? Front[END_REF]. Most studies show a negative impact of OW for both recruits and adults of Cystoseira on their survival [START_REF] Falace | Ex situ cultivation protocol for Cystoseira amentacea var. stricta (Fucales, Phaeophyceae) from a restoration perspective[END_REF][START_REF] Falace | Is the South-Mediterranean Canopy-Forming Ericaria giacconei (= Cystoseira hyblaea) a Loser From Ocean Warming? Front[END_REF][START_REF] Verdura | Localscale climatic refugia offer sanctuary for a habitat-forming species during a marine heatwaves[END_REF], resilience [START_REF] Capdevila | Warming impacts on early life stages increase the vulnerability and delay the population recovery of a long-lived habitat-forming macroalga[END_REF] and phenology [START_REF] Bevilacqua | Climatic anomalies may create a long-lasting ecological phase shift by altering the reproduction of a foundation species[END_REF][START_REF] Savonitto | Addressing reproductive stochasticity and grazing impacts in the restoration of a canopyforming brown alga by implementing mitigation solutions[END_REF]. In contrast, decreasing pH increased the productivity, antioxidant activity and production of photoprotective compounds of adult Cystoseira (Celis-Plá et al., 2017a;[START_REF] Hernández | Macroalgal response to a warmer ocean with higher CO2 concentration[END_REF]. Despite that, some species of Cystoseira (including Cystoseira compressa and Cystoseira foeniculacea) are considered CCM species whose CCM does not downregulate due to additional CO2 and, thus, could not benefit from increasing CO2 (Cornwall et al., 2017a). To our knowledge, there are no studies on the combined effects of OW and OA on early-life stages or recruits of Cystoseira, and only one study (Celis-Plá et al., 2017a) has investigated the effects of both drivers on adults.

Here, we tested the interactive effects of temperature, pH and species facilitation on the recruitment of Cystoseira compressa. This species is a common forest-forming macroalgae that can create dense populations on shallow and sheltered rocky shores around the Mediterranean Sea (Mangialajo et al., 2012). It is considered one of the most resistant Cystoseira species and it is the only one that is not protected under the Barcelona Convention (Annex II; United Nations Environment Programme/Mediterranean Action Plan-UNEP/MAP; [START_REF] Verlaque | Mediterranean seaweeds listed as threatened under the Barcelona Convention: A critical analysis[END_REF]. We designed two separate experiments to test the effects of elevated temperature, low pH and the presence of crustose coralline algae on the early-life stages of C. compressa. The first experiment focused on the effects CHAPTER 1 ……….…………………….. of the temperature on the recruits of C. compressa. Based on the results of the first experiment, we ran a second complementary experiment to assess the role of the interactive effects of temperature, pH and species facilitation (crustose coralline algae) on the recruitment of C. compressa. Because coralline algae are sensitive to OA and are a potentially favourable substrate for the recruitment of Cystoseira, we assessed the recruitment of C. compressa on living and dead Neogoniolithon brassica-florida, one of the most common species in association with shallow Cystoseira forests [START_REF] Benedetti-Cecchi | Effects of canopy cover, herbivores and substratum type on patterns of Cystoseira spp. settlement and recruitment in littoral rockpools[END_REF][START_REF] Asnaghi | Colonisation processes and the role of coralline algae in rocky shore community dynamics[END_REF]. We also compared the recruitment of Cystoseira on abiotic artificial clay substrates that have been proposed as an efficient substrate for restoration [START_REF] Falace | Ex situ cultivation protocol for Cystoseira amentacea var. stricta (Fucales, Phaeophyceae) from a restoration perspective[END_REF][START_REF] De La Fuente | First ex situ outplanting of the habitat-forming seaweed Cystoseira amentacea var. stricta from a restoration perspective[END_REF][START_REF] Orlando-Bonaca | First Restoration Experiment for Gongolaria barbata in Slovenian Coastal Waters[END_REF]. The main hypothesis of this study is that climate change will negatively affect the recruitment of C. compressa. Our hypotheses are that OW may have a direct negative impact on the recruits of C. compressa while OA may increase their growth and productivity. We also hypothesize that the settlement and survival of C. compressa might be indirectly affected by the effects of climate change on its associated understory species (crustose coralline algae) that act as a substrate.

MATERIALS AND METHODS

Experiment 1: Effects of ocean warming on the recruitment of Cystoseira compressa

Collection and obtention of recruits

Apical fertile branches of Cystoseira compressa were hand-collected on the 13th of July 2020 from a donor population situated in a rockpool (between the surface and 1m depth) in Sainte Marguerite Island (Lérins Islands, France). This site is a Nature 2000 site situated in front of the coast of Cannes and it is one of the last locations with healthy Cystoseira populations in the French Riviera [START_REF] Thibaut | Decline and local extinction of Fucales in French Riviera: the harbinger of future extinctions?[END_REF]. After visually checking that the receptacles contained fertile conceptacles, about 140 gram fresh weight (FW) of apical fertile branches were manually collected and transported in cool and dark conditions in plastic bags to the laboratory. The sampling was non-destructive, as only apical branches (roughly 5 cm long) were collected. A temperature data logger (HOBO Pendant MX Temp, ONSET), that took measurements every hour, was installed in the rock pool to monitor the temperature in the donor population site. The receptacles were conserved at 4°C in the dark overnight before placing them in experimental tanks filled with filtered seawater (20 µm) and marble substrates that acted as settlement substrates [START_REF] Falace | Ex situ cultivation protocol for Cystoseira amentacea var. stricta (Fucales, Phaeophyceae) from a restoration perspective[END_REF][START_REF] Verdura | Restoration of a Canopy-Forming Alga Based on Recruitment Enhancement: Methods and Long-Term Success Assessment[END_REF][START_REF] Orlando-Bonaca | First Restoration Experiment for Gongolaria barbata in Slovenian Coastal Waters[END_REF]. In each tank, 15g FW of receptacles were placed in a net on the surface allowing the zygotes to fall on the substrates. The receptacles were kept in the tanks for 72 hours without water circulation to facilitate the settlement of the zygotes. The temperature was maintained during the releasing and the settlement of the zygotes at the target temperature treatment. After opening the water system, the receptacles were removed and the recruits were kept CHAPTER 1 ……….…………………….. in tanks for 96 days. Submersible water pumps (NEWA) provided water motion in each experimental tank.

Experimental set-up and treatments

Three independent 5 L tanks (n=3) were set up for each temperature (24, 28 and 32°C; Table 1), for a total of 9 experimental tanks, with three square marble substrates of about 25 cm 2 placed inside each tank. Filtered seawater (20 µm) pumped from Villefranche Bay at 3 m depth was continuously delivered into the experimental tanks at a rate of 7 L h -1 . The experimental tanks were placed inside a thermoregulated bath to maintain the temperature at the targeted value. The temperature was controlled in two thermoregulated baths per treatment with a temperature controller (T CONTROLLER TWIN AQUA MEDIC). Light was provided by 37 W LED light bars (PRO² LED, Aquaristik) and the irradiance gradually increased from 0 μmol photons m -2 s -1 at 06:30 to a maximum of 110 μmol photons m -2 s -1 between 12:00 and 14:00, and gradually decreased to 0 μmol photons m -2 s -1 at 21:00 (LI-185B with an LI-190SB quantum sensor, LI-COR Biosciences, Lincoln, USA). The three temperature treatments were selected according to the temperatures registered in the donor population during the reproductive season of C. compressa and the expected increase in temperature due to global warming.

Measurements

The density and size of C. compressa recruits were selected as response variables. The density of recruits was calculated by taking pictures every 10 or 15 days and counting from the picture the total number of recruits on a 3x3 cm area in the middle of each substrate using the software ImageJ CHAPTER 1 ……….…………………….. (ImageJ, NIH US Department of Health and Human Services). The size of recruits (µm) was determined monthly on 5 recruits that were removed from each substrate (n = 45). Their total length was measured using a microscope equipped with a graduated eyepiece.

Experiment 2: Effects of climate change and species facilitation on the recruitment of Cystoseira compressa

Collection and obtention of recruits

Apical fertile branches of C. compressa were collected on the 4 th of August 2021 from the same donor population as mentioned above and following the same protocol (see section 2.1.1.). For this experiment, about 500 g FW of apical fertile branches were manually collected and transported in cool and dark conditions in plastic bags to the laboratory. The receptacles were conserved at 4°C in the dark overnight before placing them under the experimental conditions, in the experimental tanks, to obtain recruits on the different substrates and under the different conditions of temperature and acidification. In each tank, 10 g FW of receptacles placed inside a net were kept on the surface for 72 hours. During this period, the temperature and pH were maintained at the target treatment conditions. Afterwards, the receptacles were removed and the recruits were kept in tanks for 75 days.

Experimental set-up and treatments

The recruitment of Cystoseira compressa was assessed in two different conditions of temperature, 28 and 32°C, and two pH levels, ambient (pHT = 8.07) and low pH (pHT = 7.8). In order to investigate the potential facilitation effect of Neogoniolithon brassica-florida, three different substrates were used for the settlement of recruits: 1) living and 2) dead N. brassica-florida (respectively factors "coralline" and "dead coralline") and 3) artificial clay substrates (factor "artificial substrate"). The temperatures were selected according to the results obtained during the first experiment (experiment 1). The low pH condition (pHT = 7.8) corresponds to the pH value expected by the end of the century under the SSP2 -4.5 CO2 emissions scenario [START_REF] Celis-Plá | Macroalgal responses to ocean acidification depend on nutrient and light levels[END_REF][START_REF] Kwiatkowski | Twenty-first century ocean warming, acidification, deoxygenation, and upper-ocean nutrient and primary production decline from CMIP6 model projections[END_REF]. The surface of the three substrates was about 10 cm 2 (3 cm diameter and 1 cm height for the artificial substrates). The coralline algae were collected in April 2021 from Anse des Fossés (Saint-Jean-Cap-Ferrat, France) between 0.5 and 1 m depth. Samples that were the most homogeneous and the least colonized by other organisms were selected. Then, they were cleaned and epiphytes were removed using brushes and tweezers. Dead coralline substrates were obtained by putting the N. brassica-florida substrates in freshwater with bleach (1:50) for 24 hours. They were then rinsed several times with freshwater and dried, before placing them in the tanks. All the substrates were placed in the experimental tanks at ambient seawater CHAPTER 1 ……….…………………….. temperature and pH and gradually brought to the experimental temperatures and pH levels over 3 months before the start of the experiment.

The experimental set-up consisted of four independent 1.8 L tanks (n=4) for each of the 12 conditions (Substrate type × pH × Temperature) for a total of 48 experimental tanks. Each tank contained five replicates of a substrate (either artificial, coralline or dead coralline substrates). One substrate from each experimental tank was used as a control and was not seeded with zygotes of C. compressa. Seawater from the Bay of Villefranche was continuously delivered into eight 20 L header tanks that then gravity fed six 1.8 L independent experimental tanks each with a water rate of 3 L h -1 (Figure 1). Seawater pH was manipulated inside 4 20 L header tanks. pH was maintained at the target value using pH controllers (APEX, Neptune Systems) that controlled the bubbling of pure CO2 in the header tanks. The experimental tanks were placed inside thermoregulated baths (four per temperature) connected to the same control system (APEX, Neptune Systems; Figure 6) to maintain the temperature at 28 and 32°C. Submersible water pumps (NEWA) provided water motion in each experimental tank.

Light was provided by 89 W LED light bars (Aqualumix, Aquaristik) and irradiance gradually increased from 0 μmol photons m -2 s -1 at 06:30 to a maximum of 175 μmol photons m -2 s -1 between 12:00 and 14:00, and gradually decreased to 0 μmol photons m -2 s -1 at 21:00 (LI-185B with an LI-190SB quantum sensor, LI-COR Biosciences, Lincoln, USA; [START_REF] Verdura | Localscale climatic refugia offer sanctuary for a habitat-forming species during a marine heatwaves[END_REF].

Carbonate chemistry

pH in the header and experimental tanks was measured weekly using a handheld pH-meter (826 pH mobile, Metrohm) calibrated with TRIS buffer (batch #T33 provided by A. Dickson, Scripps Institution of Oceanography). Total alkalinity was measured weekly in eight randomly selected tanks and was determined by potentiometric titration using a Metrohm 888 Titrando following the method of Dickson et al. [START_REF] Dickson | Guide to Best Practices for Ocean CO2 Measurements[END_REF], the samples were measured three times and the mean value was used. Certified reference material (Batch #186) provided by A. Dickson was used to assess the accuracy of the measurements and was within 7.73 µmol kg -1 . The seawater from the Bay of Villefranche was 2565.41 ± 12.1 μmol kg -1 (mean ± SD; Table 1; Appendix A: Chapter 1 Supplementary material S1).

Measurements

The density and size of recruits of C. compressa and calcification rate of the living coralline algae substrates were selected as response variables. The density of recruits was assessed by counting directly in the tanks the total number of recruits on the substrates using a magnifying table lamp.

The total number of recruits was normalized by the surface of the substrate. The size of the recruits was determined by measuring the length of ten individuals randomly picked from each substrate (n = 160). When ten or fewer individuals were present on the substrate, all of them were measured.

The measurements were done using graph paper under a magnifying table lamp.

Total calcification rate of N. brassica-florida was assessed using the buoyant weight technique [START_REF] Davies | Short-term growth measurements of corals using an accurate buoyant weighing technique[END_REF]. Weighing was done before obtaining C. compressa recruits and at the end of the experiment, 64 days later. Changes in wet weight were converted to dry weight using the following equation:

𝐷𝑟𝑦 𝑤𝑒𝑖𝑔ℎ𝑡 = 𝑊𝑒𝑡 𝑤𝑒𝑖𝑔ℎ𝑡 (1 - 𝑊𝑎𝑡𝑒𝑟 𝑑𝑒𝑛𝑠𝑖𝑡𝑦 𝐶𝑎𝑙𝑐𝑖𝑡𝑒 𝑑𝑒𝑛𝑠𝑖𝑡𝑦 )
with a calcite density of 2.73g cm -3 . Calcification rate was determined as the change in dry weight normalized by the surface of coralline algae at the moment of the weighting and the number of days

Data analysis

Experiment 1: A Generalized Linear Mixed-Effects Model (GLMM), with a Poisson link log distribution function was used to test the effect of the temperature on the density of recruits, with temperature (three levels) and time (seven levels) as fixed factors, and substrate nested within tank as random. A two-way ANOVA was used to test the effect of temperature on the size of the recruits, with temperature (three levels) and time (three levels) as fixed factors. The assumptions of normality and equality of variance were evaluated through graphical analyses of residuals using QQ plot functions. An alpha of 0.01 was used when the assumption of equality of variance was not achieved.

Experiment 2: A GLMM with a Poisson distribution was used to test the effect of temperature, pH and substrate type on the density of recruits, with temperature (two levels), pH (two levels), substrate type (three levels) and time (five levels) as fixed factors and substrate nested within tanks as random, to account for the lack of independence between observations (repeated measures over time). The response of the variable size to the treatments was analysed using a GLMM with a Gamma error distribution function and the logit link function 'inverse', with temperature (two levels), pH (two levels), substrate type (three levels) and time (three levels) as fixed factors and tank as random. The total calcification rate of the coralline algae was analysed with a Linear Model (LM), with temperature (two levels) and pH (two levels) as fixed factors.

GLMM and LM models were fitted to analyse the effect of the variables and the AICs likelihood minimum was used to select the best model among the possible combinations. The different models were fitted using the functions "glmer" and "lm" from the package lme4 [START_REF] Bates | Fitting Linear Mixed-Effects Models using lme4[END_REF] in the statistical environment R (R: The R Project for Statistical Computing). P-values were obtained by means of a Wald χ 2 test using the "ANOVA" function from the CAR package [START_REF] Fox | An R Companion to Applied Regression[END_REF]. Finally, the function "emmeans" from the package emmeans [START_REF] Lenth | emmeans: Estimated Marginal Means, aka Least-Squares Means[END_REF] was used to perform the post-hoc analysis of the LM and GLMM models while the test "snk" (Student-Newman-Keuls) was used to perform the post-hoc analysis for the two-way ANOVA.

RESULTS

In-situ temperatures at the donor population location

The temperatures at the Cystoseira compressa donor population site during the periods that cover the first (14 th July-13 th October 2020) and second experiment (10 th August -20 th October 2021) varied between minimums and maximum values of 15.2°C (27 th October) and 29.5°C (2 nd August) in 2020 and between 19.4°C (14 th July) and 29.0°C (22 nd July) in 2021 (Figure 2).

Experiment 1: Effects of ocean warming on the recruitment of Cystoseira compressa

The temperature negatively affected the density of recruits since the beginning of the experiment (GLMM, P-value < 0.001; Appendix A: Chapter 1 Supplementary material S2). Recruit density was significantly lower at 32°C than at 28°C and 24°C since the first sampling dates (day 10; Figure 3a), while from day 36 densities at 24°C remained higher than the ones at 28°C and 32°C (Figure 3a). The temperature also affected the size of the recruits at the end of the experiment, recruits at 24°C being significantly larger than the ones grown at warmer temperatures (ANOVA, P-value < 0.001; Appendix A: Chapter 1 Supplementary material S2 and Figure 3b).

Experiment 2: Effects of climate change and species facilitation on the recruitment of Cystoseira compressa

Recruits of Cystoseira compressa

The temperature and pH, separately and together with substrate type and time, had an interactive effect on the density of recruits (GLMM, P-value < 0.001; Appendix A: Chapter 1 Supplementary material S3). There was a strong negative effect of the temperature on the density of recruits, especially in presence of Neogolithon brassica-florida (Figure 4). Low pH negatively affected the density of recruits in absence of N. brassica-florida, but no differences in density between pH levels were detected in presence of the coralline algae (Appendix A: Chapter 1 Supplementary material S3 and S4). The presence of the living and dead coralline algae negatively affected the density of recruits of C. compressa, compared to the higher densities found on artificial substrates. The interaction among temperature, pH and time (Temperature × pH × Time) also affected the density of recruits (GLMM, P-value < 0.001; Appendix A: Chapter 1 Supplementary material S3). Higher densities of recruits were found at low temperature-ambient pH while no differences between pH levels were detected at elevated temperature (Appendix A: Chapter 1 Supplementary material S3).

In general, higher densities of recruits were found at low temperature, ambient pH and in absence of N. brassica-florida. The density of recruits decreased with time which led to a homogenization of results for most of the considered factors at the end of the experiment (significant differences in pH and temperature were found only in absence of the coralline algae and at low temperature, Appendix A: Chapter 1 Supplementary material S4). This is because only a few recruits survived until the end of the experiment (Figure 4). ). The size of recruits was larger at low temperature and low pH. The size of recruits was also larger in absence of N. brassica-florida in all the treatment combinations (Figure 5). Differences in size were observed between recruits on living and dead N. brassica-florida, but only at low temperature. By the end of the experiment, recruits grown in association with the living coralline algae were smaller, while there were no differences in size between recruits grown on dead N. brassica florida and in clay substrates (Figure 5; Appendix A: Chapter 1 Supplementary material S5).

Calcification of Neogoniolithon brassica-florida

The temperature and pH negatively affected the net calcification of living Neogoniolithon brassicaflorida (LM, P-value < 0.01 for Temperature and P-value < 0.05 for pH; Appendix A: Chapter 1 Supplementary material S3). Calcification rates were statistically significantly higher at 28°C than at 32°C and ambient pH than at low pH (Figure 6).

DISCUSSION

Marine forests are in regression in many locations worldwide (Filbee-Dexter and [START_REF] Filbee-Dexter | Rise of Turfs: A New Battlefront for Globally Declining Kelp Forests[END_REF][START_REF] Christie | Shifts Between Sugar Kelp and Turf Algae in Norway: Regime Shifts or Fluctuations Between Different Opportunistic Seaweed Species? Front[END_REF][START_REF] Pessarrodona | Homogenization and miniaturization of habitat structure in temperate marine forests[END_REF] and particularly in the Mediterranean Sea [START_REF] Thibaut | Long-term decline of the populations of Fucales (Cystoseira spp. and Sargassum spp.) in the Albères coast (France, Northwestern Mediterranean)[END_REF][START_REF] Thibaut | Decline and local extinction of Fucales in French Riviera: the harbinger of future extinctions?[END_REF]Orlando-Bonaca et al., 2021a). Despite this, the response of the different life stages to climate change remains poorly known, efforts have been done in recent years to evaluate the single or combined effect of climate change on forest-forming species together with other local stressors such as pollution, highlighting and strong negative effect of OW [START_REF] Leal | Copper pollution exacerbates the effects of ocean acidification and warming on kelp microscopic early life stages[END_REF][START_REF] Capdevila | Warming impacts on early life stages increase the vulnerability and delay the population recovery of a long-lived habitat-forming macroalga[END_REF][START_REF] Fernández | Effect of environmental history on the habitat-forming kelp Macrocystis pyrifera responses to ocean acidification and warming: a physiological and molecular approach[END_REF]. Our results showed that the temperature, pH and the presence of potentially facilitating species had substantial effects on the density of recruits of Cystoseira compressa. The warmer temperature had, in agreement with our initial hypotheses, the largest negative effect on both the density and the size of recruits, as already observed on other forest-forming species, both on recruits [START_REF] Leal | Copper pollution exacerbates the effects of ocean acidification and warming on kelp microscopic early life stages[END_REF][START_REF] Capdevila | Warming impacts on early life stages increase the vulnerability and delay the population recovery of a long-lived habitat-forming macroalga[END_REF][START_REF] Verdura | Localscale climatic refugia offer sanctuary for a habitat-forming species during a marine heatwaves[END_REF] and adults [START_REF] Lind | Effects of abiotic stressors on kelp early life-history stages[END_REF][START_REF] Fernández | Nitrogen sufficiency enhances thermal tolerance in habitat-forming kelp: implications for acclimation under thermal stress[END_REF][START_REF] Smale | Impacts of ocean warming on kelp forest ecosystems[END_REF][START_REF] Falace | Is the South-Mediterranean Canopy-Forming Ericaria giacconei (= Cystoseira hyblaea) a Loser From Ocean Warming? Front[END_REF].

Warmer temperatures not only affect the survival and growth of Cystoseira but also affect their metabolism [START_REF] Celis-Plá | Macroalgal responses to ocean acidification depend on nutrient and light levels[END_REF], 2017a;[START_REF] Mancuso | Influence of ambient temperature on the photosynthetic activity and phenolic content of the intertidal Cystoseira compressa along the Italian coastline[END_REF]. A study on C. compressa reported that the maximum quantum yield (Fv/Fm) of the macroalgae started decreasing from 28°C, while the total phenolic content increased with seawater temperature [START_REF] Mancuso | Influence of ambient temperature on the photosynthetic activity and phenolic content of the intertidal Cystoseira compressa along the Italian coastline[END_REF]. Globally, the increase in seawater temperature is a direct threat to marine forests, isolating forest-forming macroalgae to refuge locations with more suitable conditions, while consequent local extinctions can be expected in the northern limits of distribution of some species (e.g. the northern limit of the Mediterranean Sea; [START_REF] Mariani | Past and present of Fucales from shallow and sheltered shores in Catalonia[END_REF][START_REF] Verdura | Localscale climatic refugia offer sanctuary for a habitat-forming species during a marine heatwaves[END_REF]. Moreover, indirect temperaturedriven effects on marine forests are not negligible, because they contribute to the tropicalization of CHAPTER 1 ……….…………………….. 34 habitats and the range expansion of warm water species that can re-shape algal communities and trophic cascades (Vergés et al., 2014a(Vergés et al., , 2014b)).

Seawater pH affected both the density and size of recruits in opposite ways, as density was negatively affected by the low pH, whereas size was positively affected. Low pH has been shown to negatively influence the settlement and early-life stages of other key species such as the giant kelp [START_REF] Gaitán-Espitia | Interactive effects of elevated temperature and pCO2 on early-life-history stages of the giant kelp Macrocystis pyrifera[END_REF][START_REF] Hollarsmith | Varying reproductive success under ocean warming and acidification across giant kelp (Macrocystis pyrifera) populations[END_REF], corals and molluscs (Kroeker et al., 2013a) which has led to the paradigm that early-life stages could be more sensitive to global change and therefore could constitute a bottleneck [START_REF] Capdevila | Warming impacts on early life stages increase the vulnerability and delay the population recovery of a long-lived habitat-forming macroalga[END_REF][START_REF] Falace | Is the South-Mediterranean Canopy-Forming Ericaria giacconei (= Cystoseira hyblaea) a Loser From Ocean Warming? Front[END_REF]. However, in our experiment, lower pH levels positively affected the size of C. compressa, potentially showing a better performance as reported in some studies for the giant kelp [START_REF] Roleda | Ocean acidification and seaweed reproduction: increased CO2 ameliorates the negative effect of lowered pH on meiospore germination in the giant kelp Macrocystis pyrifera (Laminariales, Phaeophyceae)[END_REF][START_REF] Leal | Ocean acidification and kelp development: Reduced pH has no negative effects on meiospore germination and gametophyte development of Macrocystis pyrifera and Undaria pinnatifida[END_REF]. Non-calcifying macroalgae are generally considered as not particularly sensitive to OA as they can benefit from increasing dissolved CO2, particularly the carbon-limited species that do not possess a CCM [START_REF] Hepburn | Diversity of carbon use strategies in a kelp forest community: implications for a high CO2 ocean[END_REF][START_REF] Connell | The other ocean acidification problem: CO2 as a resource among competitors for ecosystem dominance[END_REF]. The increase in dissolved CO2 with OA is therefore expected to positively affect carbon-limited species and neutrally or positively affect non-carbon-limited species with CCM [START_REF] Hepburn | Diversity of carbon use strategies in a kelp forest community: implications for a high CO2 ocean[END_REF]. Most large brown macroalgal species have attributes suggesting the presence of CCM, and yet they benefit from increased CO2 [START_REF] Celis-Plá | Macroalgal responses to ocean acidification depend on nutrient and light levels[END_REF], 2017a;[START_REF] Zhang | Elevated CO2 concentrations promote growth and photosynthesis of the brown alga Saccharina japonica[END_REF]. The same seems to happen here with the beneficial effects of high CO2 on the size of the recruits. C. compressa, possessing a high affinity CCM for dissolved inorganic carbon (DIC), would not change the activity of the CCM due to OA and, thus, would not specially benefit under elevated CO2. As a result, they would potentially end up being less competitive than other species that will benefit from elevated CO2 (Cornwall et al., 2017a). It is important to note that in some of our experimental conditions (i.e. 32°C and low pH), the biggest sizes corresponded to the lowest densities, which does not allow us to exclude the effect of density-dependent processes in addition to the effect of CO2. OA could also have indirect effects on marine forests, by favouring the increased performance of turfs that are generally carbon-limited, especially in the presence of nutrients (e.g. local nutrient pollution; [START_REF] Connell | The other ocean acidification problem: CO2 as a resource among competitors for ecosystem dominance[END_REF][START_REF] Falkenberg | Contrasting resource limitations of marine primary producers: implications for competitive interactions under enriched CO2 and nutrient regimes[END_REF]. Turf-forming species are fast-growing and therefore are great space competitors that could limit the recruitment of long-life species such as forest-forming species. As a result, turfforming species could expand and replace foundation species (e.g. Cystoseira s.l. spp.) that are already affected by global warming and other anthropogenic impacts [START_REF] Nagelkerken | Ocean acidification alters fish populations indirectly through habitat modification[END_REF][START_REF] Filbee-Dexter | Rise of Turfs: A New Battlefront for Globally Declining Kelp Forests[END_REF]. This indirect effect of OA on marine forests might decrease the structural complexity of marine forests, compromising their functioning and promoting regime shifts [START_REF] Connell | The direct effects of increasing CO2 and temperature on non-calcifying organisms: increasing the potential for phase shifts in kelp forests[END_REF][START_REF] Connell | The other ocean acidification problem: CO2 as a resource among competitors for ecosystem dominance[END_REF][START_REF] Wernberg | Climate-driven regime shift of a temperate marine ecosystem[END_REF].

Our results are in agreement with studies suggesting that OA might have a lesser direct effect on forest-forming macroalgae than warming (Kroeker et al., 2013a): the effect of the pH on the density of recruits was likely masked by the strong effect of the temperature. Nevertheless, the highest CHAPTER 1 ……….……………………..

35

densities were recorded under low temperature (28°C) and ambient pH (8.07), while the size was negatively affected by the high temperature (32°C), but positively affected by a decrease in pH (7.8). Complex interactions of abiotic and biotic factors are well known in natural systems. In our study we initially hypothesized that OW and OA would have affected the early-colonizer coralline algae Neogoniolithon brassica-florida, decreasing its potential facilitative effect on C. compressa recruitment. The calcification rate of the coralline algae was strongly affected by the temperature and pH, probably explaining the stronger effects of the temperature and pH on recruits growing in association with the coralline algae. However, the presence of both living and dead coralline algae had a negative effect on both the density and the size of the recruits, with the artificial clay substrate being the most favourable. Contrary to what is reported in the literature for other species [START_REF] Asnaghi | Colonisation processes and the role of coralline algae in rocky shore community dynamics[END_REF], the very common coralline alga N. brassica florida did not have a facilitating role in the recruitment of the later successional species C. compressa in our experiment. Since the very beginning of the experiment fewer C. compressa were observed in association with the coralline algae, living and dead. In the case of the living coralline algae substrates, the physiological state of N. brassica-florida likely does not explain this result as they exhibited calcification rates consistent with those reported in other species of coralline algae [START_REF] Martin | Response of Mediterranean coralline algae to ocean acidification and elevated temperature[END_REF]Cornwall et al., 2017b). The lower recruitment on living coralline could be due to an inhibition of the settlement and development of recruits of C. compressa caused by changes in pH or other chemical parameters in the boundary layer formed on the surface of the coralline algae [START_REF] Cornwall | Diffusion Boundary Layers Ameliorate the Negative Effects of Ocean Acidification on the Temperate Coralline Macroalga Arthrocardia corymbosa[END_REF](Cornwall et al., , 2017b)). Furthermore, crustose coralline algae have biotic interactions linked to their microbiome [START_REF] Gefen-Treves | The Microbiome Associated with the Reef Builder Neogoniolithon sp. in the Eastern Mediterranean[END_REF] and to their physical and chemical anti-fouling mechanisms to control epiphytes [START_REF] Johnson | The crustose coralline alga, Phymatolithon Foslie, inhibits the overgrowth of seaweeds without relying on herbivores[END_REF][START_REF] Keats | Antifouling effects of epithallial shedding in three crustose coralline algae (Rhodophyta, Coralinales) on a coral reef[END_REF][START_REF] Villas Bôas | Are anti-fouling effects in coralline algae species specific?[END_REF]. These characteristics of the surface of coralline algae could not be optimal for the recruitment of C. compressa and hence have reduced the settlement in our experiment. From an applicative point of view, it is worth noting that clay substrate, which is already used in many restoration actions in the Mediterranean Sea [START_REF] Falace | Ex situ cultivation protocol for Cystoseira amentacea var. stricta (Fucales, Phaeophyceae) from a restoration perspective[END_REF][START_REF] De La Fuente | First ex situ outplanting of the habitat-forming seaweed Cystoseira amentacea var. stricta from a restoration perspective[END_REF][START_REF] Orlando-Bonaca | First Restoration Experiment for Gongolaria barbata in Slovenian Coastal Waters[END_REF], was an adequate substrate for the settlement of C. compressa. Our results support that this substrate is of particular interest because it favours settlement and offers many technical practicalities (e.g. they are cheap, biodegradable, easy to produce and can be formed into any shape; [START_REF] Falace | Ex situ cultivation protocol for Cystoseira amentacea var. stricta (Fucales, Phaeophyceae) from a restoration perspective[END_REF][START_REF] De La Fuente | First ex situ outplanting of the habitat-forming seaweed Cystoseira amentacea var. stricta from a restoration perspective[END_REF][START_REF] Orlando-Bonaca | First Restoration Experiment for Gongolaria barbata in Slovenian Coastal Waters[END_REF].

In general, our results show that OW and OA additively, affect both the recruitment of C. compressa and the calcification rate of N. brassica-florida. Interestingly the interactive effect of OW and OA are likely exacerbated in presence of the coralline N. brassica-florida. This could be 1) because of the lower recruitment in the presence of coralline algae or 2) because the effects of OW and OA on coralline algae exacerbate, in turn, its inhibiting effect on C. compressa [START_REF] Maggi | Connell and Slatyer's models of succession in the biodiversity era[END_REF][START_REF] Bulleri | Facilitation and the niche: implications for coexistence, range shifts and ecosystem functioning[END_REF]. Our experiment shows that complex interactions of biotic and abiotic factors could CHAPTER 1 ……….…………………….. 36 affect the key species that shape marine forest communities, with an ultimate effect at the ecosystem level [START_REF] Smale | Climate-driven substitution of foundation species causes breakdown of a facilitation cascade with potential implications for higher trophic levels[END_REF]. As most experimental studies performed in controlled conditions, some limitations have to be highlighted. First of all, our experiment did not allow to separate the effects of the different drivers on the settlement process and the survival of recruits. Secondarily, epiphytes (turf algae) progressively appeared in our experimental tanks and their proliferation was enhanced at higher temperatures and low pH (author's personal observation), which in turn eventually affected the performance of C. compressa. But this phenomenon is likely to be observed also in natural conditions, where turf-forming species are expected to proliferate under OW and OA (Filbee-Dexter and [START_REF] Filbee-Dexter | Rise of Turfs: A New Battlefront for Globally Declining Kelp Forests[END_REF]. Many studies on recruits and adults of Cystoseira report elevated mortality under experimental conditions, which confirm the difficulty of maintaining Cystoseira in tanks [START_REF] Falace | Ex situ cultivation protocol for Cystoseira amentacea var. stricta (Fucales, Phaeophyceae) from a restoration perspective[END_REF][START_REF] De La Fuente | First ex situ outplanting of the habitat-forming seaweed Cystoseira amentacea var. stricta from a restoration perspective[END_REF][START_REF] De Caralt | Differential effects of pollution on adult and recruits of a canopy-forming alga: implications for population viability under low pollutant levels[END_REF] and of replicating the conditions of natural habitats in the laboratory. This could be due to different variability in light, temperature and flow conditions in the aquarium facilities [START_REF] Irving | Light, sediment, temperature, and the early life-history of the habitat-forming alga Cystoseira barbata[END_REF]. Different associated organisms (e.g. microbiome, epiphytic algae, invertebrates) could also explain the different performances observed when culturing these algae [START_REF] Mancuso | Diversity and Temporal Dynamics of the Epiphytic Bacterial Communities Associated with the Canopy-Forming Seaweed Cystoseira compressa (Esper) Gerloff and Nizamuddin[END_REF]. Our experiments were stopped when recruits showed signs of degradation, but the duration of our experiments is consistent with other studies on Cystoseira [START_REF] De Caralt | Differential effects of pollution on adult and recruits of a canopy-forming alga: implications for population viability under low pollutant levels[END_REF][START_REF] Falace | Is the South-Mediterranean Canopy-Forming Ericaria giacconei (= Cystoseira hyblaea) a Loser From Ocean Warming? Front[END_REF][START_REF] Verdura | Mediterranean macroalgal forests under threat: The effects of ongoing climate change and design of restoration methods[END_REF].

The densities obtained in our experiments are extremely high, reaching an average of 94 100.07 ± 89 324.78 C. compressa ind. m -2 (mean ± SD, n = 28) after two months in the first experiment and 14 213.20 ± 17.67 C. compressa ind. m -2 (mean ± SD, n = 197) after 2 months in the second experiment (min.: 1000 ind. m -2 ; max.: 320 000 ind. m -2 for the first experiment and min.: 10 000 ind. m -2 and max.: 210 000 ind. m -2 for the second). Interestingly, a parallel study performed by our team at the same time as the first experiment, seedling natural stones with the same technique presented in this study directly in the field, produced lower densities after 2 months. On the contrary, the recruits growing in the field under natural conditions were bigger (11.33 ± 3.27 mm) than in the first (5.18 ± 2.70 mm) and second experiment (2.51 ± 1.61 mm) after 2 months, showing that the conditions in tanks are not optimal for the growth of C. compressa and/or that densitydependent factors can result in high density and smaller size of recruits in the laboratory. Natural densities observed in the donor population reach only 76.36 ± 0.72 of C. compressa ind. m -2 (mean ± SD, n = 22; maximum: 128 ind. m -2 and minimum: 16 ind. m -2 ; author's personal observations, article in prep.). Even if the comparison with natural conditions often highlight some limitation for the studies in tanks, it is important to continue with this approach as it is the only one that allows testing multiple factors under controlled conditions (i.e. temperature and pH).

The results from this study demonstrate that the interactive effects of climate change have a pronounced negative impact on shallow marine forests. This result is especially striking for forest-CHAPTER 1 ……….…………………….. forming macroalgae thriving in rock pools, which are expected to be more acclimated to local factors variation (i.e. temperature and pH). Marine forests are facing several other stressors than global change (e.g. water pollution, urbanization, trampling, herbivory; [START_REF] Orfanidis | Effects of Natural and Anthropogenic Stressors on Fucalean Brown Seaweeds Across Different Spatial Scales in the Mediterranean Sea[END_REF]) that already put them on the edge. Some of these stressors have been addressed by management measures and are now mitigated (e.g. water quality; [START_REF] Blanfuné | The ups and downs of a canopy-forming seaweed over a span of more than one century[END_REF] allowing, in a few cases, the natural recovery of the forest or the feasibility of planning restoration actions [START_REF] Gorman | Recovering subtidal forests in human-dominated landscapes[END_REF][START_REF] Gianni | Conservation and restoration of marine forests in the Mediterranean Sea and the potential role of Marine Protected Areas[END_REF][START_REF] Cebrian | A Roadmap for the Restoration of Mediterranean Macroalgal Forests[END_REF][START_REF] Riquet | Highly restricted dispersal in habitat-forming seaweed may impede natural recovery of disturbed populations[END_REF]. Recently, new restoration techniques opened the door to the possibility of restoring these ecosystems, still, marine forests will be increasingly impacted by global change (IPCC, 2022) and it has to be considered to achieve long-term population sustainability and/or successful restoration actions. The next step for the protection of these key ecosystems is to understand how climate change and other drivers acting at the local scale can interact, eventually providing additive or synergetic effects (Halpern et al., 2008a), likely causing the restructure and redistribution of marine forests and affecting their ability to resist and recover under extreme conditions [START_REF] Smale | Impacts of ocean warming on kelp forest ecosystems[END_REF][START_REF] Verdura | Localscale climatic refugia offer sanctuary for a habitat-forming species during a marine heatwaves[END_REF][START_REF] Smale | Climate-driven substitution of foundation species causes breakdown of a facilitation cascade with potential implications for higher trophic levels[END_REF]. carbonate chemistry. S.C. and M.M. analysed the data. M.M. wrote the first draft of the manuscript which was then finalized by all co-authors.

CHAPTER 2: GRAZING EFFECTS ON RECRUITS OF THE FOREST-FORMING FUCOID Cystoseira compressa

INTRODUCTION

Marine forests of large brown macroalgae represented by the species from the orders Laminariales, Tylopteridales, Desmarestiales and Fucales form structurally complex habitats in cold and temperate regions around the world (Wernberg and Filbee-Dexter, 2019). These forests, commonly located on intertidal and subtidal rocky bottoms, provide important ecosystem functions, such as habitat, food and shelter to multiple species and are one of the most productive ecosystems in the world participating in the carbon sink [START_REF] Boudouresque | Where Seaweed Forests Meet Animal Forests: the Examples of Macroalgae in Coral Reefs and the Mediterranean Coralligenous Ecosystem[END_REF]Wernberg and Filbee-Dexter, 2019;[START_REF] Filbee-Dexter | Seaweed forests are carbon sinks that can mitigate CO2 emissions[END_REF][START_REF] Pessarrodona | Global seaweed productivity[END_REF]. Despite the important ecological functions marine forests provide, multiple anthropogenic stressors are pushing them to the edge (Filbee-Dexter and [START_REF] Filbee-Dexter | Rise of Turfs: A New Battlefront for Globally Declining Kelp Forests[END_REF][START_REF] Carnell | Reconstructing Historical Marine Populations Reveals Major Decline of a Kelp Forest Ecosystem in Australia[END_REF]Wernberg et al., 2019b). A worldwide decline of marine forests is occurring, mainly driven by the destruction of habitats, reduced water quality, global change and proliferation of herbivores (Wernberg et al., 2019b;[START_REF] Orfanidis | Effects of Natural and Anthropogenic Stressors on Fucalean Brown Seaweeds Across Different Spatial Scales in the Mediterranean Sea[END_REF][START_REF] Pessarrodona | Homogenization and miniaturization of habitat structure in temperate marine forests[END_REF]. In particular, overgrazing by sea urchins and herbivorous fish species is in some locations responsible for such declines [START_REF] Foster | Loss of predators and the collapse of southern California kelp forests (?): Alternatives, explanations and generalizations[END_REF][START_REF] Gianni | Threats to large brown algal forests in temperate seas: the overlooked role of native herbivorous fish[END_REF][START_REF] Sharma | Testing the efficacy of sea urchin exclusion methods for restoring kelp[END_REF][START_REF] Barrientos | Temperate Kelp Forest Collapse by Fish Herbivory: A Detailed Demographic Study[END_REF], shaping and regulating benthic macroalgal communities in a variety of natural rocky reef systems from temperate to tropical regions [START_REF] Scheibling | Destructive grazing, epiphytism, and disease: the dynamics of sea urchin -kelp interactions in Nova Scotia[END_REF][START_REF] Vanderklift | Intensity of herbivory on kelp by fish and sea urchins differs between inshore and offshore reefs[END_REF][START_REF] Vergés | Role of fish herbivory in structuring the vertical distribution of canopy algae Cystoseira spp. in the Mediterranean Sea[END_REF]. Although local stressors and regional variations can dominate marine forest dynamics (Smith et al., 2017, 22;[START_REF] Smale | Impacts of ocean warming on kelp forest ecosystems[END_REF][START_REF] Verdura | Localscale climatic refugia offer sanctuary for a habitat-forming species during a marine heatwaves[END_REF], many studies have shown how sea urchins, and native and invasive herbivorous fish, and even omnivorous fish, are a potential threat to macroalgal communities and are responsible in the formation of turfs or barren ground extensions [START_REF] Tegner | Sea urchin cavitation of giant kelp (Macrocystis pyrifera C. Agardh) holdfasts and its effects on kelp mortality across a large California forest[END_REF]Vergés et al., 2014b[START_REF] Vergés | Long-term empirical evidence of ocean warming leading to tropicalization of fish communities, increased herbivory, and loss of kelp[END_REF][START_REF] Papadakis | The neglected role of omnivore fish in the overgrazing of Mediterranean rocky reefs[END_REF]. Other smaller invertebrates associated to marine forests such as decapods, gastropods, amphipods and isopods can significantly participate in the grazing of different life stages of forest-forming species [START_REF] Arrontes | Effect of grazing by limpets on mid-shore species assemblages in northern Spain[END_REF][START_REF] Jonne | Seasonal Changes in situ Grazing of the Mesoherbivores Idotea baltica and Gammarus oceanicus on the Brown Algae Fucus vesiculosus and Pylaiella littoralis in the Central Gulf of Finland, Baltic Sea[END_REF][START_REF] Gunnarsson | The brown alga Fucus radicans suffers heavy grazing by the isopod Idotea baltica[END_REF][START_REF] Hong | Effects of sea urchin and herbivorous gastropod removal, coupled with transplantation, on seaweed forest restoration[END_REF][START_REF] Navarro-Barranco | Amphipod assemblages associated with native habitat-forming seaweeds of the Alboran Sea: Influence of environmental protection and biogeographical patterns[END_REF][START_REF] Molis | Grazing Impact of, and Indirect Interactions Between Mesograzers Associated with Kelp (laminaria Digitata)1[END_REF] but little is known about the magnitude of their possible effects.

In the Mediterranean Sea, density and abundance of macroalgae are also controlled by herbivorous [START_REF] Vergés | Role of fish herbivory in structuring the vertical distribution of canopy algae Cystoseira spp. in the Mediterranean Sea[END_REF]. The two species of sea urchin Paracentrotus lividus and Arbacia lixula are the most common benthic macro-herbivores of the sublittoral rocky bottoms [START_REF] Bulleri | Grazing by the sea urchins Arbacia lixula L. and Paracentrotus lividus Lam. in the Northwest Mediterranean[END_REF][START_REF] Agnetta | Role of two co-occurring Mediterranean sea urchins in the formation of barren from Cystoseira canopy[END_REF]. Both species of sea urchin have been responsible alongside anthropogenic stressors (e.g. date mussel harvesting), for the degradation and regime shifts of forest-forming and shrub-forming macroalgae assemblages to barren grounds, favouring the maintenance of postregime shifts stable states (e.g. bare rock) in many areas [START_REF] Sala | Fishing, Trophic Cascades, and the Structure of Algal Assemblages: Evaluation of an Old but Untested Paradigm[END_REF][START_REF] Bulleri | Grazing by the sea urchins Arbacia lixula L. and Paracentrotus lividus Lam. in the Northwest Mediterranean[END_REF][START_REF] Guidetti | Relationships among predatory fish, sea urchins and barrens in Mediterranean rocky reefs across a latitudinal gradient[END_REF][START_REF] Guidetti | The destructive date-mussel fishery and the persistence of barrens in Mediterranean rocky reefs[END_REF][START_REF] Giakoumi | Relationships between fish, sea urchins and macroalgae: The structure of shallow rocky sublittoral communities in the Cyclades, Eastern Mediterranean[END_REF].

However, sea urchins are not the only macro-herbivores that play a role in the structure of macroalgal assemblages of the Mediterranean Sea. Salema fish, (Sarpa salpa), the only true native herbivorous fish in the Mediterranean Sea, is also important in structuring macroalgae communities, and can also be responsible for the depletion of Cystoseira s.l. forests [START_REF] Gianni | Threats to large brown algal forests in temperate seas: the overlooked role of native herbivorous fish[END_REF]. More recently, the range-expansion of tropical herbivorous fishes such as the rabbitfish (Siganus spp.) has also been signalled for severely reducing the biomass and diversity of macroalgal species in the Mediterranean Sea, driving regime shifts from macroalgal dominated communities to barrens (Vergés et al., 2014b[START_REF] Vergés | Long-term empirical evidence of ocean warming leading to tropicalization of fish communities, increased herbivory, and loss of kelp[END_REF][START_REF] Zarco-Perello | Range-extending tropical herbivores increase diversity, intensity and extent of herbivory functions in temperate marine ecosystems[END_REF]. Herbivorous range-shifting fishes have also been reported for preferring to graze on turf, constituting also a menace for forest-forming species as they could graze on the microscopic life-stages present in the turf matrix [START_REF] Barrientos | Feeding preferences of range-shifting and native herbivorous fishes in temperate ecosystems[END_REF].

Cystoseira s.l. spp. in the Mediterranean Sea form prominent and dense canopies that are analogous to kelp forests of other temperate rocky coasts [START_REF] Giaccone | Écologie et chorologie des Cystoseira de Méditerranée[END_REF][START_REF] Ballesteros | Community structure and frond size distribution of a deep water stand of Cystoseira spinosa (Phaeophyta) in the Northwestern Mediterranean[END_REF], and are amongst the most productive and complex habitats of the Mediterranean Sea (Ballesteros, 1989a(Ballesteros, , 1990a;;[START_REF] Clayton | The adaptive significance of life history characters in selectedorders of marine brown maeroalgae[END_REF]. The vertical zonation of Cystoseira s.l. spp. depends on multiple depthdependent physical factors such as light, hydrodynamics, temperature, and availability of nutrients [START_REF] Ballesteros | Surface-dependent strategies and energy flux in benthic marine communities or, why corals do not exist in the Mediterranean[END_REF], but also on biotic factors such as herbivory [START_REF] Ruitton | Relationships between Algae, Benthic Herbivorous Invertebrates and Fishes in Rocky Sublittoral Communities of a Temperate Sea (Mediterranean)[END_REF][START_REF] Vergés | Role of fish herbivory in structuring the vertical distribution of canopy algae Cystoseira spp. in the Mediterranean Sea[END_REF][START_REF] Sala | Alien Marine Fishes Deplete Algal Biomass in the Eastern Mediterranean[END_REF]. Because of the important ecosystem services marine forests provide, important efforts have recently been made to promote the conservation and the viability of their restoration [START_REF] Eger | Global Kelp Forest Restoration: Past lessons, status, and future goals[END_REF].

Different restoration techniques have been used to restore marine forests from transplantation of adults to ex-situ and in-situ recruitment enhancement techniques [START_REF] Verdura | Restoration of a Canopy-Forming Alga Based on Recruitment Enhancement: Methods and Long-Term Success Assessment[END_REF]Cebrian et al., 2021;[START_REF] Falace | Ex situ cultivation protocol for Cystoseira amentacea var. stricta (Fucales, Phaeophyceae) from a restoration perspective[END_REF][START_REF] De La Fuente | First ex situ outplanting of the habitat-forming seaweed Cystoseira amentacea var. stricta from a restoration perspective[END_REF][START_REF] Savonitto | Addressing reproductive stochasticity and grazing impacts in the restoration of a canopyforming brown alga by implementing mitigation solutions[END_REF]. However, still, some problems need to be overcome (e.g. herbivory and climate change) to ensure the success of conservation and restoration actions [START_REF] Gianni | Conservation and restoration of marine forests in the Mediterranean Sea and the potential role of Marine Protected Areas[END_REF][START_REF] Cebrian | A Roadmap for the Restoration of Mediterranean Macroalgal Forests[END_REF][START_REF] Savonitto | Addressing reproductive stochasticity and grazing impacts in the restoration of a canopyforming brown alga by implementing mitigation solutions[END_REF]. Because grazing is believed to be one of the most important biological factors controlling the survival of early-life stages of fucoids [START_REF] Chapman | Functional ecology of fucoid algae: twenty-three years of progress[END_REF], it constitutes one of the major problems when planning the restoration of marine forests [START_REF] Gianni | Conservation and restoration of marine forests in the Mediterranean Sea and the potential role of Marine Protected Areas[END_REF][START_REF] Gianni | Reduction of herbivorous fish pressure can facilitate focal algal species forestation on artificial structures[END_REF][START_REF] Tamburello | Enhanced nutrient loading and herbivory do not depress the resilience of subtidal canopy forests in Mediterranean oligotrophic waters[END_REF][START_REF] Savonitto | Addressing reproductive stochasticity and grazing impacts in the restoration of a canopyforming brown alga by implementing mitigation solutions[END_REF]. Knowing which species graze on different life stages of forest-forming species could be crucial for successful conservation and restoration actions. It is of special interest to study early-life stages as they are more vulnerable than adults and high mortality rates are naturally observed [START_REF] Aberg | A Demographic Study of Two Populations of the Seaweed Ascophyllum Nodosum[END_REF][START_REF] Coelho | Impacts of anthropogenic stresses on the early development stages of seaweeds[END_REF][START_REF] Schiel | The Population Biology of Large Brown Seaweeds: Ecological Consequences of Multiphase Life Histories in Dynamic Coastal Environments[END_REF], and because the resilience of the whole population to future impacts can be largely dependent on efficient recruitment and growth of the recruits [START_REF] Ang | Natural dynamics of a Fucus distichus (Phaeophyceae, Fucales) population: reproduction and recruitment[END_REF][START_REF] Capdevila | Recruitment patterns in the Mediterranean deep-water alga Cystoseira zosteroides[END_REF].

The main objective of this study was to investigate the herbivory pressure in the field and the effect of different species (sea urchins, fishes, decapods, gastropods, amphipods and isopods) on recruits of Cystoseira compressa. In order to assess species-specific herbivory pressure on the survival and growth of recruits of C. compressa we performed experiments in the field and in mesocosms. More specifically, in the field we performed herbivory exclusion experiments on recruits of C. compressa obtained from both in-situ and ex-situ recruitment enhancement techniques. In laboratory, we assessed for the first time the potential grazing impact of non-strict herbivorous species that have not been considered before as potential herbivores (decapods, gastropods, amphipods and isopods) on recruits of C. compressa.

MATERIAL AND METHODS

Study sites and target species

Cystoseira compressa was the selected species because it is considered one of the most resistant species to manipulation and the only Cystoseira s.l. spp. not protected under Barcelona Convention Annex II and United Nations Environment Programme/Mediterranean Action Plan-UNEP/MAP [START_REF] Mangialajo | Zonation patterns and interspecific relationships of fucoids in microtidal environments[END_REF][START_REF] Verlaque | Mediterranean seaweeds listed as threatened under the Barcelona Convention: A critical analysis[END_REF]. This species, that could also have a role in facilitating the settlement of other Cystoseira s.l. spp. [START_REF] Mangialajo | Zonation patterns and interspecific relationships of fucoids in microtidal environments[END_REF], is still present in the French Riviera, where patches can be found along exposed rocky shores but dense populations are only found on rockpools in Lérins Islands (author's personal information; [START_REF] Thibaut | Decline and local extinction of Fucales in French Riviera: the harbinger of future extinctions?[END_REF]. CHAPTER 2 …………………………….. 44 Three sites were selected to study the herbivory pressure on recruits of C. compressa: one site on a Cystoseira s.l. forest (Sainte Marguerite Island) and two sites where C. compressa was reported in the past but where at present is lost (Beaulieu-sur-Mer and Passable; [START_REF] Thibaut | Decline and local extinction of Fucales in French Riviera: the harbinger of future extinctions?[END_REF]. The forest site corresponds to a rockpool system located in Sainte Marguerite Island (from now Sainte Marguerite; WN-Mediterranean Sea -Lérins Islands, Cannes, France), a protected Natura 2000 site at 1.3 km from the coast of Cannes where there is one of the last well-conserved shallow marine forests in the Côte d'Azur (Figure 1). The rockpool system (between 0.

Herbivores survey

Field experiment 1: Herbivory on in-situ recruitment substrates in a rockpool

In order to study the effectiveness of the in-situ recruitment enhancement and the herbivory pressure on recruits in the field, an herbivory exclusion experiment was performed in the rockpool system in Sainte Marguerite (Figure 1). 18 substrates were randomly fixed using Epoxy to the bottom of the rockpool system, to provide free substrate. In order to test the effect of recruitment enhancement, half of these substrates were seeded using the non-destructive in-situ recruitment enhancement technique [START_REF] Verdura | Restoration of a Canopy-Forming Alga Based on Recruitment Enhancement: Methods and Long-Term Success Assessment[END_REF]. To do so, on July 2020, after having observed mature conceptacles under the microscope, apical fertile branches (ca. 3 cm in length) were manually collected and transported in cold and dark conditions in plastic bags to the laboratory. The receptacles were conserved at 4°C in the dark overnight before placing them on the selected substrates in-situ. On each substrate, a net bag containing 5 g fresh weight (FW) of receptacles was placed floating above the substrate for three days. In order to assess the herbivory impact on the recruits, again half of the substrates were protected (factor Protection, three levels) to avoid herbivory (mainly from fishes and sea urchins, factor level protected), artefact controls and substrates with no protection (levels artefact control and open, respectively) were also included (Figure 2; Recruitment enhancement × Herbivory Protection, n = 3). The density (as number of recruits per surface) and size (cm) as length of the longest axis of each C. compressa individual growing on the substrates were monitored at 2, 9 and 12 months and used as variables. The size of the recruits was calculated by measuring 10 individuals from each substrate, in the case of substrates with less than 10 individuals, all the individuals were measured. The natural density of the C. compressa individuals present in the rockpool of Sainte Marguerite was monthly monitored from May to October 2020 using 20*20 cm quadrats (n = 25) and the global mean value was used as a reference to assess the evolution during our experiments.

The density of recruits of C. compressa on 0.04 m 2 was used as response variable. A Generalized Linear Model (GLM), with a Quasipoisson link log distribution function, was used to test the effect of the herbivory protection and recruitment enhancement on recruits of C. compressa after 2 months, with herbivory protection (three levels) and recruitment enhancement (two levels) as fixed factors. After testing the effect of the recruitment enhancement, this factor was no longer considered CHAPTER 2 …………………………….. for further analysis. Then, a Generalized Linear Mixed-Effects Model (GLMM) with a Poisson link log distribution function was used to test the effect of the factor protection over time with the factors protection and time (both three levels) as fixed factors and the factor substrate as random factor, as GLMM can cope with repeated measures over time (Pinheiro and Bates, 2000). A GLMM with an inverse gaussian link log distribution was used to test the effects of the herbivory protection on the size of the recruits (cm), with the factors herbivory protection and time (both three levels) as fixed factors and the substrate as a random factor.

Field experiment 2: Herbivory on ex-situ recruitment substrates on the open coast

In order to study the herbivory pressure in the field and the success of the ex-situ recruitment enhancement technique in absence of a Cystoseira s.l. spp. forest, an herbivory exclusion experiment was performed in the two locations situated in the open coast: Beaulieu and Passable (Figure 1). In August 2021, after having observed mature conceptacles under the microscope, about 180 g FW apical branches of C. compressa containing fertile receptacles, were collected from the same C. compressa donor population located in the rockpool system in Sainte Marguerite, following the same non-destructive technique described before (see section 2.3;[START_REF] Falace | Ex situ cultivation protocol for Cystoseira amentacea var. stricta (Fucales, Phaeophyceae) from a restoration perspective[END_REF][START_REF] Verdura | Restoration of a Canopy-Forming Alga Based on Recruitment Enhancement: Methods and Long-Term Success Assessment[END_REF][START_REF] De La Fuente | First ex situ outplanting of the habitat-forming seaweed Cystoseira amentacea var. stricta from a restoration perspective[END_REF]. The apical branches were transported in cold and dark conditions and once in the laboratory they were gently cleaned with tweezers. To stimulate the release of gametes, the apical branches were maintained at 4°C in the dark overnight before placing them on the surface of three 30 L closed system tanks (60 g FW of fertile receptacles per tanks) to obtain recruits on substrates placed on the bottom of the tanks. The receptacles were kept for four days on the surface of the tanks while releasing the zygotes. The recruits of C. compressa were kept for 1 month in the aquarium facilities, with air pumps and under natural light conditions, before the transplant to the two coastal locations was done. The seawater (filtered using a 200 µm mesh) of the tanks was changed every two days and no culture medium was provided to the cultures. After 2.5 months in the tanks, 24 substrates with similar densities of C. compressa recruits were transplanted and fixed using Epoxy at the sea bottom of the two open coast locations in October 2021. Some substrates with recruits were maintained in the aquarium facilities and used as controls (n = 6). As for experiment 1, half of the transplanted substrates were protected to avoid herbivory (mainly fishes and sea urchins), while the others were not (n=6; Figure 2).

The density of recruits of C. compressa on 0.04 m 2 was used as response variable. A Generalized Linear Mixed-Effect Model (GLMM) with a Poisson link log distribution function, was used to test the effect of the herbivory protection on recruits of C. compressa, with the herbivory protection (two levels) and time (two levels) as fixed factors, and the site (two levels) and substrate as random to cope with repeated measures over time (Pinheiro and Bates, 2000).

Mesocosm experiment: Potential herbivory of different species

Four experiments were done in a mesocosm in the laboratory to study the potential herbivory role of some invertebrates on recruits of C. compressa. The selected species consisted of a decapod (Clibanarius erythropus), a gastropod (Cerithium vulgatum), an amphipod (Gammarus sp.) and an isopod (Idotea balthica). The species and their densities were selected in function of the observations in the field and in function of the information reported in the literature for other herbivory experiments performed on fucoid species [START_REF] Engkvist | Density dependent grazing effects of the isopod Idotea baltica Pallas on Fucus vesiculosus L in the Baltic Sea[END_REF][START_REF] Gunnarsson | The brown alga Fucus radicans suffers heavy grazing by the isopod Idotea baltica[END_REF][START_REF] Suzuki | Effects of herbivorous gastropod grazing on the sedimentation and succession of subtidal macroalgal assemblages[END_REF][START_REF] Hong | Effects of sea urchin and herbivorous gastropod removal, coupled with transplantation, on seaweed forest restoration[END_REF][START_REF] Navarro-Barranco | Amphipod assemblages associated with native habitat-forming seaweeds of the Alboran Sea: Influence of environmental protection and biogeographical patterns[END_REF]. All the GLMM and LM models were fitted to analyse the effect of the variables and the AICs likelihood minimum was used to select the best model among the possible combinations. All the different models were fitted using the functions "glmer" and "lm" from the package lme4 [START_REF] Bates | Fitting Linear Mixed-Effects Models using lme4[END_REF] in the statistical environment R (R Core Team, 2019). For all the models, the assumptions of normality and equality of variance were evaluated through graphical analyses of residuals using QQ plot functions. P-values were obtained by means of a Wald χ2 test using the 'ANOVA' function from the car package [START_REF] Fox | An R Companion to Applied Regression[END_REF]. Finally, the function 'emmeans' from the package emmeans [START_REF] Lenth | emmeans: Estimated Marginal Means, aka Least-Squares Means[END_REF] was used to perform the post-hoc analysis.

RESULTS

Herbivores survey

In the open coast sites mean densities of Sarpa salpa of 1.1 ± 1.59 ind. 39.27 m -2 and of 1.2 ± 0.84 ind. 39.27 m -2 (mean ± SD, hereafter) were respectively found in Beualieu and Passable. Sea urchins (Paracentrotus lividus and Arbacia lixula) were present at densities of 6.00 ± 6.86 ind. m -2 and of 3.14 ± 3.89 ind. m -2 ; Paguroidea decapods were present at densities of 2.91 ± 6.32 ind. m -2 and of 49.45 ± 69.29 ind. m -2 ; and Cerithium vulgatum was present at densities of 5.09 ± 18.07 ind.m -2 and of 96.73 ± 116.27 ind. m -2 , respectively for Beaulieu and Passable (Figure 3). In the rockpool system in Sainte Marguerite, neither Sarpa salpa (>5 cm) or sea urchins, Paracentrotus lividus or Arbacia lixula, were observed during the experiment. In this location mean density of Paguroidea decapods was 45.90 ± 102.18 ind. m -2 and of Cerithium vulgatum was 36.46 ± 73.73 ind. m -2 (mean ± SD). Paguroidea decapods in the three locations consisted mainly in Clibanarius erythropus but individuals of Diogenes pugilator and few of Calcinus tubularis were also observed. In general, higher densities of herbivores were observed in Passable (Figure 3).

Field experiment 1: Herbivory on in-situ recruitment substrates in a rockpool

The density of recruits (ind. 0.04 m -2 ) on the substrates in Sainte Marguerite, were affected at two months only by the herbivory protection, and not the recruitment enhancement (GLMM, P-value < 0.001; Figure 4 and Table 1). At this time, the post-hoc analysis found higher densities of recruits on protected substrates than on open or artefact control substrates. Due to the non-significance of the factor recruitment enhancement on the density of recruits at two months, this factor was no longer considered and the data was pooled together in the next analysis. When considering the three sampling dates, the herbivory protection and the time exerted an interactive effect on the density of recruits (GLMM, P-value < 0.001 for the interaction Herbivory protection × Time; Figure 5 and Table 1). The post-hoc analysis detected differences in densities of C. compressa between the levels of herbivory protection for all sampling days. At month 2 and 12 densities of C. compressa were higher on protected substrates, while at month 7 there were no differences between protected and open substrates. For all the herbivory protection levels, the densities of C. compressa decreased over time, but at month 12 they were still higher than the ones observed in the natural population (3.05 ± 0.14 ind. C. compressa 0.04 m -2 on natural populations; Figure 4). Only time, and not the herbivory protection had a significant effect on the size of the recruits (GLMM, P-value < 0.001 for the factor Time; Figure 6 and Table 1). The size was increasing during the sampling dates with a maximum at month 12 (July 2021), with a mean size of 9 ± 4.18 cm (mean ± SD), and maximum and minimum of 18 and 3 cm, respectively (Figure 6 and Table 1).

The natural population density of Cystoseira compressa in Sainte Marguerite was 3.05 ± 0.14 ind.

CHAPTER 2 …………………………….. 52 0.04 m -2 (76.36 ± 0.72 individuals m -2 ; mean ± SD), with maximum densities of 5 and a minimum of 1 individual 0.04 m -2 .

Field experiment 2: Herbivory on ex-situ recruitment substrates on the open coast

The herbivory protection and time had an interactive effect on the density of recruits of C.

compressa on the open coast (GLMM, P-value < 0.001 for the interaction Herbivory protection × Time; Figure 7 and Table 1). The post-hoc analysis found higher densities of recruits of C. compressa on protected substrates after 1 and 12 days. However most of the recruits (> 95%), even on protected substrates, disappeared after 24h, while densities of the recruits that remained in the tanks in the laboratory (substrates that were not transplanted to the field) remained stable after 12 days (Figure 7). on the two open coast sites and in the laboratory controls. The density on the laboratory control substrates was lower as the most recruited substrates were selected to be transplanted to the field. The errors bars show the standard error.

Mesocosm experiment: Potential herbivory of different species

All the studied herbivorous species, except Gammarus sp., consumed recruits of C. compressa. For

Clibanarius erythropus, Cerithium vulgatum and Idotea balthica the post-hoc analysis found significant differences in the consumption of C. compressa biomass when compared to the controls (factor number of individuals; GLMM, P-value < 0.001; Figure 8 and Table 1). No consumption and neither significant mortality of C. compressa was found in the tanks with Gammarus sp. (Figure 8).

For C. erythropus, the only differences were between the control and the presence of the herbivores, whatever the density was (without significant differences according to the number of individuals).

While for C. vulgatum there were significant differences between the highest and the lowest number of individuals (5 individuals versus 1 individual; Figure 8 and Table 1). 24h after the start of the experiment, I. balthica individuals consumed 100% of the biomass of recruits available in four of the seven experimental tanks, and more than the 85% in two of the seven experimental tanks (Figure 8).

DISCUSSION

Our study is a first step to elucidate the herbivory pressure in the field and direct effects of different species on the grazing of recruits in the framework of shallow Cystoseira s.l. restoration, as grazing have been observed and quantified on transplanted adult individuals and recruits [START_REF] Susini | Development of a transplantation technique of Cystoseira amentacea var. stricta and Cystoseira compressa[END_REF]; 0.00 0.04 0.08 0.12 Clibanarius erythropus [START_REF] Mangialajo | Zonation patterns and interspecific relationships of fucoids in microtidal environments[END_REF][START_REF] Tamburello | Enhanced nutrient loading and herbivory do not depress the resilience of subtidal canopy forests in Mediterranean oligotrophic waters[END_REF][START_REF] Savonitto | Addressing reproductive stochasticity and grazing impacts in the restoration of a canopyforming brown alga by implementing mitigation solutions[END_REF]. During the experiments Cerithium vulgatum and decapods (mainly Clibanarius erythropus) were the most abundant herbivorous observed on all our sampling sites in exception of Beaulieu, and higher densities were observed on patches of bare rock just next to the macroalgal communities (author's personal observation). In the rockpool system in Sainte Marguerite the density of Cystoseira compressa recruits changed in function of herbivory protection, showing how important it is to control herbivory on early-life stages for increasing restoration success. The first assessment of density and sizes of recruits on this location was performed 2 months after the substrates provision and recruitment enhancement because of the difficulty of identifying the early-life stages. Even if in this location the densities of recruits drastically decreased on month 7, on month 12 there were still significantly more recruits on the protected substrates than on the artefact control and open substrates. The variations on the density of recruits could be because of seasonal changes or also due to density-dependant effects: the density decreases as the size increases [START_REF] Chapman | Functional ecology of fucoid algae: twenty-three years of progress[END_REF].

Nevertheless, on all the substrates from the three treatments (protected, artefact control, and open) the density of recruits of C. compressa after 12 months was still higher than the density observed in the surrounding natural population. On the experiment in the field we did not observe an effect of the factor recruitment enhancement, probably due to the presence of fertile C. compressa adults in the rockpool, which could have recruit on all our substrates. Also, the fact of not observing an effect of the recruitment enhancement factor could be because the recruitment enhancement performed on half of our substrates had a larger scale effect, affecting all the substrates and not only the substrate just underneath. In addition, there were no differences on the size of the recruits independently of the herbivory protection as the size of the recruits increased over time in all the substrates. At month 12, a slightly higher number of recruits was reported than at month 7, those new recruits were probably settled later during the season.

In the two open coast locations, Beaulieu and Passable, the dramatic decrease in density of the transplanted recruits after only 24 h could be due to several reasons. Unlike in Sainte Marguerite, where the conditions for the growth of Cystoseira s.l. spp. were ensured (presence of a healthy forest), the environmental conditions for the survival of recruits of C. compressa in the two open coast locations might have not been adequate (e.g. water quality, hydrodynamics, irradiation, absence of a canopy… [START_REF] Irving | Light, sediment, temperature, and the early life-history of the habitat-forming alga Cystoseira barbata[END_REF]. Another reason for this ex-situ technique to fail in these locations could be the methodology used for obtaining recruits: the recruits that were adapted to the stable conditions in the tanks for 2.5 months might have been sensitive to different environmental parameters such as hydrodynamics, variations in temperature, higher irradiance, different water chemistry etc. This problem could be solved by transplanting the recruits sooner to the sea, as suggested in other studies [START_REF] Falace | Ex situ cultivation protocol for Cystoseira amentacea var. stricta (Fucales, Phaeophyceae) from a restoration perspective[END_REF][START_REF] De La Fuente | First ex situ outplanting of the habitat-forming seaweed Cystoseira amentacea var. stricta from a restoration perspective[END_REF][START_REF] Savonitto | Addressing reproductive stochasticity and grazing impacts in the restoration of a canopyforming brown alga by implementing mitigation solutions[END_REF].

Replicating natural conditions in experimental tanks is difficult, and during the 2.5 month that the CHAPTER 2 …………………………….. 56 recruits were in the laboratory we could observe that they grew less sizes than the recruits in Sainte Marguerite, obtained from the in-situ recruitment enhancement technique (author's personal observation). Even if it is not considered a major stressor for the recruits [START_REF] Falace | Ex situ cultivation protocol for Cystoseira amentacea var. stricta (Fucales, Phaeophyceae) from a restoration perspective[END_REF][START_REF] De La Fuente | First ex situ outplanting of the habitat-forming seaweed Cystoseira amentacea var. stricta from a restoration perspective[END_REF], the transport of the recruits from the laboratory facilities to the experimental locations on the coast could have affected the recruits. However, the fact of not observing mortality of recruits on the substrates that were kept in the laboratory (substrates that were not transplanted in Beaulieu and Passable and were kept as laboratory controls), could be due to the lower densities of recruits they had in comparison to the transplanted substrates in the field. As said before, substrates with higher densities of recruits could have experiences mortality due to densitydependant effects, known as self-thinning [START_REF] Chapman | Functional ecology of fucoid algae: twenty-three years of progress[END_REF].

The recruits for the two field experiments were obtained using different techniques in function of the characteristic of each location [START_REF] Cebrian | A Roadmap for the Restoration of Mediterranean Macroalgal Forests[END_REF] which prevent from comparing the results obtained between Sainte Marguerite, and Beaulieu and Passable. In the rockpool system in Sainte Marguerite, the in-situ technique with substrate provision and natural and artificial seedling was used, as it is the one preferred for sheltered locations with low herbivory pressure [START_REF] Cebrian | A Roadmap for the Restoration of Mediterranean Macroalgal Forests[END_REF]. While in the open coast in Beaulieu and Passable, the ex-situ technique was used as it is the one recommended for deforested locations [START_REF] Cebrian | A Roadmap for the Restoration of Mediterranean Macroalgal Forests[END_REF]. However, herbivory pressure change in function of the physical environmental conditions and the structure of the benthic communities [START_REF] Lubchenco | Relative importance of competition and predation: early colonization by seaweeds in New England[END_REF]. One study [START_REF] Gianni | Reduction of herbivorous fish pressure can facilitate focal algal species forestation on artificial structures[END_REF] performed on the open infralittoral fringe of the French Riviera (N-W Mediterranean Sea), where sea urchins were not present, concluded that S. salpa was the most efficient grazer on adult Cystoseira amentacea and able to limit the success of restoration actions. However, [START_REF] Ferrario | The overlooked role of biotic factors in controlling the ecological performance of artificial marine habitats[END_REF] The mesocosm experiment performed in this study is a first approach for elucidating which other species, apart from the most studied ones (mainly urchins and herbivorous fish), can have a role in the grazing of recruits of Cystoseira s.l. spp. [START_REF] Ferrario | The overlooked role of biotic factors in controlling the ecological performance of artificial marine habitats[END_REF] already highlighted that most species consuming or interacting with Cystoseira s.l. spp. are usually classified as omnivorous rather than herbivorous. During our experiment, the species that had the highest and fastest consumption of recruits of C. compressa was Idotea balthica, followed by Clibanarius erythropus and Cerithium vulgatum. Even if in our experiment the amphipod (Gammarus sp.) did not graze on CHAPTER 2 …….….……………..…..… 57 recruits of C. compressa, it is reported that amphipods heavily feed on brown macroalgae [START_REF] Duffy | Global kelp forest restoration: past lessons, present status, and future directions[END_REF] including Fucales [START_REF] Jonne | Seasonal Changes in situ Grazing of the Mesoherbivores Idotea baltica and Gammarus oceanicus on the Brown Algae Fucus vesiculosus and Pylaiella littoralis in the Central Gulf of Finland, Baltic Sea[END_REF], but they preferences could be species specific. Some studies have already shown how molluscs, and decapods can graze or interact (i.e. clipping and cutting the thalli) with recruits and adults of some forest-forming macroalgae including C. compressa [START_REF] Lubchenco | Littornia and Fucus: Effects of Herbivores, Substratum Heterogeneity, and Plant Escapes During Succession[END_REF][START_REF] Perkol-Finkel | Conservation challenges in urban seascapes: promoting the growth of threatened species on coastal infrastructures[END_REF][START_REF] Ferrario | The overlooked role of biotic factors in controlling the ecological performance of artificial marine habitats[END_REF][START_REF] Gianni | Threats to large brown algal forests in temperate seas: the overlooked role of native herbivorous fish[END_REF]. I. balthica is the most widespread Idoteidae in European Seas [START_REF] Guarino | BIOLOGY AND POPULATION DYNAMICS OF IDOTEA BALTICA (CRUSTACEA, ISOPODA) IN THE GULF OF NAPLES, THE TYRRHENIAN SEA[END_REF] that is frequently used as model species to study the interactions between macroalgae and herbivores [START_REF] Gutow | Ocean acidification affects growth but not nutritional quality of the seaweed Fucus vesiculosus (Phaeophyceae, Fucales)[END_REF][START_REF] Lavaut | Pollinators of the sea: A discovery of animal-mediated fertilization in seaweed[END_REF]. This species can be found living in macroalgal communities and marine forests [START_REF] Guarino | BIOLOGY AND POPULATION DYNAMICS OF IDOTEA BALTICA (CRUSTACEA, ISOPODA) IN THE GULF OF NAPLES, THE TYRRHENIAN SEA[END_REF][START_REF] Lavaut | Pollinators of the sea: A discovery of animal-mediated fertilization in seaweed[END_REF] and it is an important consumer of forest-forming brown macroalgae such as the shallow Fucus vesiculosus in the Atlantic [START_REF] Kotta | Major Changes in Macroalgae Community Composition Affectthe Food and Habitat Preference of Idotea baltica[END_REF][START_REF] Jonne | Seasonal Changes in situ Grazing of the Mesoherbivores Idotea baltica and Gammarus oceanicus on the Brown Algae Fucus vesiculosus and Pylaiella littoralis in the Central Gulf of Finland, Baltic Sea[END_REF][START_REF] Vesakoski | Sexual and local divergence in host exploitation in the marine herbivore Idotea baltica (Isopoda)[END_REF][START_REF] Molis | Grazing Impact of, and Indirect Interactions Between Mesograzers Associated with Kelp (laminaria Digitata)1[END_REF][START_REF] Schaal | Biodiversity and trophic structure of invertebrate assemblages associated with understorey red algae in a Laminaria digitata bed[END_REF]. I. balthica has an important grazing effect on structuring F. vesiculosus populations in the Baltic Sea, preferring younger tissue over older [START_REF] Engkvist | Density dependent grazing effects of the isopod Idotea baltica Pallas on Fucus vesiculosus L in the Baltic Sea[END_REF][START_REF] Boström | Effects of isopod grazing: an experimental comparison in temperate (Idotea balthica, Baltic Sea, Finland) and subtropical (Erichsonella attenuata, Gulf of Mexico, U.S.A.) ecosystems[END_REF]. It could also play an important role in grazing recruits of shallow Cystoseria s.l. spp. in the Mediterranean Sea according to our results. Even if macroalgae species, including Cystoseira s.l. spp., contain metabolites such as phenolic compounds, that deter feeding in macroalgae and especially on young tissue, this chemical defence may not be enough to prevent grazing by some herbivorous species. For example, in another mesocosm experiment [START_REF] Vergés | Experimental evidence of chemical deterrence against multiple herbivores in the seagrass Posidonia oceanica[END_REF] C. vulgatum was the only invertebrate unaffected by the chemical deterrence extracts from the phanerogam Posidonia oceanica, and it was one of the species with the highest densities in our study location. It is however important to note that the lack of other potentially more palatable species of macroalgae or the limited amount of biomass of C. compressa in each tank could have impacted the consumptions rates of C. compressa reported here during the mesocosm experiment.

We highlight that non-strict herbivorous species can potentially graze on recruits of Cystoseira s.l.

spp. affecting the success of marine forests conservation and restoration actions. The protected substrates in the studied locations had significantly more densities of recruits of C. compressa, showing the negative effect of herbivory on early-life stages obtained from different recruitment techniques. However, we cannot exclude that others characteristics of the locations had an effect on the density of recruits. Despite adapting the restoration techniques to the particularities of each locations, the degree of success of the restoration is not assured. We conclude that, in addition to making sure that abiotic conditions (e.g. water quality, seawater temperature, anthropogenic pressures) are favourable for the restoration of marine forest, biotic factors such as herbivory should be well understood as a great variety of species could impact the recruits (e.g. in function of the size of the recruits). In locations with high herbivory pressure, anti-herbivory devices should be included when restoring marine forests [START_REF] Cebrian | A Roadmap for the Restoration of Mediterranean Macroalgal Forests[END_REF] and these devices should be adequate CHAPTER 2 …………………………….. 58 to the diversity of potential herbivorous species present in the site. More studies are needed to predict how the feeding behaviour and preferences of different native and invasive species could change under climate change to evaluate the threat they pose for marine forests conservation in the future [START_REF] Tomas | Differential herbivory of invasive algae by native fish in the Mediterranean Sea[END_REF][START_REF] Asnaghi | Cascading Effects of Ocean Acidification in a Rocky Subtidal Community[END_REF][START_REF] Gutow | Ocean acidification affects growth but not nutritional quality of the seaweed Fucus vesiculosus (Phaeophyceae, Fucales)[END_REF]Vergés et al., 2014b;[START_REF] Mitterwallner | Exposure to moderately elevated temperatures changes food preferences in the tropical marine herbivore Haliotis squamata[END_REF].

INTRODUCTION

Over the past decades, coastal areas throughout the world experienced an accelerating trend of harmful algal blooms (HABs) events [START_REF] Anderson | Scaling up from regional case studies to a global harmful algal bloom observing system[END_REF] including the ones due to benthic species [START_REF] Parsons | Gambierdiscus and Ostreopsis: Reassessment of the state of knowledge of their taxonomy, geography, ecophysiology, and toxicology[END_REF]. A significant proportion of studies concerning benthic HABs is focused on tropical benthic dinoflagellate ecology, mostly because of the incidence of ciguatera fish poisoning (CFP) in tropical and subtropical areas [START_REF] Litaker | Taxonomy of Gambierdiscus including four new species, Gambierdiscus caribaeus, Gambierdiscus carolinianus, Gambierdiscus carpenteri and Gambierdiscus ruetzleri (Gonyaulacales, Dinophyceae)[END_REF] and the geographic expansion of some toxic species in temperate waters [START_REF] Rhodes | World-wide occurrence of the toxic dinoflagellate genus Ostreopsis Schmidt[END_REF]. Among them, recurrent Ostreopsis spp. blooms have been recorded throughout the globe: in the Mediterranean Sea (Vila et al., 2001;[START_REF] Penna | Characterization of Ostreopsis and Coolia (Dinophyceae) isolates in the western Mediterranean Sea based on morphology, toxicity and internal transcribed spacer 5.8s rDNA sequences[END_REF]Turki, 2005;Aligizaki and Nikolaidis, 2006;Mangialajo et al., 2008a;Totti et al., 2010;Cohu et al., 2011a;Ismael and Halim, 2012;Manca et al., 2015;Açaf et al., 2020;Gémin et al., 2020), the East Atlantic Ocean [START_REF] Solino | Toxin Profile of Ostreopsis cf. ovata from Continental Portuguese Coast and Selvagens Islands (Madeira, Portugal)[END_REF], the West Atlantic Ocean (Nascimento et al., 2012b;Tibirica et al., 2019), the South-West Pacific (Chang et al., 2000;[START_REF] Rhodes | Toxic marine epiphytic dinoflagellates, Ostreopsis siamensis and Coolia monotis (Dinophyceae)[END_REF]Shears and Ross, 2009), the North-West Pacific (Yamaguchi et al., 2012a;Zou et al., 2020), the South-West Indian Ocean (Lenoir et al., 2004) and the Red Sea (Gomaa et al., 2018). At least five out of the eleven identified species (Fukuyo, 1981;[START_REF] Norris | Benthic dinoflagellates associated with ciguatera from the Florida Keys. 1. Ostreopsis heptagona sp[END_REF]Quod, 1994;Faust and Morton, 1995;Faust, 1999) produce palytoxin-like compounds, one of the most toxic marine compounds (Usami et al., 1995;[START_REF] Ukena | Structure elucidation of ostreocin D, a palytoxin analog isolated from the dinoflagellate Ostreopsis siamensis[END_REF][START_REF] Ciminiello | Complex palytoxin-like profile of Ostreopsis ovata. identification of four new ovatoxins by high-resolution liquid chromatography/mass spectrometry[END_REF][START_REF] Suzuki | LC-MS/MS analysis of novel ovatoxin isomers in several Ostreopsis strains collected in Japan[END_REF]Varela et al., 2021), causing respiratory disease after inhalation of marine aerosols [START_REF] Tichadou | Health impact of unicellular algae of the Ostreopsis genus blooms in the Mediterranean Sea: Experience of the French Mediterranean coast surveillance network from 2006 to 2009[END_REF][START_REF] Tubaro | Case definitions for human poisonings postulated to palytoxins exposure[END_REF]Vila et al., 2016). The whole ecosystem is affected by Ostreopsis spp. blooms [START_REF] Turner | Marine invertebrate interactions with Harmful Algal Blooms -implications for One Health[END_REF], due to mass mortalities of invertebrates such as bivalves, gastropods, crustaceans, and echinoderms (Shears and Ross, 2009;[START_REF] Ramos | Palytoxin and analogs: Biological and ecological effects[END_REF]Guidi-Guilvard et al., 2012;[START_REF] Parsons | Gambierdiscus and Ostreopsis: Reassessment of the state of knowledge of their taxonomy, geography, ecophysiology, and toxicology[END_REF]Accoroni and Totti, 2016;Migliaccio et al., 2016;Neves et al., 2018). In order to effectively manage and mitigate Ostreopsis spp. blooms, a thorough understanding of the bloom dynamics of these species is needed. Samplings have been worldwide restricted mostly to shallow waters as it has been demonstrated in both tropical and temperate areas that Ostreopsis spp. cell abundances are negatively correlated with depth (Richlen and Lobel, 2011;Cohu and Lemee, 2012). The mechanisms that affect Ostreopsis spp. populations are unclear, but most studies relate larger abundances of Ostreopsis spp. to sheltered zones or in low hydrodynamic conditions [START_REF] Di Turi | Ostreopsidiaceae (Dinophyceae) presence along the coastal area of Bari[END_REF]Chiantore et al., 2008;Shears and Ross, 2009;Battocchi et al., 2010;Cabrini et al., 2010;Richlen and Lobel, 2011;Accoroni and Totti, 2016;Mohammad-Noor et al., 2016;Boisnoir et al., 2018;Hachani et al., 2018;Meroni et al., 2018), while others suggest larger abundances of Ostreopsis spp. in higher or slightly higher levels of water motion (Vila et al., 2001;Selina et al., 2014). The inshore occurrence of Ostreopsis spp. has also raised the question concerning their association with nutrient enrichments due to human activities (Faust et al., 1996), but there are mixed conclusions on how nutrient concentrations influence population dynamics of Ostreopsis spp. (Ungaro et al., 2010;Cohu et al., 2011a;Accoroni et al., 2012;Asnaghi et al., 2012). A concise summary of the effects of surface seawater temperature, salinity and nutrient concentrations on Ostreopsis spp. populations in temperate areas can be found in Accoroni and Totti's (2016) and Tester's et al. (2020) reviews. Results from the above-mentioned studies are sometimes contradictory, likely due to the fact that Ostreopsis spp. blooms are a global phenomenon controlled by several factors in very different contexts and, as a consequence, blooms are very variable in space and time, making difficult defining general trends (Mangialajo et al., 2011).

Due to their benthic nature, Ostreopsis spp. develop in close relation with a substrate. Ostreopsis spp. are often described as epiphytic on macroalgae and seagrasses [START_REF] Rhodes | World-wide occurrence of the toxic dinoflagellate genus Ostreopsis Schmidt[END_REF], but can also be found on dead corals, sediments, rocks, and in the water column [START_REF] Bomber | Genetic variability in toxin potencies among seventeen clones of Gambierdiscus toxicus (dinophyceae). Genet. Var. Toxin Potencies Seventeen Clones Gamb[END_REF]Vila et al., 2001;Shears and[START_REF] Shears | Toxic cascades: Multiple anthropogenic stressors have complex and unanticipated interactive effects on temperate reefs[END_REF]Totti et al., 2010). However, the substrate preferences of Ostreopsis spp. are still ambiguous (Vila et al., 2001;Cohu et al., 2013;[START_REF] Sparrow | Effects of temperature, salinity and composition of the dinoflagellate assemblage on the growth of Gambierdiscus carpenteri isolated from the Great Barrier Reef[END_REF]Ternon et al., 2020;[START_REF] Tester | Climate change and harmful benthic microalgae[END_REF], although some studies reported larger abundances on some substrates compared to others (Vila et al., 2001(Vila et al., , 2012;;Mohammad-Noor et al., 2007;Widiarti, 2008;Cabrini et al., 2010;Totti et al., 2010;Accoroni et al., 2011Accoroni et al., , 2012;;Mangialajo et al., 2011;Accoroni and Totti, 2016;Yong et al., 2018;Boisnoir et al., 2019;Bravo et al., 2020;Lee et al., 2020). Such comparisons are difficult because of the limits of cell quantification in function of the type of substrate (i.e. surface for rocks, fresh weight for macrophytes), but also because the measures depend often on some characteristics of the substrate (i.e. the specific weight of sampled species). As an example, some macroalgae species seem to host larger abundance of Ostreopsis spp. than most other species, e.g. Corallina spp. (Simoni et al., 2004;Aligizaki and Nikolaidis, 2006;Monti et al., 2007;Chiantore et al., 2008) and Padina spp. (Aligizaki and Nikolaidis, 2006;Cabrini et al., 2010;Hachani et al., 2018;Gémin et al., 2020). Nonetheless, other species provide discordant patterns such as: Ulva spp. (Aligizaki and Nikolaidis, 2006;Monti et al., 2007;Okolodkov et al., 2007;Totti et al., 2010;Ismael and Halim, 2012), Dictyota spp. (Cohu et al., 2013;[START_REF] Blanfuné | Distribution and abundance of Ostreopsis spp. and associated species (Dinophyceae) in the northwestern Mediterranean: the region and the macroalgal substrate matter[END_REF]González et al., 2019;Gémin et al., 2020;Ternon et al., 2020), and Cystoseira sensu lato spp. (Aligizaki andNikolaidis, 2006, 2008;Monti et al., 2007;[START_REF] Blanfuné | Distribution and abundance of Ostreopsis spp. and associated species (Dinophyceae) in the northwestern Mediterranean: the region and the macroalgal substrate matter[END_REF][START_REF] Catania | The influence of macroalgae on the proliferation and regulation of the benthic dinoflagellate Ostreopsis cf[END_REF]Moncer et al., 2017;Meroni et al., 2018). Ostreopsis abundances on different substrates are highly variable and, despite the technical issues linked to the measurement cited above, several biotic factors acting at different spatial scales can be responsible for such variability: morphology and palatability of macrophytes, herbivory and allelopathic interactions due to the production of secondary metabolites by living substrates, among others [START_REF] Cruz-Rivera | Macroalgal palatability and the flux of ciguatera toxins through marine food webs[END_REF]Totti et al., 2010;Accoroni et al., 2015Accoroni et al., , 2016;;Ternon et al., 2020).

Benthic ecosystems are dynamic systems and, especially due to human impacts, can experience regime shifts, resulting in profound changes in the structure and composition of communities [START_REF] Hughes | Catastrophes, Phase Shifts, and Large-Scale Degradation of a Caribbean Coral Reef[END_REF][START_REF] Benedetti-Cecchi | Predicting the consequences of anthropogenic disturbance: large-scale effects of loss of canopy algae on rocky shores[END_REF][START_REF] Chemello | Regime shifts and alternative stable states in intertidal rocky habitats: State of the art and new trends of research[END_REF][START_REF] Filbee-Dexter | Rise of Turfs: A New Battlefront for Globally Declining Kelp Forests[END_REF][START_REF] Johns | Macroalgal feedbacks and substrate properties maintain a coral reef regime shift[END_REF][START_REF] Melis | Macroalgal forests and sea urchin barrens: Structural complexity loss, fisheries exploitation and catastrophic regime shifts[END_REF][START_REF] Pessarrodona | Homogenization and miniaturization of habitat structure in temperate marine forests[END_REF]. In both temperate and tropical reefs, regime shifts cause a change from healthy communities dominated by foundation species to communities dominated by less structurally complex species (Filbee-Dexter and [START_REF] Filbee-Dexter | Sea urchin barrens as alternative stable states of collapsed kelp ecosystems[END_REF][START_REF] Jouffray | Identifying multiple coral reef regimes and their drivers across the Hawaiian archipelago[END_REF][START_REF] Wernberg | Climate-driven regime shift of a temperate marine ecosystem[END_REF][START_REF] O'brien | Turf wars: competition between foundation and turfforming species on temperate and tropical reefs and its role in regime shifts[END_REF][START_REF] Pessarrodona | Homogenization and miniaturization of habitat structure in temperate marine forests[END_REF]. In temperate areas forest-forming macroalgae are lost and replaced by less complex communities [START_REF] Strain | Identifying the interacting roles of stressors in driving the global loss of canopy-forming to matforming algae in marine ecosystems[END_REF]Vergés et al., 2014a;[START_REF] Bulleri | Facilitation and the niche: implications for coexistence, range shifts and ecosystem functioning[END_REF]; several studies relate major densities of Ostreopsis spp. on algal turfs (Bravo et al., 2020) and in particular on highly urbanised coasts that are usually characterized by post-regime shift algal communities (Mangialajo et al., 2008a;Widiarti, 2008;Cohu et al., 2013), compared to healthier habitats dominated by forest-forming brown algae (Meroni et al., 2018). In tropical areas, ciguatera fish poisoning episodes recurrently follow disturbances to coral reefs by natural and artificial events such as heat waves, hurricanes, dredging, and shipwrecks, among others, where regime shifts from coral to algae are observed [START_REF] De Sylva | Distribution and Ecology of Ciguatera Fish Poisoning in Florida, with Emphasis on the Florida Keys[END_REF][START_REF] Jouffray | Identifying multiple coral reef regimes and their drivers across the Hawaiian archipelago[END_REF][START_REF] Rains | Gambierdiscus species exhibit different epiphytic behaviors toward a variety of macroalgal hosts[END_REF][START_REF] Johns | Macroalgal feedbacks and substrate properties maintain a coral reef regime shift[END_REF]. In this case, higher benthic dinoflagellate abundances are recorded, both on turfforming (Yong et al., 2018;Lee et al., 2020) and forest-forming species, such as Sargassum spp. (Chinain et al., 2020).

To our best knowledge, only few studies assessed the variability of Ostreopsis spp. at a level higher than the substrate (Cohu et al., 2013;Meroni et al., 2018;Yong et al., 2018;Bravo et al., 2020;Lee et al., 2020) with the aim of comparing different habitats to help understanding Ostreopsis spp. variability. To ascertain how the habitat positively or negatively affects Ostreospis spp. blooms, a literature review was conducted in order to describe trends at different spatial scales. In this review we define: (i) the microhabitat (substrate, spatial scale of cm 2 ), as the biotic or abiotic object sampled to quantify/collect Ostreopsis spp. cells (including living or dead organisms and abiotic substrates), (ii) the mesohabitat, as the community or the abiotic material patch where the substrate is sampled (spatial scale of few to a dozen of m 2 , e.g. the macroalgal turf where a certain macroalgal species is sampled as a substrate or the sandy bottom where a pebble is sampled as a substrate; Meroni et al., 2018;Bravo et al., 2020;Lee et al., 2020), and (iii) the macrohabitat, as the ecosystem or the seascape (spatial scales of hundreds/thousands of m 2 ) where the mesohabitat is found (i.e. coral reefs, macroalgal-dominated rocky shores; Yong et al., 2018).

MATERIALS AND METHODS

A bibliographic review assessment was performed using three databases: Aquatic Science and Fisheries Abstracts (ASFA), Web of Science (WoS) and SCOPUS (last update at 1 st February 2021). Search parameters were kept as wide as possible to ensure that all the relevant publications on the topic were found in the database search. The keyword used for the search was "Ostreopsis" and the parameters of the search were to find the word "Ostreopsis" 'everywhere' in the body or text of the publications, between anytime and February 1 st , 2021.

To quantify and compare the relationship between Ostreopsis spp. and the micro-meso and macrohabitats, only articles involving field collection of Ostreopsis spp. were included in this review. The key information extracted from each article included, when available: microhabitat (e.g. sampled substrates such as macrophytes, rocks, shells, artificial substrates, others), mesohabitat (community/abiotic patch type), macrohabitat (ecosystem/seascape), ocean and sea of study, country, temperate or tropical area (tropical areas were defined as areas between -35° and 35°, including both tropical and subtropical climate), geographic coordinates (if not available, they were obtained from Google Earth, whenever possible) of the sampling location where Ostreopsis spp. were sampled, maximum recorded abundance in the study and sampling dates. The maximum abundances of Ostreopsis spp. recorded in the studies were classified in three categories: No bloom CHAPTER 3 ……………………………… 65 (< 200 × 10 3 cells per gram fresh weight (FW) of macrophyte), Bloom (> 200 × 10 3 cells per gram FW of macrophyte) and Major bloom (> 1000 × 10 3 cells per gram FW of macrophyte) as in Mangialajo et al. (2017). Several studies (132) were conducted in two or more separate regions around the globe, seas, countries, locations and/or involving different substrates. In this case, a different line was inserted in the table for each different score, therefore it is possible that one article was accounted more than once. All the graphs have been produced using R Project version 3.6.2.

(R: The R Project for Statistical Computing).

Macroalgal species are characterised by different sizes and shapes, and the communities dominated by different species can be more or less structurally complex, comparable to continental forests, shrubs and turfs, terms often used in the scientific literature (see below). Because abundances of Ostreopsis spp. on macroalgae are subjected to differences in the structure, surface, and fresh weight, the different algal species sampled as a substrate in the reviewed studies were grouped in the following three categories according to their structural morphology:

-Turf-forming algae: algae with tightly packed fronds and filamentous thin cylindrical axes, with a horizontal coverage several times higher than their height forming a dense and compact mat [START_REF] Stewart | Fluctuations in the quantity of sediments trapped among algal thalli on intertidal rock platforms in southern California[END_REF][START_REF] Tittley | The marine algal flora of the Azores and its biogeographical affinities[END_REF][START_REF] Sales | Long-term comparison of algal assemblages dominated by Cystoseira crinita (Fucales, Heterokontophyta) from Cap Corse (Corsica, North Western Mediterranean)[END_REF]Sala et al., 2012;[START_REF] Connell | What are algal turfs? Towards a better description of turfs[END_REF][START_REF] Thiriet | Abundance and Diversity of Crypto-and Necto-Benthic Coastal Fish Are Higher in Marine Forests than in Structurally Less Complex Macroalgal Assemblages[END_REF][START_REF] Mauffrey | Seaweed functional diversity revisited: Confronting traditional groups with quantitative traits[END_REF].

-Shrub-forming algae: foliose laminar, ribbon-like, massive or fan-like thallus and erect arborescent tufts. Algae usually having a smaller size and forming less complex communities than forest-forming species. At the same time, in their region, they generally have a bigger size and form communities characterised by higher complexity than turf forming algae [START_REF] Sala | The Structure of Mediterranean Rocky Reef Ecosystems across Environmental and Human Gradients, and Conservation Implications[END_REF][START_REF] Thiriet | Abundance and Diversity of Crypto-and Necto-Benthic Coastal Fish Are Higher in Marine Forests than in Structurally Less Complex Macroalgal Assemblages[END_REF][START_REF] Bertolini | Can secondary species maintain a primary role? Consistent inter-regional effects of understory algae on diversity[END_REF].

-Forest-forming algae: cylindrical axes, branched and tree-like characterised by a complex tri-dimensional structure, generating a canopy. Algae having a large size compared to other algae growing in the same region. This term is generally used for Laminariales, Fucales and some Tilopteridales [START_REF] Sala | The Structure of Mediterranean Rocky Reef Ecosystems across Environmental and Human Gradients, and Conservation Implications[END_REF][START_REF] Strain | Identifying the interacting roles of stressors in driving the global loss of canopy-forming to matforming algae in marine ecosystems[END_REF][START_REF] Thiriet | How 3D Complexity of Macrophyte-Formed Habitats Affect the Processes Structuring Fish Assemblages Within Coastal Temperate Seascapes?[END_REF][START_REF] Thiriet | Abundance and Diversity of Crypto-and Necto-Benthic Coastal Fish Are Higher in Marine Forests than in Structurally Less Complex Macroalgal Assemblages[END_REF][START_REF] Bertolini | Can secondary species maintain a primary role? Consistent inter-regional effects of understory algae on diversity[END_REF][START_REF] Shelamoff | Ecosystem engineering by a canopy-forming kelp facilitates the recruitment of native oysters[END_REF][START_REF] Assis | A finetuned global distribution dataset of marine forests[END_REF].

RESULTS

The bibliometric search resulted in 1157 publications including the word "Ostreopsis" in the text

Spatial scales

Macrohabitat

The macrohabitat was detailed in a total of 197 out of the 249 papers (79.1%). In 12 of them (6.1% studies) at least two different ecosystems/seascapes were sampled. From the 957 sampled locations, 897 (93.7%) included or allowed extrapolation of the information at the large scale (ecosystem/seascape; Figure 3). In temperate areas, Ostreopsis spp. is mostly sampled in rocky reefs (usually dominated by macroalgal communities), while in tropical areas Ostreopsis spp. is generally sampled in coral reefs. Soft bottoms are also regularly sampled, especially in tropical areas.

Mesohabitat

The mesohabitat was detailed in a total of 87 out of the 249 papers (34.9%). In 30 of them (12% studies) at least two different mesohabitats were sampled. From the 957 sampled locations, 301 (31.4%) included or allowed extrapolation of the information at the intermediate scale (Figure 4).

In temperate and in tropical areas, Ostreopsis spp. is mostly sampled in algal communities (6.2%

Microhabitat

The microhabitat was detailed in a total of 243 out of the 249 (97.6%) studies considered. In almost half of the studies (113 studies, 45.4%) at least two different substrates were sampled. From the 957 sampled locations, 950 (99.3%) included information about the sampled substrate. On 527 (55.1%) sampled locations out of the 957, two or more substrates were sampled, resulting in a total of 1644 different sampled substrates. The frequency of the total sampled substrates is reported in Figure 5. Three additional substrates were sampled and are not accounted for in the present study: a Bryozoan (Di Pippo and Congestri, 2017), the gut contents of the herbivorous fish Sarpa salpa (Bellassoued et al., 2013) and floating plastic debris (Masó et al., 2003;Casabianca et al., 2019;Tibirica et al., 2019Tibirica et al., , 2019)).

Artificial substrates, that allow an easier standardisation of Ostreopsis spp. abundances (Jauzein et al., 2018;Yong et al., 2018;Fernandez-Zabala et al., 2019;Lee et al., 2020); were used in 24 out of the 249 studies (3.6% in temperate areas and 6% in tropical), and in 136 out of the 1644 sampled substrates (2.3% in temperate areas and a 6% in tropical). Seawater is also commonly sampled and, in some countries, thresholds alerts are based on cell concentrations in seawater at dozen of centimetres above the sea bottom [START_REF] Funari | Ostreospis cf. ovata blooms in coastal water: Italian guidelines to assess and manage the risk associated to bathing waters and recreational activities[END_REF]. Water was sampled in 100 out of the 249 studies (32.5% in temperate and 7.6% in tropical areas) and represent 332 out of the 1644 sampled substrates (15.7% in temperate and 4.5% in tropical areas). Without considering seawater, macroalgae are by far the preferred substrate for sampling Ostreopsis spp. Out of the 249 studies, 179 (71.9 %) included at least a macroalgal sample. A total of 792 macroalgae specimens were sampled in tropical and temperate areas. Macroalgae are commonly sampled both in temperate (where they dominate rocky reefs habitats) and in tropical areas (where, on the contrary, habitats are usually dominated by corals). The second preferred substrate is represented by seagrasses, followed by sand and sediments, coral fragments, pebbles/rocks and dead mollusc shells. Most studies (56.8%) sampling macrophytes identified the sample at the species level, while 20.1% out of them identified the macrophyte at genus level. Some studies (2.9%) identified the species sampled only at a level higher than genus or using nontaxonomic classification (i.e. turf). The remaining fraction (20.2%) did not provide any taxonomic information. It is worth noting that quantification of Ostreopsis spp. cell abundances on invertebrates (corals or molluscs) is generally performed on dead coral fragments or shells (Faust et al., 1996;Faust, 1999;Mohammad-Noor et al., 2007;Okolodkov et al., 2007;Xu et al., 2014).

Macroalgae samples, that involved the collection of 792 different specimens, represent nearly half of the sampled substrates (47.7%) and are a polyphyletic and extremely diversified group of organisms. The different macroalgal phyla, Rhodophyta, Ochrophyta and Chlorophyta [START_REF] Ruggiero | A Higher Level Classification of All Living Organisms[END_REF] were sampled in different proportions, in particular in temperate areas where Rhodophyta (49.2%) seem to be sampled preferentially, followed by Ochrophyta (36.7%) and Chlorophyta (14%). In tropical areas, the sampling frequency of each phylum is similar (39.4%, 36.7% and 23.8%, respectively; Figure 6). One of the major limitations and controversies for studying macrophyte preferences is the difficulty of standardising cell densities. In order to compare and assess the role of each macroalgae species in structuring the community, we classified the macroalgae sampled as a substrate in three different groups according to their physical structure. The algae sampled in the considered studies were classified, based to our knowledge about their mean size and their appearance in "forest-forming", "shrub-forming" and "turf-forming", as defined in the "Materials and methods" section. Following this classification, most of the macroalgae sampled in both temperate and tropical areas correspond to the category shrub-forming (55.9%), followed by the turf-forming (31.7%) and the forestforming (12.3%; Figure 7). Of the 249 studies, 82 (32.9%) reported abundances of Ostreopsis spp. cells per gram FW of macrophyte. Such quantification is dependent on the specific weight and morphology of the different species (Mangialajo et al., 2017) and would not, ideally, allow a direct comparison of CHAPTER 3 ……………………………… 71 abundances on different macroalgal species. At present, the few alternative methods allowing a standardised quantification (Tester et al., 2014;Jauzein et al., 2016;Mangialajo et al., 2017) are only sporadically applied and large-scale comparisons are performed on the measures of cells per gram FW. In order to have a global vision of the Ostreopsis spp. abundances, the maximal abundance reported on each study indicating the host macrophyte has been reported, when available, in Table 1 (expressed as cells/g of macroalga FW). From the data reported on Table 1, the maximum cell densities of Ostreopsis spp. (cells g -1 FW macroalgae) are grouped according to the type of mesohabitat the macroalgae sampled can form and the intensity of the bloom (N = 54; Figure 8; Mangialajo et al., 2017). In the case of Bloom and No bloom scenarios (according to Mangialajo et al. (2017)), the abundance of Ostreopsis spp. cells seems to be higher on shrub and turf-forming species. For the Major bloom scenario (see classification in Mangialajo et al. (2017)), the abundance of Ostreopsis spp. seems to be higher on shrub-forming species in temperate areas (followed by high values in forest-forming) and on forest-forming species in tropical areas.

DISCUSSION

The first indexed papers involving Ostreopsis spp. sampling have been published in tropical areas.

The bibliometric study shows a rise in scientific publications presumably linked to the increasing HABs incidences in recent decades [START_REF] Anderson | Scaling up from regional case studies to a global harmful algal bloom observing system[END_REF], and in particular since Ostreopsis spp.

blooms have spread in temperate areas (Shears and Ross, 2009) The same considerations could be valid for shells, pebbles, and stones, but there is no specific literature on these particular cases to our knowledge. On the contrary, the description of the mesohabitat (e.g. macroalgal turfs, macroalgal forests, coral-dominated communities, seagrasses, or sandy patches) is rarely reported (35% of the considered studies). As highlighted in Pavaux et al. (2020) it is undeniable that only a small part of the research on Ostreopsis spp. is focused on ecological aspects. Interestingly, recent studies, mainly focused on tropical species related to ciguatera, suggest that habitat heterogeneity and complexity affect benthic dinoflagellate communities [START_REF] Rains | Gambierdiscus species exhibit different epiphytic behaviors toward a variety of macroalgal hosts[END_REF]Meroni et al., 2018;Yong et al., 2018;Boisnoir et al., 2019;Bravo et al., 2020;Lee et al., 2020). In particular Yong et al. (2018) found higher Gambierdiscus, Prorocentrum and Amphidinium cell abundances on reefs with high turf algal cover and coral rubble, while Ostreopsis preferred mesohabitats with high coral cover. Lee et al. (2020) found that the mesohabitat influence the distribution of benthic harmful algal blooms and that Ostreopsis and Gambierdiscus are both associated with turf algal communities and hard coral. Bravo et al. (2020) observed an influence of the macrophyte's thallus architecture on dinoflagellates abundances and also reported higher abundances of Ostreopsis on turf Rhodophyta, while Gambierdiscus and Sinophysis seem to be more abundant on Rhodophyta and Ochrophyta. More information at the global scale on how habitat affects benthic dinoflagellate blooms would allow to better disentangle the different factors playing a role in Ostreopsis spp. blooms.

Results from this literature review highlight that most data available on benthic substrates are from studies focusing on Ostreopsis spp. attached to biotic substrates and in particular, macroalgae.

Interestingly, independently of the macrohabitat sampled, most of the studies in both temperate and tropical areas chose macroalgae (mainly visually obvious macroalgae species) as substrate. Even in studies conducted in coral reefs (where macroalgae are usually not dominant), macroalgae, rather than living corals or dead coral fragments, are selected as sampled substrate, meaning that i) macroalgae could be very abundant in this particular site and/or ii) scientists believe there are larger abundances of Ostreopsis spp. on macroalgae rather than on other substrates. However, choosing macroalgae over corals as substrate to be sampled could be for ethical reasons (coral species may be protected under international or national environmental law) or other practicalities of the sampling. Concerning the different species, Park et al. (2020) report higher Ostreopsis spp. abundances on red (Spyridia filamentosa and Laurencia complex) and brown macroalgae (i.e. Dictyota spp., Halopteris scoparia, Dictyopteris polypodioides and Carpophyllum plumosum) that are also the most sampled macroalgal groups while lower densities are reported in Chlorophyta, and in particular Ulva spp. Conversely, some of these results contrast with the results from studies on the effect of metabolites from macroalgae on the growth and settlement of Ostreopsis cf. ovata (Accoroni et al., 2015;Ternon et al., 2020) that suggest a strong negative effect of some Dictyotales on O. cf. ovata. However, [START_REF] Pavaux | Chemical Ecology of the Benthic Dinoflagellate Genus Ostreopsis : Review of Progress and Future Directions[END_REF] and Ternon et al. (2020) suggest that macroalgae surface chemistry may be not enough to explain Ostreopsis spp. settlement preferences and that other factors such as other organisms of the epiphytic community (bacteria, viruses, parasites, microalgae, and fauna) and the whole surrounding community are likely to modulate the growth of Ostreopsis spp.

With respect to the mesohabitat, the information in the literature is scarce. The most sampled macroalgae are species forming medium to low complexity communities, such as shrubs formed by Padina spp., Dictyota spp., Halopteris spp. (Cabrini et al., 2010;Widiarti and Anggraini, 2012;Cohu et al., 2013;[START_REF] Blanfuné | Distribution and abundance of Ostreopsis spp. and associated species (Dinophyceae) in the northwestern Mediterranean: the region and the macroalgal substrate matter[END_REF]Boisnoir et al., 2019) and turfs formed by Corallina spp., Jania spp., Laurencia spp., Hypnea spp. (Simoni et al., 2004;Monti et al., 2007;Totti et al., 2010;Kim et al., 2011;[START_REF] Abbate | A new method for sampling potentially toxic benthic dinoflagellates[END_REF]Ismael and Halim, 2012;[START_REF] Blanfuné | Distribution and abundance of Ostreopsis spp. and associated species (Dinophyceae) in the northwestern Mediterranean: the region and the macroalgal substrate matter[END_REF]Yong et al., 2018). The sampling is often performed on the most abundant or predominant macroalgal species (Mangialajo et al., 2008a(Mangialajo et al., , 2011;;Accoroni et al., 2011;[START_REF] Lemee | Interactions between scientists, managers and policy makers in the framework of the French MediOs project on Ostreopsis (2008-2010)[END_REF]Rhodes et al., 2017;Jauzein et al., 2018;Boisnoir et al., 2019;Chinain et al., 2020;Gémin et al., 2020) and it can therefore be deduced that the representative mesohabitats in the sites where scientists study Ostreopsis spp. blooms are potentially characterized by algal shrubs and turfs. This is in agreement with the results of the few ecological studies considering the mesohabitat scale, where larger Ostreopsis spp. densities are found on low-complexity macroalgal communities (shrubs or turfs) characteristic of post-regime shift scenarios in highly impacted locations (Mangialajo et al., 2008a;Meroni et al., 2018;Bravo et al., 2020;Lee et al., 2020). This is particularly true in temperate areas, where macroalgal communities dominate rocky reefs, and suggest that lower abundances of Ostreopsis spp. could be found on large brown algae forests of fucoids and kelps, e.g. Cystoseira sensu lato spp., Sargassum spp., etc; (Mangialajo et al., 2008b;Catania, 2017, 2;Meroni et al., 2018). Nevertheless, blooms are observed on Ericaria crinita in the Adriatic Sea (Pfannkuchen et al., 2012) and on Carpophyllum plumosum populations in New Zealand (Shears and Ross, 2009).

In tropical areas important blooms of Ostreopsis spp. can be observed on forest-forming macroalgal species (Sargassum spp., Chinain et al., 2020), where regime shifts from coral to algae (often with a dominance of large size species; [START_REF] Payri | Variations entre 1971 et 1980 de la biomasse et de la composition des populations de macroalgues sur le récif corallien de Tiahura (Île de Moorea, Polynésie française)[END_REF][START_REF] Stiger | Spatial and Seasonal Variations in the Biological Characteristics of Two Invasive Brown Algae, Turbinaria ornata (Turner) J. Agardh and Sargassum mangarevense (Grunow) Setchell (Sargassaceae, Fucales) Spreading on the Reefs of Tahiti (French Polynesia)[END_REF] and subsequent coral reef degradation, are usually associated to human impacts [START_REF] Stiger | Spatial and Seasonal Variations in the Biological Characteristics of Two Invasive Brown Algae, Turbinaria ornata (Turner) J. Agardh and Sargassum mangarevense (Grunow) Setchell (Sargassaceae, Fucales) Spreading on the Reefs of Tahiti (French Polynesia)[END_REF]. But such results, based on the maximum Ostreopsis spp. abundances occurring at a study site have to be considered with care, because of the potential bias linked to the classic sampling method, that quantify cells/gram of macroalga, providing an estimate that is species dependent (i.e. specific weight, Mangialajo et al., 2017).

The present review reveals that most studies do not provide detailed descriptions of the mesohabitats (or benthic communities) where the studied blooms occur and how the mesohabitat could play a role on Ostreopsis spp. bloom dynamics. Benthic dinoflagellate preferences at different habitat scales (substrate, community, and ecosystem) and biotic interactions present untapped fields of research with great potential which still need to be addressed in the future. Our results seem to predict a larger risk exposure to toxic effects for humans in post-regime shifts communities such as urban or degraded areas, where marine forests of large brown algae and/or reef building corals are often lost and replaced by less complex species that could host large densities of Ostreopsis spp. [START_REF] Hughes | Catastrophes, Phase Shifts, and Large-Scale Degradation of a Caribbean Coral Reef[END_REF][START_REF] Benedetti-Cecchi | Predicting the consequences of anthropogenic disturbance: large-scale effects of loss of canopy algae on rocky shores[END_REF]Mangialajo et al., 2008a;[START_REF] Catania | The influence of macroalgae on the proliferation and regulation of the benthic dinoflagellate Ostreopsis cf[END_REF][START_REF] Filbee-Dexter | Rise of Turfs: A New Battlefront for Globally Declining Kelp Forests[END_REF][START_REF] Johns | Macroalgal feedbacks and substrate properties maintain a coral reef regime shift[END_REF]Meroni et al., 2018). Understanding the underlying mechanisms on how the habitat can affect Ostreopsis spp. blooms, would allow a better risk assessment of the phenomenon, identifying areas at risk simply on the base of the benthic habitats.

These findings underline the need of a better understanding of Ostreopsis spp. ecology to prevent socio-economic damage, reduce human health risks in coastal regions, and reduce ecological impacts to marine coastal ecosystems.

INTRODUCTION

Harmful algal bloom (HABs) events have been increasing in coastal areas around the globe in the last decades [START_REF] Anderson | Scaling up from regional case studies to a global harmful algal bloom observing system[END_REF]. Such increase is referred to ocean warming resulting from global change that would favour the expansion and growth rates of tropical and sub-tropical benthic harmful dinoflagellates including the genera Ostreopsis, Gambierdiscus and Fukoya in areas where habitat requirements are satisfied [START_REF] Tester | Climate change and harmful benthic microalgae[END_REF]. The expansion and faster growth rates of toxic benthic dinoflagellates could multiply the number and intensity of benthic harmful algal blooms (BHABs) events in temperate areas, which could have negative repercussions on human and marine ecosystems health [START_REF] Tester | Climate change and harmful benthic microalgae[END_REF].

The genus Ostreopsis, belonging to the family of Ostreopsidaceae (Gonyaulacales, Dinophyceae, Dinozoa) includes several species largely distributed from tropical to temperate marine coastal areas worldwide. Ostreopsis is a benthic dinoflagellate that has been reported for a long time in tropical ciguatera endemic areas and, for some decades has become common in temperate areas as well [START_REF] Litaker | Taxonomy of Gambierdiscus including four new species, Gambierdiscus caribaeus, Gambierdiscus carolinianus, Gambierdiscus carpenteri and Gambierdiscus ruetzleri (Gonyaulacales, Dinophyceae)[END_REF][START_REF] Rhodes | World-wide occurrence of the toxic dinoflagellate genus Ostreopsis Schmidt[END_REF][START_REF] Zingone | Toxic marine microalgae and noxious blooms in the Mediterranean Sea: A contribution to the Global HAB Status Report[END_REF]. In temperate areas, its blooms are CHAPTER 4 ………………….………….

82

often associated with toxic effects on humans and benthic marine ecosystems (Accoroni and Totti, 2016). Some species of Ostreopsis produce palytoxin-like compounds, one of the most toxic marine compounds [START_REF] Ukena | Structure elucidation of ostreocin D, a palytoxin analog isolated from the dinoflagellate Ostreopsis siamensis[END_REF][START_REF] Suzuki | LC-MS/MS analysis of novel ovatoxin isomers in several Ostreopsis strains collected in Japan[END_REF]Varela et al., 2021), causing respiratory disease after inhalation of marine aerosols in humans (Vila et al., 2016) and mass mortalities of invertebrates in marine ecosystems [START_REF] Parsons | Gambierdiscus and Ostreopsis: Reassessment of the state of knowledge of their taxonomy, geography, ecophysiology, and toxicology[END_REF]Accoroni and Totti, 2016). With the aim of better understanding and modelling blooms of Ostreopsis spp. an important research effort has been done for assessing the role of the effects of environmental factors (e.g. nutrients, salinity and seawater temperature) on the occurrence and dynamics of HABs. The results of such studies are often contrasting, showing a highly variable response of Ostreopsis spp. to these factors (reviewed in [START_REF] Parsons | Gambierdiscus and Ostreopsis: Reassessment of the state of knowledge of their taxonomy, geography, ecophysiology, and toxicology[END_REF]Accoroni and Totti, 2016).

Ostreopsis spp. blooms develop in close relationship with the substrate and different interactions at different special scales (i.e. microhabitat, mesohabitat and macrohabitat) could regulate Ostreopsis spp. development (Cohu et al., 2013;[START_REF] Monserrat | The role of habitat in the facilitation of Ostreopsis spp. blooms[END_REF]. Even if an antagonistic relationship between macroalgae and microalgae has been reported in both natural and experimental conditions [START_REF] Lee | Eutrophication and Management Initiatives for the Control of Nutrient Inputs to Rhode Island Coastal Lagoons[END_REF][START_REF] Fong | Competition with macroalgae and benthic cyanobacterial mats limits phytoplankton abundance in experimental microcosms[END_REF], Ostreopsis spp. are known to develop on the surface of macroalgae forming a biofilm (Totti et al., 2010;Accoroni et al., 2015). In both temperate and tropical areas Ostreopsis spp. are mainly sampled on macroalgal species that form low or medium complex communities (turf and shrub-like macroalgal communities), typical of post-regime shift scenarios and degraded habitats (Mangialajo et al., 2008a;[START_REF] Sales | Long-term comparison of algal assemblages dominated by Cystoseira crinita (Fucales, Heterokontophyta) from Cap Corse (Corsica, North Western Mediterranean)[END_REF]Meroni et al., 2018;Yong et al., 2018;Bravo et al., 2020;Lee et al., 2020). Recent studies highlighted the importance of the macrophyte morphology on the abundance of dinoflagellates: filamentous morphotypes and macroalgae forming entangled groups (turf-forming macroalgae) are suggested to host higher abundances, especially from the genus Ostreopsis (Totti et al., 2010;[START_REF] Mustapa | Growth and epiphytic behavior of three Gambierdiscus species (Dinophyceae) associated with various macroalgal substrates[END_REF]Bravo et al., 2020). The fact that BHABs could be associated with structurally less complex macroalgal communities (turf and shrubs) is of special interest, due to the generalized phenomenon of habitat miniaturization result of regime shifts in temperate ecosystems [START_REF] Pessarrodona | Homogenization and miniaturization of habitat structure in temperate marine forests[END_REF]. Mainly due to anthropogenic pressures (e.g. local pollution, habitat destruction, climate change), structurally complex macroalgal communities, such as marine forests, are being substituted by less complex macroalgal communities, such as turfs, with the consequent loss of ecosystem functions and perturbations at the ecosystem level that could favour BHAB [START_REF] Parsons | Gambierdiscus and Ostreopsis: Reassessment of the state of knowledge of their taxonomy, geography, ecophysiology, and toxicology[END_REF][START_REF] Filbee-Dexter | Rise of Turfs: A New Battlefront for Globally Declining Kelp Forests[END_REF][START_REF] Benedetti-Cecchi | Species Interactions and Regime Shifts in Intertidal and Subtidal Rocky Reefs of the Mediterranean Sea[END_REF][START_REF] Orfanidis | Effects of Natural and Anthropogenic Stressors on Fucalean Brown Seaweeds Across Different Spatial Scales in the Mediterranean Sea[END_REF][START_REF] Smale | Climate-driven substitution of foundation species causes breakdown of a facilitation cascade with potential implications for higher trophic levels[END_REF]. Furthermore, it is thought that the metabolites produced by some macroalgal species could also influence the settlement and growth of Ostreopsis spp. (Accoroni et al., 2015;Ternon et al., 2020). Some macroalgal species such as brown macroalgae are known to produce secondary metabolites with an important chemical effect against marine herbivores [START_REF] Schnitzler | Chemical defense of brown algae (Dictyopteris spp.) against the herbivorous amphipod Ampithoe longimana[END_REF][START_REF] Wiesemeier | The potential role of wound-activated volatile release in the chemical defence of the brown alga Dictyota dichotoma: Blend recognition by marine herbivores[END_REF][START_REF] Jormalainen | Resistance of the brown alga Fucus vesiculosus to herbivory[END_REF]. Some studies even suggest the CHAPTER 4 ……………………………… 83 use of macroalgae in the prevention, control and mitigation of BHABs as some macroalgal species have been proven to exert negative effects against harmful benthic dinoflagellates, including Ostreopsis cf. ovata (Accoroni et al., 2015;Ternon et al., 2020). Nevertheless, the substrate and habitat preferences of Ostreopsis spp. are still unclear, and possibly linked to other biotic and abiotic factors including other components of the epiphytic community [START_REF] Pavaux | Chemical Ecology of the Benthic Dinoflagellate Genus Ostreopsis : Review of Progress and Future Directions[END_REF]Ternon et al., 2020;[START_REF] Roselli | Harmful algae and pressure-impact relationship: Noxious blooms and toxic microalgae occurrence from coastal waters of the Apulia region (Adriatic and Ionian Seas, Mediterranean)[END_REF]. Because of that, the existence of direct and indirect effects at smaller (e.g. epiphytic community) and larger (e.g. the macroalgal community) spatial scales than the substrate that affect Ostreopsis spp. proliferation are expected.

Yet, direct comparisons among different macroalgae, the most common sampled substrate [START_REF] Monserrat | The role of habitat in the facilitation of Ostreopsis spp. blooms[END_REF], are hindered by the quantification procedure that, most of the time takes into account the number of microalgal species as cells per gram of fresh weight (FW) of the macroalgae. Such quantification is strongly affected by the specific weight and morphology of the macroalgae, therefore not allowing appropriate comparisons between macroalgal species (Totti et al., 2010;Mangialajo et al., 2017;[START_REF] Tester | Using artificial substrates to quantify Gambierdiscus and other toxic benthic dinoflagellates for monitoring purposes[END_REF]. Even if efforts have been performed using alternative methodologies independent from the specific weight of the substrate allowing a standardized quantification, they are not yet applied at a large scale (Tester et al., 2014;Jauzein et al., 2016;Mangialajo et al., 2017), hindering finding trends for Ostreopsis spp. preferences.

There is currently a need for a better understanding of the ecology of Ostreopsis spp. and to reduce human health and ecological risks in coastal regions, preventing indirect socioeconomic damage [START_REF] Berdalet | Marine harmful algal blooms, human health and wellbeing: challenges and opportunities in the 21st century[END_REF]. Understanding the mechanisms affecting Ostreopsis spp. blooms, would allow a better risk assessment of BHABs, and eventually the identification of areas at risk in function of their benthic characteristics. The objective of this study is to assess the effect of different macroalgal species (small scale or microhabitat) and macroalgal communities (medium scale or mesohabitat) on the abundances of Ostreopsis spp. We hypothesise that (i) individual forestforming macroalgae and marine forests, the most structurally complex macroalgal communities (dominated by large brown forest-forming macroalgae) will host lower abundances of Ostreopsis spp.; while (ii) medium and low structurally complex macroalgal species and related macroalgal communities (dominated by fast-growing macroalgal species with a larger number of filaments and entangled clumps, (i.e. shrub and turf-forming macroalgae) characteristic in post-regime shift scenarios and degraded habitats; will foster the highest abundances of Ostreopsis spp.

As Ostreopsis spp. abundances on the biofilm associated to macroalgae can vary in function of different factors at the microhabitat (macroalgae) scale (i.e. morphology, surface, allelopathy) and at the mesohabitat (macroalgal community) scale (i.e. resistance to wave energy, presence of herbivores), in each location we sampled: (i) different macroalgal species according to their structural morphology (i.e. turf-forming, shrub-forming and forest-forming); (ii) artificial macroalgae also according to their structural morphology, dismissing any chemical or allelopathic effect; and (iii) different macroalgal communities, characterized by the same dominant species sampled for the microhabitat scale preference assessment.

MATERIAL AND METHODS

Sampling sites

Two locations with known annual summer blooms of Ostreopsis spp. were selected for the study.

The two sampling sites, Rochambeau and Vernazzola, are situated on the continental N-W Mediterranean coast more than 150 km apart. The sampling site Rochambeau (Villefranche-sur-Mer, France; 43°41′34.83′′ N and 7°18′31.66′′ E), consists in a small creek of the Bay of Villefranche-sur-Mer characterized by a sheltered rocky coast. The sampling site Vernazzola (Genova, Italy; 44°23′24.23′′ N and 8°58′40.13′′ E), consists in a shallow exposed rocky coast next to the centre of Genova (Figure 1). The marine vegetation in both locations is characterized by turf and shrub-forming macroalgae, mainly Halopteris scoparia and Padina pavonica and to a lesser extent Dictyota spp., Ulva spp., Caulerpa cylindracea and Jania rubens. In each sampling site, we sampled different types of natural and artificial macroalgae species (for the small scale or microhabitat scale) and macroalgal communities (for the medium or mesohabitat scale) according to their structural morphology: turf, shrub and forest-forming macroalgae species were sampled.

However, not all the different types of macroalgae species were present in the sampling locations, in this case, we transplanted individuals and stones colonized with the missing macroalgal species one week before starting the experiment.

For the microhabitat preference assessment, in each sampling location five species (or groups of species) of individual natural macroalgae were sampled (n = 4; Cohu et al., 2013;Jauzein et al., 2018). The sampled macroalgal species are reported in Table 1; as much as possible the same species were selected in the two sampling locations. Artificial macroalgae made of latex with the same surface area but different sizes and number of ramifications were also sampled for the microhabitat preference assessment. The artificial macroalgae were attached to a plastic lid and fixed to the sea bottom with cable tides as in Fricke et al. (2018) in both sampling locations (n = 4; see Appendix D Chapter 4, Supplementary material S1). Four different types of artificial macroalgae were used and classified according to their similarity to natural macroalgae, from a smaller and higher number of ramifications to larger and fewer number of ramifications: artificial Turf, Dictyota, Halopteris and Cystoseira like, and an empty lid was used as a control treatment (Appendix D Chapter 4 Supplementary material S1). * By the end of the experiment the two transplanted macroalgal communities in Rochambeau corresponding to Cystoseira compressa and Gongolaria barbata, experienced a reduction in the macroalgal biomass.

The sampled macroalgal communities (mesohabitat scale) were dominated by the same species of macroalgae selected for the microhabitat assessment (Table 1). However, because of the lack of forest-forming species in Rochambeau, stones colonized by Cystoseira compressa and Gongolaria barbata were transplanted to Rochambeau for the duration of the experiment from Sainte Marguerite Island (Lérins Islands, Cannes, France). Sainte Marguerite Island is one of the last locations with healthy Cystoseira sensu lato forests in the French Riviera [START_REF] Thibaut | Decline and local extinction of Fucales in French Riviera: the harbinger of future extinctions?[END_REF]. In Rochambeau, a modified BEDI device (Benthic Dinoflagellate Integrator; Mangialajo et al., 2017) was used in order to sample the selected communities (see section 2.2. Collection and processing of samples). In both sites, 0.04 m 2 scrapings (n=3) of the selected communities were performed in CHAPTER 4 ……………………………… 87 order to extrapolate the abundances of Ostreopsis spp. from the communities, expressed as cells per surface of seabottom (see section 2.2. Collection and processing of samples).

For each site, also abundances of Ostreopsis spp. cells on the surrounding water were sampled (n = 4 in Rochambeau and n = 2 in Vernazzola). The temperature was monitored every hour during the duration of the experiment at both sampling sites at 0.5 m depth using a temperature data logger (HOBO Pendant MX Temp, ONSET).

Collection and processing of samples

Ostreopsis spp. cells on the surrounding water were determined from seawater samples collected in 250 mL plastic flasks 20 cm above the natural macroalgal substrate, in particular turf (Table 1), as described in Cohu et al. (2013). Samples of individual natural macroalgae (microhabitat scale samples) were carefully collected into a 250 mL flask with the surrounding water, as described by Cohu et al. (2013). Abundances of Ostreopsis spp. on artificial macroalgae, with the surrounding water, were sampled in 1 L flasks coiled to the lid at the base of the artificial macroalgae, in a similar way as in Fricke et al. (2018), closing the flask underwater. Then, once the flask was hermetically closed, the lid with the macroalgae and the flask were detached from the base cutting the cable tides and pulled out of the water. Afterwards the flask was shaken to detach the cells of Ostreopsis spp. from the artificial macroalgae and the lid. Finally, the 1 L flask with the sample was closed using another lid and the lid with the artificial macroalgae was fixed again at the same point on the sea bottom with cable tides.

The cells at the mesohabitat scale were assessed in two ways: (i) using the BEDI device and (ii) extrapolated from the total biomass of macroalgae obtained from the scraping. In the first assessment, a modified adapted BEDI device (Mangialajo et al., 2017) consisting of a hollow plastic cylinder open at both ends was used. In order to avoid cell loss, the bottom part was provided of a fixed rubber seal and the upper part was kept out of the water. The adapted BEDI device used in this study covered an area of 314.16 cm 2 of the sea bottom and had a total volume of 9.42 L. To homogenize cells of Ostreopsis spp. in the area and volume covered, a propeller was installed on the inside upper part of the BEDI. Once the device was placed on the macroalgal communities, the sea water level inside the BEDI was noted and the propeller was rotated 10 times from the outside of the BEDI thanks to a crank. Then a 60 mL sample was extracted from the inside of the adapted BEDI device using a syringe. For the second assessment, the total biomass of macroalgae for each macroalgal community was calculated from the total biomass of macroalgae obtained from the scrapings. In both sites, 0.04 m 2 of all the considered communities were sampled (n = 3). Once in the laboratory, the dominant species were centrifugated for 30 seconds and weighted (FW). In both cases (BEDI sampling and extrapolation from scrapings) the results can be expressed as the number CHAPTER 4 ………………….…………. of cells per seabottom surface area (i.e. 0,04 m 2 ) independently of the specific weight of the substrate (Mangialajo et al., 2017).

All the samples were fixed with acidic Lugol (2% v/v). For the seawater samples the Utermöhl method [START_REF] Utermöhl | Methods of collecting plankton for various purposes are discussed[END_REF] was used to estimate abundances (in number of cells / L). The fixation of the natural macroalgae samples was followed by vigorous shaking and filtration of the sample through a 500 μm mesh to separate the macroalgae from the seawater containing Ostreopsis spp. cells and the macroalgae were weighed (± 0.01 g; Jauzein et al., 2018). The abundances of Ostreopsis spp. cells on natural macroalgae, artificial macroalgae and macroalgal communities were estimated under a microscope (Axio Observer D1, Zeiss) using a Sedgewick Rafter counting chamber. The abundance of cells at the microhabitat scale were expressed as number of cells per gram of fresh weight of macroalgae (cells / g FW). The density of cells of Ostreopsis spp. on artificial macroalgae was expressed as cells / cm 2 of surface of artificial macroalgae (Appendix D Chapter 4 Supplementary material S1). At the mesohabitat scale abundances were expressed as cells / 0.04 m 2 .

Our study was conducted during the different phases of the bloom (exponential, peak and decreasing phase) to assess eventual changes in Ostreopsis spp. preferences in function of the dynamic of the bloom. In order to define the magnitude of the bloom, the sampled abundances of Ostreopsis spp. were classified in Major bloom (>1000 × 10 3 cells / g FW), Bloom (>200 × 10 3 cells / g FW), Alert (>30 × 10 3 cells / L), and No bloom (<200 × 10 3 cells / g FW) according to the definition from [START_REF] Funari | Ostreospis cf. ovata blooms in coastal water: Italian guidelines to assess and manage the risk associated to bathing waters and recreational activities[END_REF] and Mangialajo et al. (2017).

Data analysis

Abundances of Ostreopsis spp. on natural macroalgae (microhabitat scale): A Generalized Linear Mixed-Effects Model (GLMM), with a Gamma link log distribution function was used to test the effect of the macroalgal species on the abundance of cells of Ostreopsis spp. The variable tested was cells per gram of macroalgal FW in Rochambeau and Vernazzola, with (i) macroalgal species (five levels) as a fixed factor, and (ii) the sampling day (5 levels in Rochambeau and 6 in Vernazzola) as random.

Abundances of Ostreopsis spp. on artificial macroalgae (microhabitat scale): A GLMM with a Gamma link log distribution function was used to test the effect of the macroalgal morphology on the abundance of cells of Ostreopsis spp. The variable was cells per surface (cm 2 ) in Rochambeau and Vernazzola with (i) macroalgal species (five levels) as a fixed factor, and (ii) the sampling day (5 levels) and the replicate as random to account for repeated measures over time.

Abundances of Ostreopsis spp. on macroalgal communities (mesohabitat scale): A GLMM with a Gamma link log distribution function was used to test the effect of the benthic macroalgal community on the abundance of cells of Ostreopsis spp. The tested variable were cells in 0.04 m 2 of sea bottom using the adapted BEDI device in Rochambeau and cells in 0.04 m 2 of sea bottom extrapolated from the biomass of macroalgae obtained from the scrapings in Rochambeau and Vernazzola, with (i) the macroalgal community type (five levels) as a fixed factor, and (ii) the sampling day (6 levels) and the replicate as random to account for repeated measures over time.

All GLMM were fitted to analyse the effect of the variables and the AICs likelihood minimum was used to select the best model among the possible combinations. The different models were fitted using the functions "glmer" and "lm" from the package lme4 [START_REF] Bates | Fitting Linear Mixed-Effects Models using lme4[END_REF] in the statistical environment R in the statistical environment R (R: The R Project for Statistical Computing). Pvalues were obtained by means of a Wald χ 2 test using the "ANOVA" function from the CAR package [START_REF] Fox | An R Companion to Applied Regression[END_REF]. Finally, the function "emmeans" from the package emmeans [START_REF] Lenth | emmeans: Estimated Marginal Means, aka Least-Squares Means[END_REF] was used to perform the post-hoc analysis of the GLMM models.

RESULTS

The mean seawater temperature was 24.99 ± 1.25°C and 26.77 ± 1.68°C (mean ± SD), respectively in Rochambeau and Vernazzola, during the duration of the experiment, with maximum and minimum temperatures of 28.27°C (on the 30 th of July 2021) and 21.49°C (on the 2 nd July 2021) in Rochambeau and of 30.54°C (on the 15 th of August 2021) and 23.59°C (on the 28 th of June 2021) in Vernazzola.

Ostreopsis spp. concentrations in seawater

Concentrations of Ostreopsis spp. in the water in Rochambeau (n = 4) situate the peak of the bloom on 15/07 (day 3, mean cell abundance of 5.25 × 10 4 Ostreopsis spp. cells / L), reaching a maximum concentration of 7.90 × 10 4 Ostreopsis spp. cells / L (the maximum cell concentration in seawater measured in this study; Figure 2A). In Vernazzola, the seawater samples of Ostreopsis spp. (n = 2)

show the peak of the bloom on day 3 (21/07, mean cell abundance of 8.47 × 10 3 Ostreopsis spp. cells / L), reaching a maximum concentration of 1.26 × 10 4 Ostreopsis spp. cells / L (Figure 2B).

According to the definition of bloom [START_REF] Funari | Ostreospis cf. ovata blooms in coastal water: Italian guidelines to assess and manage the risk associated to bathing waters and recreational activities[END_REF]Mangialajo et al., 2017), the seawater samples from Rochambeau reached concentrations corresponding to a state of alert (>30 × 10 3 cells / L) the sampling days 1 (07/07), 3 (15/07) and 4 (21/07), while seawater abundances of Ostreopsis spp. in Vernazzola never surpassed the values of routine monitoring.

Abundances of Ostreopsis spp. on natural macroalgae (microhabitat scale)

The ANOVA from the GLMM model did not find differences in abundances of Ostreopsis spp.

among the different macroalgal species sampled in Rochambeau (Table 2). For this sampling site, the peak of the benthic bloom on natural macroalgae was on day 2 (11/07, mean cell abundance of 2.06 × 10 6 cells / g FW), with maximum benthic abundances of Ostreopsis spp. reaching 4.00 × 10 6 cells / g FW (the maximum benthonic cell abundances measured in this study) on turf (day 2, 11/07; Figure 3A). Major bloom abundances were reached on days 1 (07/07), 2 (11/07) and 3 (15/07; on 35 out of 83 total samples), and bloom abundances on days 1 (07/07), 2 (11/07), and 5 (05/08; on 29 out of 83 total samples). Abundances of no bloom on macroalgae were recorded on days 4 (21/07) and 5 (05/08; on 19 out of 83 total samples). In Vernazzola, significant differences on abundances of Ostreopsis spp. were found among macroalgal species (Figure 3B and Table 2). Significantly higher abundances of Ostreopsis spp.

were found on Dictyota fasciola than in Sargassum vulgare, Cystoseira compressa and turf.

Halopteris scoparia hosted significantly higher abundances of Ostreopsis spp. than C. compressa (Figure 3B). For this sampling site, the peak of the benthic bloom on natural macroalgae was on day 3 (21/07, mean abundance of 1.03 × 10 6 cells / g FW), and the maximum abundances were observed on D. fasciola. (3.76 × 10 6 cells / g FW; Figure 3B). In Vernazzola major bloom and bloom abundances [START_REF] Funari | Ostreospis cf. ovata blooms in coastal water: Italian guidelines to assess and manage the risk associated to bathing waters and recreational activities[END_REF]Mangialajo et al., 2017) were reached on days 2 (12/07), 3 (21/07), 4 (30/07), 5 (11/08) and 6 (01/09; on 28 and 49 out of 97 total samples for the major bloom and the bloom scenarios respectively), and no bloom abundances on days 1 (23/06), 4 (30/07) and 5 ( 11/08; on 20 out of 97 total samples). However, significant differences among types of artificial macroalgae were found in Vernazzola, where the control hosted lower abundances of Ostreopsis spp. than the artificial algal types Cystoseira, Dictyota and Turf (Figure 5B and Table 2). In this sampling location, abundances of Ostreopsis spp. on artificial macroalgae remain high on the two first sampling days (12/07 and 21/07) when they start decreasing (Figure 5B). Maximum abundances on artificial macroalgae in Vernazzola were lower than in Rochambeau, reaching only 2.33 × 10 4 cells / cm 2 on turf on day 1 (12/07), the highest mean abundances were recorded on day 2 (mean abundance of 32/07; 8.54 × 10 3 cells / cm 2 ).

Abundances of Ostreopsis spp. on macroalgal communities (mesohabitat scale)

The ANOVA from the GLMM model did not find significant differences in abundances of Ostreopsis spp. among the different macroalgal communities sampled with the adapted BEDI device in Rochambeau (Table 2). The peak of the bloom on natural communities (mesohabitat scale) in Rochambeau was day 3 (15/07, mean abundance of 3.98 × 10 6 cells / 0.04 m 2 ), and after this day the abundances started decreasing (Figure 5). The maximum abundances measured reached 8.95 × 10 6 cells / 0.04 m 2 on the turf community on day 4 (21/07). Ostreopsis spp. abundances on

94

natural communities in Rochambeau reached values corresponding to a major bloom (> 1319 cells / cm 2 ) on the sampling days 1 (07/07), 2 (11/07), 3 (15/07) and 4 (21/07), globally on 76 out of 100 total samples. Bloom values (> 279 cells/ cm 2 ) were reached on days 4 (21/07) and 5 (05/08), and globally on 13 out of 100 total samples (for major bloom and bloom definition, see Mangialajo et al., 2017). Regarding the cells of Ostreopsis spp. on 0.04 m 2 of sea bottom (mesohabitat scale), extrapolated from the total biomass of macroalgae obtained from the scrapings, significant differences were found among the different communities sampled in both Rochambeau and Vernazzola (Table 2).

In Rochambeau, the abundances of cells was significantly higher on Halopteris scoparia than on Gongolaria barbata dominated communities, without differences among the other species (Figure 6A). The highest mean abundances of cells of were recorded the day 2 (11/07; mean abundance of 1.90 x 10 8 cells / 0.04 m 2 ). In Vernazzola significantly higher abundances were found on Dictyota spp. dominated communities than on all the other communities (Figure 6B). The highest mean abundances were recorded the day 3 (21/07; mean abundance of 1.45 x 10 8 cells / 0.04 m 2 ).

DISCUSSION

The objective of this study was to expand our understanding of the role that macroalgal species (microhabitat) and/or communities (mesohabitat) can play facilitating blooms of Ostreopsis spp.

To do so, samples of Ostreopsis spp. were taken at different spatial scales and on different macroalgal species and communities, using different sampling techniques and methodologies to quantify abundances of Ostreopsis spp.

The maximum concentrations of cells of Ostreopsis spp. in the seawater observed in this study are comparable with the ones reported in Rochambeau (10.4 × 10 4 cells / L), and the ones reported in several sites in Genova, including Vernazzola (8.4 × 10 4 cells / L; Mangialajo et al., 2008aMangialajo et al., , 2011)).

On both sampling sites, the number of cells of Ostreopsis spp. on natural macroalgae (cells / g FW) reached abundances characteristic of a major bloom [START_REF] Funari | Ostreospis cf. ovata blooms in coastal water: Italian guidelines to assess and manage the risk associated to bathing waters and recreational activities[END_REF]Mangialajo et al., 2017) and are comparable with the maximum abundances reported in other studies in or near the same locations. In Rochambeau in July 2008 one of the maximal epiphytic abundances reached 8.54 × 10 6 cells / g FW on Dictyota spp. (Cohu et al., 2013)

96

The present study highlights the high variability of Ostreopsis spp. preferences when using different sampling techniques to assess the abundances of cells. At the microhabitat scale, in Rochambeau abundances of cells of Ostreopsis spp. per gram FW of macroalgae were not significantly different for the sampled species. Contrarily, significant patterns were observed in Vernazzola, where the cells of Ostreopsis spp. per gram FW of macroalgae were higher on Dictyota fasciola and Halopteris scoparia than on most of the other sampled macroalgal species. Such results are in agreement with our hypotheses that higher abundances are found on shrub-forming macroalgae (D.

fasciola and H. scoparia) than on forest-forming (Cystoseira compressa and Sargassum vulgare; Meroni et al., 2018;Yong et al., 2018;Bravo et al., 2020;Lee et al., 2020). Some recent studies already highlighted the importance of the macrophyte morphology on the abundance of dinoflagellates (Vila et al., 2001;Parsons and Preskitt, 2007;Totti et al., 2010), and the need for standardized methods allowing direct comparisons, as some of the ones used in the present study (i.e. cells per surface of artificial macroalgae or cells per surface of sea bottom). Our study did not highlight significant differences among the different artificial macroalgae. In Vernazzola the control was hosting a significantly lower number of cells in 3 out of the 4 treatments (except Halopteris), while in Rochambeau even the control was not significantly different from the other treatments. Therefore, highlighting that the considered different morphologies of artificial macroalgae are not influencing Ostreopsis spp. abundances (no differences among treatments); that allelopathic interactions may be at the origin of the patterns observed; and that the small scale variability can be very high (no significant differences with the control in Rochambeau). The maximum abundances of Ostreopsis spp. recorded on the artificial macroalgae were found in Rochambeau, and were of the same order of magnitude of the abundances observed on natural macroalgae (Totti et al., 2010) in the Adriatic Sea and higher than the abundances reported on other artificial substrates in the N-W Mediterranean Sea (Fricke et al., 2018).

At the mesohabitat scale, no differences in Ostreopsis spp. per cm 2 of sea bottom were found among the sampled macroalgal communities using the adapted BEDI device in Rochambeau. This result can be explained by the high variability in cell abundances observed in this location. When extrapolating the abundances of cells of Ostreopsis spp. per surface of sea bottom using the total biomass of macroalgae obtained from the scrapings, significant differences at the mesohabitat scale among communities were found both in Rochambeau (Halopteris scoparia higher than Gongolaria barbata) and in Vernazzola (Dictyota spp. higher than all the other communities). Yet, while no differences among communities were highlighted by the cell's abundances obtained with the BEDI device, the extrapolation of the same variable using the total biomass of macroalgae obtained from the scrapings allowed to observe higher abundances on a shrub-forming species (H. scoparia) than on a forest-forming (G. barbata) in agreement with our hypothesis. Nevertheless, such results have to be taken with care as the abundances on the other considered communities were not significantly different. We cannot exclude that such results could be due to a high variability associated to the BEDI sampling, that can be difficult to apply on irregular sea bottoms and that integrates the cells of Ostreopsis spp. on the macroalgal community and the cells present in the water column, potentially confusing the absolute values of the cells in the macroalgal biofilm, especially in the situations of "bloom" or "major bloom". Nevertheless, our calculations suggest that the mean cells of Ostreopsis spp. sampled in seawater, represent less than the 10 % of the mean cells in the macroalgal biofilm. Concerning the transplanted communities, we observed that the C. compressa and G. barbata dominated communities moved to Rochambeau from Sainte Marguerite Island lost about a 30 % of their biomass by the end of the experiment, possibly reducing the facilitating or deterrent effects of these communities on Ostreopsis spp. proliferation. The loss of biomass of the transplanted forest-forming communities was partially due to fish grazing, but sign of degradation were also observed, potentially due to unfavourable environmental conditions in Rochambeau or to the effects of the bloom, that have been suggested to negatively impact other Cystoseira s.l. spp. [START_REF] Jauzein | Uptake of dissolved inorganic and organic nitrogen by the benthic toxic dinoflagellate Ostreopsis cf. ovata[END_REF][START_REF] Iveša | Increased ammonium levels occurring during benthic algal blooms are potentially toxic to fucalean algae[END_REF].

In Vernazzola, patterns were more clear, observing higher abundances of cells at the mesohabitat scale on Dictyota spp. dominated communities, in agreement with the samplings at the microhabitat scale (cells / g FW macroalgae) and with other studies on other locations in the Mediterranean Sea [START_REF] Blanfuné | Distribution and abundance of Ostreopsis spp. and associated species (Dinophyceae) in the northwestern Mediterranean: the region and the macroalgal substrate matter[END_REF]González et al., 2019;Gémin et al., 2020;Ternon et al., 2020). It is worth noting that one of the maximal epiphytic abundance ever reported for Ostreopsis cf. ovata in the N-W Mediterranean was found on species belonging to this genus (Cohu et al., 2013). Nonetheless, studies in mesocosm found that Dictyota spp. could inhibit the growth of Ostreopsis cf. ovata (Accoroni et al., 2015;Ternon et al., 2020). These differences among studies in the field and in laboratory could be due to the changes in macroalgal metabolites when exposed to culture conditions or because of the lack of other effects at larger scale that could modify the preferences of Ostreopsis spp. [START_REF] Mancuso | Diversity and Temporal Dynamics of the Epiphytic Bacterial Communities Associated with the Canopy-Forming Seaweed Cystoseira compressa (Esper) Gerloff and Nizamuddin[END_REF]. Moreover, older identifications of Dictyota spp. as substrate for the development of Ostreopsis spp. should be reviewed as this genus has suffered taxonomic changes [START_REF] Tronholm | Species Delimitation, Taxonomy, and Biogeography of Dictyota in Europe (dictyotales, Phaeophyceae)1[END_REF].

Overall, the results obtained extrapolating the Ostreopsis spp. cells in 0.04 m 2 of sea bottom from the cells per gram FW of macroalgae are in agreement with the results from other studies reporting higher abundances of cells of harmful benthic dinoflagellates in shrubs and turfs (Meroni et al., 2018b;Yong et al., 2018;Bravo et al., 2020) and from a recent review [START_REF] Monserrat | The role of habitat in the facilitation of Ostreopsis spp. blooms[END_REF]. Some studies (Yong et al., 2018;Lee et al., 2020) highlight the importance of the mesohabitat for the proliferation of BHAB and developed methodologies (artificial substrates) to allow the comparison between different macroalgal communities. Unfortunately, studies using either artificial substrates or quantification of cells per surface of sea bottom (using the BEDI device or CHAPTER 4 ………………….…………. 98 extrapolating from macroalgal biomass) are scarce and do not allow us to compare the results obtained in this study.

The comparison of cell abundances between different macroalgal species from the literature is controversial, especially due to differences on macrophyte morphologies and surfaces (Totti et al., 2010;Mangialajo et al., 2017). Our results, as the ones reported in other studies (Totti et al., 2010;[START_REF] Blanfuné | Distribution and abundance of Ostreopsis spp. and associated species (Dinophyceae) in the northwestern Mediterranean: the region and the macroalgal substrate matter[END_REF]Meroni et al., 2018b;Bravo et al., 2020) show an important variability in the abundance of Ostreopsis spp. in function of the sampled macroalgal species and communities, sometimes not allowing to find clear patterns of preference for Ostreopsis spp. [START_REF] Berdalet | Harmful Algal Blooms in Benthic Systems Recent Progress and Future Research[END_REF]Jauzein et al., 2018;[START_REF] Tester | Using artificial substrates to quantify Gambierdiscus and other toxic benthic dinoflagellates for monitoring purposes[END_REF] as in the case of Rochambeau. The abundance of cells per area of sea bottom could be a measure better estimating the public health risk of blooms, as it represents the total number of cells in a determinate area (Mangialajo et al., 2017). The methodologies and standardisations better representing the reality of the ecosystem (e.g. real macroalgal biomass and area available for the settlement of Ostreopsis spp. per sea bottom surface) should be further studied and, if possible, preferred [START_REF] Tester | Using artificial substrates to quantify Gambierdiscus and other toxic benthic dinoflagellates for monitoring purposes[END_REF].

In general, the results obtained in this study highlight the wide and highly variable distribution of Ostreopsis spp. on all macroalgal species and communities sampled. Nevertheless, the fact of not having found in some cases clear preferences on cell abundances among natural or artificial macroalgae could be due to the high abundances of Ostreopsis spp. during the bloom that could homogeneously colonize all the available substrates. This could be the case of Rochambeau, the sampling location where higher abundances of Ostreopsis spp. were observed in this study, and where the preferences of Ostreopsis spp. were less clear. In this site, the variability on Ostreopsis spp. abundances could also be due to the characteristic morphology of the coast, consisting in a small bay with different expositions affecting in different ways important environmental factors such as hydrodynamics and light. On the opposite, in Vernazzola the coast is more homogenous and the whole sampling location is equally exposed, possibly leading to smaller variability in cell abundances (Figure 1). Furthermore, Ostreopsis spp. abundance variability in Rochambeau could be spatial and temporal, as the exposition of the coast can condition the sooner or later development of the bloom (Chang et al., 2000;Shears and Ross, 2009b;Accoroni and Totti, 2016;Fricke et al., 2018).

Our results highlight that preferences of Ostreopsis spp. can be species-, site-and period of the bloom-specific. Effects at a larger scale (i.e. at the macrohabitat scale, not assessed in this study) could in turn directly influence Ostreopsis spp. abundances or have indirect effects influencing the dominance of specific macroalgal communities that could facilitate or regulate Ostreopsis spp. blooms (Mangialajo et al., 2008b). Results from this study show how variable Ostreopsis spp. cell assessment can be in function of the scale (micro and mesohabitat) and of the quantification method CHAPTER 4 ……………………………… 99 used, opening new research topics such as the study of blooms at a bigger scale (macrohabitat) and testing different methods for the standardization of the cells independently on the substrate. Some patterns of substrate's preferences for Ostreopsis spp. proliferation have been observed, indicating the role that benthic communities can potentially play in the frequency and magnitude of blooms.

The effect of the dominant macroalgal communities has to be further investigated in order to: (i) mitigate BHABs on current and future regime shift scenarios, where the most complex forestforming macroalgal communities, possibly hosting fewer abundances of benthic toxic dinoflagellates, are disappearing [START_REF] Rindi | Long-term changes in the benthic macroalgal flora of a coastal area affected by urban impacts (Conero Riviera, Mediterranean Sea)[END_REF][START_REF] Pessarrodona | Homogenization and miniaturization of habitat structure in temperate marine forests[END_REF]; (ii) better assess locations with high public health risk of BHABs and (iii) estimate priority locations for possible actions of mitigation (remediation and/or restoration).

GENERAL DISCUSSION AND CONCLUSIONS

The aim of this dissertation is to unravel some of the effects of global change on shallow Cystoseira s.l. forests, using a cosmopolitan habitat-forming species, Cystoseira compressa, as a model species and to assess if the proliferation of blooms of Ostreopsis spp. could be a consequence of Cystoseira s.l. forests loss. Considering that in each chapter a specific discussion was addressed, we present here the main findings of the experiments focussing on some causes involved in the loss of Cystoseira s.l. forests and the consequences, such as the proliferation of benthic harmful algal blooms (BHAB) of Ostreopsis spp. In this section, we provide a discussion on the implications of the loss of coastal Mediterranean marine forests and the expansion of Ostreopsis spp. in the conservation of shallow coastal habitats, as well as a view of the application and contribution of the thesis to coastal conservation, management and restoration.

THE DECLINE OF MARINE FORESTS AND REGIME SHIFTS

Forest-forming brown macroalgae, covering ~28% of the coastline worldwide have been valued at over 10 6 USD km -1 year -1 (Filbee-Dexter and [START_REF] Filbee-Dexter | Rise of Turfs: A New Battlefront for Globally Declining Kelp Forests[END_REF] and their conservation and restoration is clearly justified by the importance of these species in hosting high biodiversity, providing several ecosystem services and being highly productive (Filbee-Dexter and [START_REF] Filbee-Dexter | Rise of Turfs: A New Battlefront for Globally Declining Kelp Forests[END_REF]Wernberg et al., 2019a;[START_REF] Pessarrodona | Global seaweed productivity[END_REF]. It is then critical to understand the factors promoting the decline of forest-forming brown macroalgae and its consequences for ecosystem functioning, such as fostering harmful algal blooms. Therefore, we investigated different ecological interactions, such as species facilitation (Chapters 1, 3 and 4), and herbivory (Chapter 2), which are key factors in structuring communities and ecosystems [START_REF] Stachowicz | Mutualism, Facilitation, and the Structure of Ecological Communities: Positive interactions play a critical, but underappreciated, role in ecological communities by reducing physical or biotic stresses in existing habitats and by creating new habitats on which many species depend[END_REF][START_REF] Kordas | Community ecology in a warming world: The influence of temperature on interspecific interactions in marine systems[END_REF][START_REF] Kroeker | Ocean acidification causes ecosystem shifts via altered competitive interactions[END_REF].

Microalgae and macroalgae are the base of marine trophic nets and any perturbation in their populations can have an enormous effect on the whole ecosystem [START_REF] Cardona | Pulses of phytoplanktonic productivity may enhance sea urchin abundance and induce state shifts in Mediterranean rocky reefs[END_REF]Smale et al., 2022). Human impacts on forest-forming macroalgae are expected to continue or increase in the future [START_REF] Halpern | Evaluating and Ranking the Vulnerability of Global Marine Ecosystems to Anthropogenic Threats[END_REF][START_REF] Halpern | Recent pace of change in human impact on the world's ocean[END_REF], potentially leading to changes in the macroalgal composition and its structural complexity (i.e. regime shifts). The substitution of long-lived large brown forestforming species, including Cystoseira s.l. spp., by fast-growing opportunistic turf-forming macroalgae and even barren grounds [START_REF] Chemello | Regime shifts and alternative stable states in intertidal rocky habitats: State of the art and new trends of research[END_REF][START_REF] Filbee-Dexter | Rise of Turfs: A New Battlefront for Globally Declining Kelp Forests[END_REF][START_REF] O'brien | Turf wars: competition between foundation and turfforming species on temperate and tropical reefs and its role in regime shifts[END_REF] have already been reported along hundreds of kilometres along Canadian, European, and Australian coasts (Filbee-Dexter and [START_REF] Filbee-Dexter | Rise of Turfs: A New Battlefront for Globally Declining Kelp Forests[END_REF]. Regime shifts have also been observed for Cystoseira s.l. spp. in several areas of the Mediterranean Sea. It is expected that only the most tolerant populations or species will be able to resist under future global change scenarios [START_REF] Thibaut | Long-term decline of the populations of Fucales (Cystoseira spp. and Sargassum spp.) in the Albères coast (France, Northwestern Mediterranean)[END_REF][START_REF] Thibaut | Decline and local extinction of Fucales in French Riviera: the harbinger of future extinctions?[END_REF][START_REF] Blanfuné | The fate of Cystoseira crinita, a forest-forming Fucale (Phaeophyceae, Stramenopiles), in France (North Western Mediterranean Sea)[END_REF][START_REF] Rindi | Long-term changes in the benthic macroalgal flora of a coastal area affected by urban impacts (Conero Riviera, Mediterranean Sea)[END_REF]Orlando-Bonaca et al., 2021a). For example, in the Mediterranean Sea, [START_REF] Thibaut | Long-term decline of the populations of Fucales (Cystoseira spp. and Sargassum spp.) in the Albères coast (France, Northwestern Mediterranean)[END_REF](2015) report the high regression and fragmentation of Cystoseira s.l. populations in the French Riviera and Alberes Coast (North Western Mediterranean), where only five of the fifteen historically present species were still thriving and only C. compressa presented no signs of regression. The causes responsible for regime shifts and miniaturisation of habitats are multiple and ultimately related to anthropogenic activities [START_REF] Benedetti-Cecchi | Predicting the consequences of anthropogenic disturbance: large-scale effects of loss of canopy algae on rocky shores[END_REF]Mangialajo et al., 2008b;Sala et al., 2012;[START_REF] Chemello | Regime shifts and alternative stable states in intertidal rocky habitats: State of the art and new trends of research[END_REF]. Some of the abiotic and biotic stressors related to marine forests decline have been addressed in this thesis, using the Mediterranean widely distributed Cystoseira compressa as a model for forest-forming macroalgae. C. compressa, independently of being the only non-protected Cystoseira s.l. spp. [START_REF] Verlaque | Mediterranean seaweeds listed as threatened under the Barcelona Convention: A critical analysis[END_REF], is considered very tolerant to manipulation and interesting in the restoration of Mediterranean marine forests due to its potential facilitation role in the recruitment of other forest-forming species [START_REF] Mangialajo | Zonation patterns and interspecific relationships of fucoids in microtidal environments[END_REF].

ABIOTIC INTERACTIONS ON MARINE FORESTS: CLIMATE CHANGE

Abiotic factors such as habitat destruction, local pollution, eutrophication and overfishing contribute to the loss of marine forests [START_REF] Connell | The other ocean acidification problem: CO2 as a resource among competitors for ecosystem dominance[END_REF][START_REF] De Caralt | Differential effects of pollution on adult and recruits of a canopy-forming alga: implications for population viability under low pollutant levels[END_REF][START_REF] Smale | Impacts of ocean warming on kelp forest ecosystems[END_REF][START_REF] Verdura | Localscale climatic refugia offer sanctuary for a habitat-forming species during a marine heatwaves[END_REF], but climate change is expected to be the ultimate impact that all species around the world will need to overcome, especially macroalgae [START_REF] Doney | Climate Change Impacts on Marine Ecosystems[END_REF][START_REF] Harvey | Meta-analysis reveals complex marine biological responses to the interactive effects of ocean acidification and warming[END_REF][START_REF] Smale | Impacts of ocean warming on kelp forest ecosystems[END_REF].

Ocean warming

Ocean warming (OW) is expected to affect forest-forming macroalgae around the globe, including Cystoseira s.l. spp. [START_REF] Wernberg | Climate-driven regime shift of a temperate marine ecosystem[END_REF][START_REF] Capdevila | Warming impacts on early life stages increase the vulnerability and delay the population recovery of a long-lived habitat-forming macroalga[END_REF][START_REF] Smale | Impacts of ocean warming on kelp forest ecosystems[END_REF]. Several studies in the Mediterranean Sea already linked the increase in seawater temperatures with (i) perturbations in the phenology of Cystoseira s.l. [START_REF] Bevilacqua | Climatic anomalies may create a long-lasting ecological phase shift by altering the reproduction of a foundation species[END_REF], (ii) a decrease in the resilience of adults [START_REF] Capdevila | Warming impacts on early life stages increase the vulnerability and delay the population recovery of a long-lived habitat-forming macroalga[END_REF], and, in agreement with the results presented in this thesis, (iii) a higher mortality of early-life stages in the laboratory and in the field [START_REF] Falace | Ex situ cultivation protocol for Cystoseira amentacea var. stricta (Fucales, Phaeophyceae) from a restoration perspective[END_REF][START_REF] Falace | Is the South-Mediterranean Canopy-Forming Ericaria giacconei (= Cystoseira hyblaea) a Loser From Ocean Warming? Front[END_REF][START_REF] Capdevila | Warming impacts on early life stages increase the vulnerability and delay the population recovery of a long-lived habitat-forming macroalga[END_REF][START_REF] Verdura | Localscale climatic refugia offer sanctuary for a habitat-forming species during a marine heatwaves[END_REF]. Because early-life stages are more sensitive than adults to any change in environmental conditions [START_REF] Coelho | Impacts of anthropogenic stresses on the early development stages of seaweeds[END_REF][START_REF] De Caralt | Differential effects of pollution on adult and recruits of a canopy-forming alga: implications for population viability under low pollutant levels[END_REF][START_REF] Verdura | Localscale climatic refugia offer sanctuary for a habitat-forming species during a marine heatwaves[END_REF], and present high mortality rates [START_REF] Chapman | Functional ecology of fucoid algae: twenty-three years of progress[END_REF], they are considered as a bottleneck for the continuity of the populations. Therefore, further studies on the recruits of these species are needed for the conservation and restoration of marine forests of Cystoseira s.l. spp.

In our experiment, the warmest temperature (32°C) had a negative effect on the density of recruits since the start of the experiment, while differences in the density of recruits at 28°C and 24°C were only observed after one month and a half. However, no differences in the size of recruits relative to the different temperature treatments were observed until the third month, potentially indicating that the temperature was not affecting the growth of the surviving individuals [START_REF] Celis-Plá | Macroalgal responses to ocean acidification depend on nutrient and light levels[END_REF], 2017a;[START_REF] Mancuso | Influence of ambient temperature on the photosynthetic activity and phenolic content of the intertidal Cystoseira compressa along the Italian coastline[END_REF]. These results on C. compressa show how recruits of this species could be more tolerant and resistant to climate change, and thus more widely distributed, than early-life stages of other Cystoseria s.l. spp. [START_REF] Bennett | Thermal Performance of Seaweeds and Seagrasses Across a Regional Climate Gradient[END_REF]. For example, Ericaria crinita, drastically decreased in density after 5 days in experimental tanks at 28°C [START_REF] Verdura | Localscale climatic refugia offer sanctuary for a habitat-forming species during a marine heatwaves[END_REF] or Ericaria giacconei, which zygotes did not survived at 28°C in the laboratory [START_REF] Falace | Is the South-Mediterranean Canopy-Forming Ericaria giacconei (= Cystoseira hyblaea) a Loser From Ocean Warming? Front[END_REF].

The contrasting responses against OW of different Cystoseira s.l. spp. manifest that not all the Cystoseira s.l. spp. are ecological analogues [START_REF] Bruno De Sousa | Improved phylogeny of brown algae Cystoseira (Fucales) from the Atlantic-Mediterranean region based on mitochondrial sequences[END_REF][START_REF] Orellana | Diversity of Cystoseira sensu lato (Fucales, Phaeophyceae) in the eastern Atlantic and Mediterranean based on morphological and DNA evidence, including Carpodesmia gen. emend. and Treptacantha gen. emend[END_REF]. In general, the negative effects of OW have already been observed on other forest-forming species, both on recruits [START_REF] Leal | Copper pollution exacerbates the effects of ocean acidification and warming on kelp microscopic early life stages[END_REF][START_REF] Capdevila | Warming impacts on early life stages increase the vulnerability and delay the population recovery of a long-lived habitat-forming macroalga[END_REF][START_REF] Verdura | Localscale climatic refugia offer sanctuary for a habitat-forming species during a marine heatwaves[END_REF] and adults [START_REF] Lind | Effects of abiotic stressors on kelp early life-history stages[END_REF][START_REF] Fernández | Nitrogen sufficiency enhances thermal tolerance in habitat-forming kelp: implications for acclimation under thermal stress[END_REF][START_REF] Smale | Impacts of ocean warming on kelp forest ecosystems[END_REF][START_REF] Falace | Is the South-Mediterranean Canopy-Forming Ericaria giacconei (= Cystoseira hyblaea) a Loser From Ocean Warming? Front[END_REF].

Specific studies on the effect of OW on adults of C. compressa in the Mediterranean Sea have also been performed. In a mesocosm experiment, [START_REF] Bennett | Thermal Performance of Seaweeds and Seagrasses Across a Regional Climate Gradient[END_REF] found the maximum gross primary production of C. compressa from Cyprus and Mallorca (Balearic Islands) at 28 ± 4 °C, and a thermal limit where the net production become negative at 23 ± 0.5 °C. The results reported by [START_REF] Bennett | Thermal Performance of Seaweeds and Seagrasses Across a Regional Climate Gradient[END_REF] on adult C. compressa, presumably less sensitive than recruits, are comparable with the mortality we observed in the tanks at similar temperatures. Our results in the treatments at warmer temperatures (28 °C and 32°C) are also in concordance with the results reported by [START_REF] Mancuso | Influence of ambient temperature on the photosynthetic activity and phenolic content of the intertidal Cystoseira compressa along the Italian coastline[END_REF] who observed a decrease in the photosynthetic activity of C. compressa from 28°C. Moreover, C. compressa shows a reduction in net production [START_REF] Bennett | Thermal Performance of Seaweeds and Seagrasses Across a Regional Climate Gradient[END_REF] and an increase in phenolic contents [START_REF] Mancuso | Influence of ambient temperature on the photosynthetic activity and phenolic content of the intertidal Cystoseira compressa along the Italian coastline[END_REF] with the increment in temperature. The impacts of OW and extreme climatic events such as marine heat waves have also been claimed as responsible for the decline and local extinction of Cystoseira s.l. populations in the Mediterranean Sea [START_REF] Mariani | Past and present of Fucales from shallow and sheltered shores in Catalonia[END_REF][START_REF] Verdura | Localscale climatic refugia offer sanctuary for a habitat-forming species during a marine heatwaves[END_REF]. In fact, OW can, in some cases, limit populations to specific refuge locations with more favourable conditions [START_REF] Smale | Impacts of ocean warming on kelp forest ecosystems[END_REF][START_REF] Verdura | Localscale climatic refugia offer sanctuary for a habitat-forming species during a marine heatwaves[END_REF].

Finally, indirect temperature-driven effects, such as the range expansion of warm water, fastgrowing or opportunistic species, on marine forests are not negligible, because they contribute to the tropicalization of habitats and can re-shape macroalgal communities (Vergés et al., 2014a(Vergés et al., , 2014b)).

Ocean acidification

Ocean acidification (OA) due to increasing anthropogenically-derived atmospheric CO2 levels is leading to an increase in dissolved inorganic carbon (DIC), a decrease in pH, CO3 2-and carbonate saturation state, having adverse consequences for marine calcifiers, such as crustose coralline algae, and probably affecting the organisms' cellular homeostasis [START_REF] Hurd | Ocean acidification as a multiple driver: how interactions between changing seawater carbonate parameters affect marine life[END_REF]. Nevertheless, the increase in CO2 and DIC levels can be beneficial for some non-calcifying macroalgae (Connell et al., 2013;Cornwall et al., 2017a). For instance, OA could stimulate photosynthesis [START_REF] Connell | The other ocean acidification problem: CO2 as a resource among competitors for ecosystem dominance[END_REF][START_REF] Fernández | Effects of ocean acidification on the photosynthetic performance, carbonic anhydrase activity and growth of the giant kelp Macrocystis pyrifera[END_REF][START_REF] Rautenberger | Saturating light and not increased carbon dioxide under ocean acidification drives photosynthesis and growth in Ulva rigida (Chlorophyta)[END_REF][START_REF] Zhang | Elevated CO2 concentrations promote growth and photosynthesis of the brown alga Saccharina japonica[END_REF], thus favouring carbonlimited macroalgae and macroalgae with the ability to down-regulate their carbon concentration mechanisms (CCM; Cornwall et al., 2017a;[START_REF] Cornwall | Variability in the benefits of ocean acidification to photosynthetic rates of macroalgae without CO2-concentrating mechanisms[END_REF], altering the dominant species and re-shaping macroalgal communities [START_REF] Hepburn | Diversity of carbon use strategies in a kelp forest community: implications for a high CO2 ocean[END_REF][START_REF] Connell | The other ocean acidification problem: CO2 as a resource among competitors for ecosystem dominance[END_REF][START_REF] Koch | Climate change and ocean acidification effects on seagrasses and marine macroalgae[END_REF][START_REF] Sunday | Ocean acidification can mediate biodiversity shifts by changing biogenic habitat[END_REF].

Besides the evident impact of climate change on forest-forming species around the world, studies on the interactive effects of OW and OA have focused on Laminariales rather than on Fucales [START_REF] Poore | Direct and indirect effects of ocean acidification and warming on a marine plant-herbivore interaction[END_REF][START_REF] Gaitán-Espitia | Interactive effects of elevated temperature and pCO2 on early-life-history stages of the giant kelp Macrocystis pyrifera[END_REF][START_REF] Leal | Copper pollution exacerbates the effects of ocean acidification and warming on kelp microscopic early life stages[END_REF][START_REF] Hollarsmith | Varying reproductive success under ocean warming and acidification across giant kelp (Macrocystis pyrifera) populations[END_REF][START_REF] Fernández | Effect of environmental history on the habitat-forming kelp Macrocystis pyrifera responses to ocean acidification and warming: a physiological and molecular approach[END_REF] and to my knowledge, this study represents the first quantification of the effects of OA on the recruitment of Cystoseira s.l. spp. In the first chapter of this thesis, and as reported in other studies, OA seems to have a less pronounced effect on recruits of C. compressa. than OW.

Experiments on the effects of OA on adults of Cystoseira s.l. spp. (Celis-Plá et al., 2017b, 2017a) showed that OA could be beneficial when there are enough light and nutrients and in moderate temperature ranges. In agreement with our results, the few studies investigating the interactive effects of OW and OA on adult Cystoseira s.l. spp., performed on Ericaria selaginoides by Celis-Plà et al. (2017b, 2017a), found that the negative effects were due to the increase in temperature.

The same authors showed that low pH increased biomass, maximal electron transport rate (ETRmax), polyphenol content and antioxidant activity (EC50) of E. selaginoides in ambient temperature (20°C). They also concluded that ongoing OW and OA can increase photoprotection and photosynthesis of shallow forests of Cystoseira s.l. spp.

In our experiment, lower pH levels positively affected the size of C. compressa, potentially showing a better performance under elevated CO2. Indeed, OA seems to favour the size of Cystoseira s.l.

spp. (Celis-Plá et al., 2017b, 2017a), including C. compressa, even if they are considered CCM species that cannot down regulate their CCM and thus are not expected to be as favoured by OA as other carbon-limited macroalgae or macroalgae that can down regulate their CCM (Cornwall et al., 2017a). In the Mediterranean Sea, surveys of coastal CO2 seeps have repeatedly shown how brown macroalgae, such as Cystoseira s.l. spp., Dictyota spp., Sargassum vulgare and Padina pavonica, proliferate as CO2 levels rise [START_REF] Porzio | Effects of ocean acidification on macroalgal communities[END_REF][START_REF] Baggini | Seasonality Affects Macroalgal Community Response to Increases in pCO2[END_REF]. Moreover, one specific study performed in CO2 vents investigating the effects of OA on C. compressa, reports that in elevated CO2 and shaded conditions, C. compressa had higher carbon content and antioxidant activity, independently of nutrients concentrations [START_REF] Celis-Plá | Macroalgal responses to ocean acidification depend on nutrient and light levels[END_REF].

However, future responses of Cystoseira s.l. spp. to OA may depend on multiple factors, such as temperature, light and nutrient availability [START_REF] Celis-Plá | Short-term ecophysiological and biochemical responses of Cystoseira tamariscifolia and Ellisolandia elongata to environmental changes[END_REF], 2015, 2017b, 2017a). For example, [START_REF] Britton | Ocean acidification reverses the positive effects of seawater pH fluctuations on growth and photosynthesis of the habitat-forming kelp[END_REF] reported that Eklonia radiata, a CCM-enabled species, relies on diffusive CO2 as a carbon source for photosynthesis when exposed to lower daytime pH, but did not show an increase in growth rates, as expected when CCMs are down-regulated. Some studies on the giant kelp Macrocystis pyrifera [START_REF] Roleda | Ocean acidification and seaweed reproduction: increased CO2 ameliorates the negative effect of lowered pH on meiospore germination in the giant kelp Macrocystis pyrifera (Laminariales, Phaeophyceae)[END_REF][START_REF] Leal | Ocean acidification and kelp development: Reduced pH has no negative effects on meiospore germination and gametophyte development of Macrocystis pyrifera and Undaria pinnatifida[END_REF] reported better performance and increased growth rates of the macroalgae under OA treatments, while others did not report any benefit of elevated CO2 on the photosynthesis and growth of the same species [START_REF] Gaitán-Espitia | Interactive effects of elevated temperature and pCO2 on early-life-history stages of the giant kelp Macrocystis pyrifera[END_REF][START_REF] Fernández | Effect of environmental history on the habitat-forming kelp Macrocystis pyrifera responses to ocean acidification and warming: a physiological and molecular approach[END_REF]. This suggests that the benefits of OA on noncalcifying macroalgae depend on multiple drivers or combined effects of multiple environmental factors, like temperature and nutrient availability, [START_REF] Fernández | Effects of ocean acidification on the photosynthetic performance, carbonic anhydrase activity and growth of the giant kelp Macrocystis pyrifera[END_REF] and also on the possible effects of the seawater carbonate system on physiological processes [START_REF] Hurd | Ocean acidification as a multiple driver: how interactions between changing seawater carbonate parameters affect marine life[END_REF].

In the first chapter, the effects of the low pH on the density of recruits of C. compressa seemed to be negatively stronger during the first experimental days. Similarly, OA showed a negative influence on the settlement and early-life stages of other forest-forming macroalgae such as giant kelp [START_REF] Gaitán-Espitia | Interactive effects of elevated temperature and pCO2 on early-life-history stages of the giant kelp Macrocystis pyrifera[END_REF][START_REF] Hollarsmith | Varying reproductive success under ocean warming and acidification across giant kelp (Macrocystis pyrifera) populations[END_REF], and other key species such as corals and molluscs (Kroeker et al., 2013a). These results together with the magnified effects of OW on recruits of Cystoseira s.l. spp. [START_REF] Capdevila | Warming impacts on early life stages increase the vulnerability and delay the population recovery of a long-lived habitat-forming macroalga[END_REF][START_REF] Falace | Is the South-Mediterranean Canopy-Forming Ericaria giacconei (= Cystoseira hyblaea) a Loser From Ocean Warming? Front[END_REF][START_REF] Verdura | Localscale climatic refugia offer sanctuary for a habitat-forming species during a marine heatwaves[END_REF] has led to the paradigm that early-life stages could be more sensitive to global change and therefore could constitute a bottleneck [START_REF] Chapman | Functional ecology of fucoid algae: twenty-three years of progress[END_REF][START_REF] Coelho | Impacts of anthropogenic stresses on the early development stages of seaweeds[END_REF].

Overall, OA could act both as a stressor and as a resource [START_REF] Connell | The other ocean acidification problem: CO2 as a resource among competitors for ecosystem dominance[END_REF], favouring the replacement of some species by others more resistant and with different functional roles, compromising ecosystem function and stability [START_REF] Kroeker | Ocean acidification causes ecosystem shifts via altered competitive interactions[END_REF][START_REF] Teixidó | Functional biodiversity loss along natural CO2 gradients[END_REF].

Changes in the biomass of macroalgal species can directly affect their consumers [START_REF] Ghedini | Organismal homeostasis buffers the effects of abiotic change on community dynamics[END_REF][START_REF] Nagelkerken | Ocean acidification drives global reshuffling of ecological communities[END_REF], leading to bottom-up driven alterations to the food web [START_REF] Nagelkerken | Trophic pyramids reorganize when food web architecture fails to adjust to ocean change[END_REF][START_REF] Smale | Climate-driven substitution of foundation species causes breakdown of a facilitation cascade with potential implications for higher trophic levels[END_REF]. Under climate change, key species could be replaced by opportunistic species able to survive in disturbed environments (O'Brien and [START_REF] O'brien | Turf wars: competition between foundation and turfforming species on temperate and tropical reefs and its role in regime shifts[END_REF][START_REF] Álvarez-Losada | A regime shift in intertidal assemblages triggered by loss of algal canopies: A multidecadal survey[END_REF] and physiologically less sensitive to the effects of OA [START_REF] Connell | The other ocean acidification problem: CO2 as a resource among competitors for ecosystem dominance[END_REF][START_REF] Wittmann | Sensitivities of extant animal taxa to ocean acidification[END_REF][START_REF] Sunday | Evolution in an acidifying ocean[END_REF].

Our results on OW and OA are striking for C. compressa thriving in rock pools, which are expected to be more acclimated to large variability in environmental factors (i.e. temperature, pH, salinity).

Rockpools can experience strong fluctuations of biotic and abiotic factors due to biological activity, irradiance variation, tides, weather-driven changes in water level, etc [START_REF] Truchot | Oxygen and carbon dioxide in the marine intertidal environment: Diurnal and tidal changes in rockpools[END_REF][START_REF] Morris | Diurnal and seasonal variation in physico-chemical conditions within intertidal rock pools[END_REF][START_REF] Olabarria | Response of macroalgal assemblages from rockpools to climate change: effects of persistent increase in temperature and CO2[END_REF]. C. compressa can express a high plasticity in function of the origin location [START_REF] Bennett | Thermal Performance of Seaweeds and Seagrasses Across a Regional Climate Gradient[END_REF], and our recruits, obtained from a donor population situated in a rockpool, are expected to be adapted to a high range of temperatures, pH and salinity. Populations usually exposed to larger extremes in temperatures, such as our population, could have higher thermal tolerances than populations of the same species not acclimated to such environmental variations, however, the effects of pH variability are less clear to provide a better adaptation to future OA [START_REF] Rivest | The Role of Natural Variability in Shaping the Response of Coral Reef Organisms to Climate Change[END_REF][START_REF] Cornwall | Resistance of corals and coralline algae to ocean acidification: physiological control of calcification under natural pH variability[END_REF][START_REF] Legrand | Impact of ocean acidification and warming on the productivity of a rock pool community[END_REF]. It has already been demonstrated how intraspecific variability of other Cystoseira s.l. spp. to certain environmental parameters such as irradiance allows these species to be better adapted to them (Sant andBallesteros, 2020b, 2021). Nonetheless, in habitats where organisms are near, or at their physiological limits, environmental variability could exacerbate even more the future effects of climate change [START_REF] Lawson | Environmental variation and population responses to global change[END_REF][START_REF] Pansch | A new mesocosm system to study the effects of environmental variability on marine species and communities[END_REF][START_REF] Kroeker | Ecological change in dynamic environments: Accounting for temporal environmental variability in studies of ocean change biology[END_REF]. Therefore, it is very important to know the variability species are dealing with at present as this could affect their responses under climate change scenarios or ongoing marine heat waves.

Assessing the interactive effects of OW and OA on marine forests is, therefore, essential for their conservation and restoration. Next steps to evaluate the responses of Mediterranean marine forests under climate change could be: (i) testing the interactive effects of OW and OA on recruits of other Cystoseira s.l. spp., including deep-sea species, as some species could be more sensitive than others [START_REF] Capdevila | Warming impacts on early life stages increase the vulnerability and delay the population recovery of a long-lived habitat-forming macroalga[END_REF][START_REF] Falace | Is the South-Mediterranean Canopy-Forming Ericaria giacconei (= Cystoseira hyblaea) a Loser From Ocean Warming? Front[END_REF][START_REF] Verdura | Localscale climatic refugia offer sanctuary for a habitat-forming species during a marine heatwaves[END_REF]; (ii) studying the effects of climate change on recruits of the same species form different locations, e.g. from the open coast and from rockpools, as the effects of their environmental story could make them more resistant to variations in environmental factors [START_REF] Fernández | Effect of environmental history on the habitat-forming kelp Macrocystis pyrifera responses to ocean acidification and warming: a physiological and molecular approach[END_REF][START_REF] Sant | The canopy-forming alga Ericaria brachycarpa (J. Agardh) Molinari-Novoa & Guiry (Fucales, Phaeophyceae) shows seasonal and depth adaptation to the incoming light levels[END_REF][START_REF] Verdura | Mediterranean macroalgal forests under threat: The effects of ongoing climate change and design of restoration methods[END_REF][START_REF] Bennett | Thermal Performance of Seaweeds and Seagrasses Across a Regional Climate Gradient[END_REF]; (iii) elucidating the interactive effects of climate change on the development and settlement of the recruits [START_REF] Leal | Ocean acidification and kelp development: Reduced pH has no negative effects on meiospore germination and gametophyte development of Macrocystis pyrifera and Undaria pinnatifida[END_REF][START_REF] Savonitto | Reproductive phenology, zygote embryology and germling development of the threatened Carpodesmia barbatula (= Cystoseira barbatula) (Fucales, Phaeophyta) towards its possible restoration[END_REF]; and (iv) assessing how the photosynthetic activity and metabolome of the recruits change under future climate change scenarios [START_REF] Fernández | Effects of ocean acidification on the photosynthetic performance, carbonic anhydrase activity and growth of the giant kelp Macrocystis pyrifera[END_REF]Celis-Plá et al., 2017a;[START_REF] Mancuso | Influence of ambient temperature on the photosynthetic activity and phenolic content of the intertidal Cystoseira compressa along the Italian coastline[END_REF].

BIOTIC INTERACTIONS ON MARINE FORESTS: SPECIES FACILITATION AND HERBIVORY

Species facilitation

Marine forests are complex and rich ecosystems interacting (i.e. predation, competition, facilitation, allelopathy) with other species of macroalgae, microalgae, vertebrates, and invertebrates. Therefore, in the first chapter we considered it important to evaluate the direct and indirect effects of climate change in presence of other species. Several studies have already investigated the role the canopy has on the maintenance of the understory crustose coralline algae [START_REF] Melville | Experimental effects of kelp canopies on subtidal coralline algae[END_REF][START_REF] Connell | The monopolization of understorey habitat by subtidal encrusting coralline algae: a test of the combined effects of canopy-mediated light and sedimentation[END_REF][START_REF] Irving | Effects of kelp canopies on bleaching and photosynthetic activity of encrusting coralline algae[END_REF][START_REF] Irving | The response of encrusting coralline algae to canopy loss: an independent test of predictions on an Antarctic coast[END_REF][START_REF] Irving | Predicting understorey structure from the presence and composition of canopies: an assembly rule for marine algae[END_REF], however, the role of understory crustose coralline algae on the latter successional forest-forming species is less well understood [START_REF] Benedetti-Cecchi | Effects of canopy cover, herbivores and substratum type on patterns of Cystoseira spp. settlement and recruitment in littoral rockpools[END_REF][START_REF] Asnaghi | Colonisation processes and the role of coralline algae in rocky shore community dynamics[END_REF]. We evaluated the role of one of the most common crustose coralline algae in shallow Mediterranean forests, Neogoniolithon brassica-florida [START_REF] Benedetti-Cecchi | Effects of canopy cover, herbivores and substratum type on patterns of Cystoseira spp. settlement and recruitment in littoral rockpools[END_REF][START_REF] Braga | Coralline algae (Corallinales, Rhodophyta) in western and central Mediterranean Messinian reefs[END_REF], on the recruitment of C. compressa under different OW and OA treatments. Our results show strong effects of OW and OA on the calcification rate of N. brassica-florida which could have cascading effects on later successional species such as C. compressa. From the start of our experiment, C. compressa showed lower densities when associated with the crustose coralline algae, either living or dead. Studies at CO2 seeps [START_REF] Porzio | Effects of ocean acidification on macroalgal communities[END_REF][START_REF] Baggini | Seasonality Affects Macroalgal Community Response to Increases in pCO2[END_REF] have reported a decrease in coralline algae abundance with decreasing pH and CO3 2-, while brown algae, such as Cystoseira s.l. spp., proliferated. This suggests that the disappearance of crustose coralline algae does not affect the maintenance of Cystoseira s.l. spp. Even if some studies [START_REF] Asnaghi | Colonisation processes and the role of coralline algae in rocky shore community dynamics[END_REF] found a facilitation effect of N. brassica-florida on Cystoseria s.l. spp., our experiment showed lower densities of recruits of C. compressa in association with this crustose coralline algae, independently of the treatment. This result potentially relates to the anti-fouling mechanisms of crustose coralline algae [START_REF] Johnson | The crustose coralline alga, Phymatolithon Foslie, inhibits the overgrowth of seaweeds without relying on herbivores[END_REF][START_REF] Keats | Antifouling effects of epithallial shedding in three crustose coralline algae (Rhodophyta, Coralinales) on a coral reef[END_REF][START_REF] Villas Bôas | Are anti-fouling effects in coralline algae species specific?[END_REF] and is of major importance for future restoration actions, because it suggest that natural biotic substrate are not necessarily the best option. Supporting the suitability of the bare substrate, including the widely used artificial clay substrate in the Mediterranean Sea [START_REF] Falace | Ex situ cultivation protocol for Cystoseira amentacea var. stricta (Fucales, Phaeophyceae) from a restoration perspective[END_REF][START_REF] De La Fuente | First ex situ outplanting of the habitat-forming seaweed Cystoseira amentacea var. stricta from a restoration perspective[END_REF] for the settlement of Cystoseira s.l. spp.

Forest-forming macroalgae as refuge from ocean acidification

Forest-forming macroalgae can metabolically induce diel pH fluctuations through photosynthesis and respiration (higher pH during the daytime, lower at night) modifying local seawater chemistry and therefore, potentially acting as a refuge from OA in the vicinity of these habitats for sensitive species such as calcifying organisms [START_REF] Middelboe | High pH in shallow-water macroalgal habitats[END_REF][START_REF] Wootton | Dynamic patterns and ecological impacts of declining ocean pH in a high-resolution multi-year dataset[END_REF][START_REF] Krause-Jensen | Macroalgae contribute to nested mosaics of pH variability in a subarctic fjord[END_REF], 2016). Nevertheless, it is not established if these refuges from OA could still be effective under future OA scenarios [START_REF] Britton | Ocean acidification reverses the positive effects of seawater pH fluctuations on growth and photosynthesis of the habitat-forming kelp[END_REF][START_REF] Murie | Fragmented kelp forest canopies retain their ability to alter local seawater chemistry[END_REF], especially considering the above mentioned negative effects of climate change on marine forests. Fucus vesiculosus can induce diurnal pH fluctuations of about one pH unit [START_REF] Middelboe | High pH in shallow-water macroalgal habitats[END_REF][START_REF] Saderne | Differential Responses of Calcifying and Non-Calcifying Epibionts of a Brown Macroalga to Present-Day and Future Upwelling pCO2[END_REF], but the magnitude of the fluctuations would be inversely proportional to the spatial scale considered, being more important in locations with little water turnover rates such as rockpools [START_REF] Wahl | How good are we at assessing the impact of ocean acidification in coastal systems? Limitations, omissions and strengths of commonly used experimental approaches with special emphasis on the neglected role of fluctuations[END_REF][START_REF] Wahl | Macroalgae may mitigate ocean acidification effects on mussel calcification by increasing pH and its fluctuations[END_REF]. Moreover, pH fluctuations are not temporally and spatially uniform at the community and organismal scales [START_REF] Britton | Ocean acidification reverses the positive effects of seawater pH fluctuations on growth and photosynthesis of the habitat-forming kelp[END_REF]. For example, [START_REF] Houlihan | Diffusive Boundary Layers and Ocean Acidification: Implications for Sea Urchin Settlement and Growth[END_REF] show that an increase in irradiance and thus, photosynthesis, and low hydrodynamics, increase pH and the thickness of the diffusion boundary layer [START_REF] Hurd | Metabolically induced pH fluctuations by some coastal calcifiers exceed projected 22nd century ocean acidification: a mechanism for differential susceptibility?[END_REF][START_REF] Cornwall | Canopy macroalgae influence understorey corallines' metabolic control of near-surface pH and oxygen concentration[END_REF] of crustose coralline algae, affecting an habitat at the dozen of microns to millimetres scale, in which sea urchin larvae, and other species of invertebrates and algae can settle. However, this increase in pH would not perdure during the night due to strong variations in the pH at such a small scale because of respiration processes [START_REF] Hurd | Metabolically induced pH fluctuations by some coastal calcifiers exceed projected 22nd century ocean acidification: a mechanism for differential susceptibility?[END_REF][START_REF] Cornwall | Canopy macroalgae influence understorey corallines' metabolic control of near-surface pH and oxygen concentration[END_REF]. In fact, [START_REF] Cornwall | Diurnal fluctuations in seawater pH influence the response of a calcifying macroalga to ocean acidification[END_REF] report pH fluctuations of 0.94 units within a shallow Macrocystis pyrifera forest demonstrating that diurnal variability in pH was as important as a mean decrease on pH due to OA in controlling the growth rates of understory coralline algae.

Herbivory

Biotic factors such as the proliferation of herbivorous species can be responsible for driving complex systems toward range shifts (Sala et al., 1998, 199;[START_REF] Chemello | Regime shifts and alternative stable states in intertidal rocky habitats: State of the art and new trends of research[END_REF]. In tropical areas, the impact of benthic herbivores such as sea urchins is well documented [START_REF] Lewis | Herbivore abundance and grazing intensity on a Caribbean coral reef[END_REF][START_REF] Foster | The relative impacts of grazing by caribbean coral reef fishes and Diadema: effects of habitat and surge[END_REF][START_REF] Coyer | Interactions between corals and algae on a temperate zone rocky reef: mediation by sea urchins[END_REF][START_REF] Burkepile | Herbivore species richness and feeding complementarity affect community structure and function on a coral reef[END_REF] although omnivorous and herbivorous fishes exert the greatest control on macroalgal abundance and distribution [START_REF] Hiatt | Ecological Relationships of the Fish Fauna on Coral Reefs of the Marshall Islands[END_REF][START_REF] Cronin | Are Tropical Herbivores More Resistant Than Temperate Herbivores to Seaweed Chemical Defenses? Diterpenoid Metobolites from Dictyota acutiloba as Feeding Deterrents for Tropical Versus Temperate Fishes and Urchins[END_REF][START_REF] Vergés | Long-term empirical evidence of ocean warming leading to tropicalization of fish communities, increased herbivory, and loss of kelp[END_REF]. In contrast, in temperate areas, herbivory by benthic invertebrates, especially sea urchins and gastropods seems to be predominant [START_REF] Hagen | Destructive grazing of kelp beds by sea urchins in Vestfjorden, northern Norway[END_REF][START_REF] Harrold | Food Availability, Sea Urchin Grazing, and Kelp Forest Community Structure[END_REF][START_REF] Norderhaug | Sea urchin grazing and kelp re-vegetation in the NE Atlantic[END_REF][START_REF] Filbee-Dexter | Sea urchin barrens as alternative stable states of collapsed kelp ecosystems[END_REF]. However, while there is a considerable amount of information showing an inverse correlation between the abundance of macroalgae and sea urchin densities [START_REF] Giakoumi | Relationships between fish, sea urchins and macroalgae: The structure of shallow rocky sublittoral communities in the Cyclades, Eastern Mediterranean[END_REF][START_REF] Sala | The Structure of Mediterranean Rocky Reef Ecosystems across Environmental and Human Gradients, and Conservation Implications[END_REF], the relationship between herbivorous fish and other invertebrates (decapods, gastropods, amphipods, isopods) and the conservational status of Cystoseira s.l. forests has not been assessed. Recently, the proliferation of herbivorous fish such as Sarpa salpa, and also omnivorous species such as Diplodus vulgaris, Coris julis or Thalassoma pavo, in the Mediterranean Sea has been related to the formation of shrubs and turfs, where forest-forming species are substituted by erect and/or turf-forming macroalgae [START_REF] Vergés | Role of fish herbivory in structuring the vertical distribution of canopy algae Cystoseira spp. in the Mediterranean Sea[END_REF][START_REF] Gianni | Threats to large brown algal forests in temperate seas: the overlooked role of native herbivorous fish[END_REF][START_REF] Papadakis | The neglected role of omnivore fish in the overgrazing of Mediterranean rocky reefs[END_REF]. These post regime shift scenarios (i.e. shrubs, turfs or barren grounds), either created by the proliferation of sea urchins, herbivorous fish or other herbivores, are considered stable states because of the difficulty to reverse the ecosystem to its original state of healthy marine forests, mainly due to the demographic characteristics of the forest-forming macroalgae (i.e. long lived and short dispersal; [START_REF] Chapman | Functional ecology of fucoid algae: twenty-three years of progress[END_REF][START_REF] Capdevila | Effective dispersal and density-dependence in mesophotic macroalgal forests: Insights from the Mediterranean species Cystoseira zosteroides[END_REF][START_REF] Riquet | Highly restricted dispersal in habitat-forming seaweed may impede natural recovery of disturbed populations[END_REF]. The effects of gastropods, decapods, amphipods and isopods have also been investigated on forest-forming species, revealing a variety of behaviours of the herbivores towards the macroalgae such as physical damage or clipping, facilitation effects by removing sediments or herbivory [START_REF] Pavia | Inducible chemical resistance to herbivory in the brown seaweed Ascophyllum nodosum[END_REF][START_REF] Perkol-Finkel | Conservation challenges in urban seascapes: promoting the growth of threatened species on coastal infrastructures[END_REF][START_REF] Gutow | Ocean acidification affects growth but not nutritional quality of the seaweed Fucus vesiculosus (Phaeophyceae, Fucales)[END_REF][START_REF] Suzuki | Effects of herbivorous gastropod grazing on the sedimentation and succession of subtidal macroalgal assemblages[END_REF].

However, the herbivory pressure is more studied on adults of Cystoseira s.l. spp. [START_REF] Vergés | Role of fish herbivory in structuring the vertical distribution of canopy algae Cystoseira spp. in the Mediterranean Sea[END_REF](Vergés et al., , 2014b;;[START_REF] Gianni | Threats to large brown algal forests in temperate seas: the overlooked role of native herbivorous fish[END_REF][START_REF] Papadakis | The neglected role of omnivore fish in the overgrazing of Mediterranean rocky reefs[END_REF] than on recruits. Since some studies found that a variety of invertebrates could graze on early-life stages of forest-forming macroalgae [START_REF] Lubchenco | Littornia and Fucus: Effects of Herbivores, Substratum Heterogeneity, and Plant Escapes During Succession[END_REF][START_REF] Korpinen | Grazing and nutrients reduce recruitment success of Fucus vesiculosus L. (Fucales: Phaeophyceae)[END_REF][START_REF] Alestra | Effects of opportunistic algae on the early life history of a habitat-forming fucoid: influence of temperature, nutrient enrichment and grazing pressure[END_REF][START_REF] Suzuki | Effects of herbivorous gastropod grazing on the sedimentation and succession of subtidal macroalgal assemblages[END_REF][START_REF] Barrientos | Feeding preferences of range-shifting and native herbivorous fishes in temperate ecosystems[END_REF][START_REF] Savonitto | Addressing reproductive stochasticity and grazing impacts in the restoration of a canopyforming brown alga by implementing mitigation solutions[END_REF] we wanted to study the potential herbivory effect of species usually present within shallow Cystoseira s.l. forests on recruits of these species.

In the Mediterranean Sea, the grazing on Cystoseira s.l. spp. has been principally studied on adults and for sea urchins and Sarpa salpa [START_REF] Vergés | Role of fish herbivory in structuring the vertical distribution of canopy algae Cystoseira spp. in the Mediterranean Sea[END_REF][START_REF] Agnetta | Role of two co-occurring Mediterranean sea urchins in the formation of barren from Cystoseira canopy[END_REF][START_REF] Gianni | Threats to large brown algal forests in temperate seas: the overlooked role of native herbivorous fish[END_REF][START_REF] Medrano | From marine deserts to algal beds: Treptacantha elegans revegetation to reverse stable degraded ecosystems inside and outside a No-Take marine reserve[END_REF][START_REF] Miller | Sea Urchin Removal as a Tool for Macroalgal Restoration: A Review on Removing "the Spiny Enemies[END_REF]. But as herbivory is believed to contribute to the high mortality observed during early-life stages [START_REF] Chapman | Functional ecology of fucoid algae: twenty-three years of progress[END_REF], in the second chapter we performed a series of experiments in the field and in the laboratory to better understand the herbivory pressure on Cystoseira s.l. recruits and for the first time, we selected small size non-strict herbivorous species (Clibanarius erythropus, Cerithium vulgatum, Idotea balthica and Gammarus sp.). The experiment in the field allowed us the elucidate the herbivory pressure in locations with different characteristics (e.g. presence or past presence of a Cystoseira s.l. forest). We observed a high mortality rate of the early-life stages, possibly linked to density-dependant processes, but also a significant effect of the herbivory protection. However, the effects of herbivory on recruits in the field should be reviewed and monitored during larger periods of time, as Cystoseira s.l. spp. are long-lived species and macroalgal populations that need decades to become functional and self-sustaining.

The experiments in the laboratory reported in the second chapter of this thesis also demonstrate that a variety of other non-strict herbivorous species usually found in shallow macroalgal communities (Clibanarius erythropus, Cerithium vulgatum and Idotea balthica) can graze on recruits of C. compressa, potentially contributing to the mortality of recruits observed in the field. These experiments represent a first approach to evaluate the role of potential herbivores and to elucidate which species must be considered to reduce or avoid grazing on recruits of Cystoseira s.l. spp. The results observed during our experiment are in agreement with the results performed with other species of fucoids and other macrophytes, reporting the important role Clibanarius erythropus, Cerithium vulgatum and Idotea balthica exert in structuring macroalgal assemblages [START_REF] Jonne | Seasonal Changes in situ Grazing of the Mesoherbivores Idotea baltica and Gammarus oceanicus on the Brown Algae Fucus vesiculosus and Pylaiella littoralis in the Central Gulf of Finland, Baltic Sea[END_REF][START_REF] Vergés | Experimental evidence of chemical deterrence against multiple herbivores in the seagrass Posidonia oceanica[END_REF][START_REF] Gunnarsson | The brown alga Fucus radicans suffers heavy grazing by the isopod Idotea baltica[END_REF][START_REF] Suzuki | Effects of herbivorous gastropod grazing on the sedimentation and succession of subtidal macroalgal assemblages[END_REF]. Our results showed that fish, sea urchins, decapods, gastropods and isopods can exert a top-down control potentially limiting the recruitment of shallow Cystoseira s.l. forests by consuming early-life stages of these species.

Contrarly to the effects of OW and OA on marine forests, the effects of the proliferation of herbivores are usually local and, therefore, could be addressed to prevent the loss of key species with effective management of affected adult or young restored populations [START_REF] Gianni | Reduction of herbivorous fish pressure can facilitate focal algal species forestation on artificial structures[END_REF][START_REF] Gianni | Optimizing canopy-forming algae conservation and restoration with a new herbivorous fish deterrent device[END_REF][START_REF] Suzuki | Effects of herbivorous gastropod grazing on the sedimentation and succession of subtidal macroalgal assemblages[END_REF][START_REF] Tamburello | Can we preserve and restore overlooked macroalgal forests?[END_REF]. To achieve this, recruits and mature adults of threatened populations should be protected against herbivory by using deterrent devices to ensure the continuity of the population [START_REF] Gianni | Optimizing canopy-forming algae conservation and restoration with a new herbivorous fish deterrent device[END_REF][START_REF] Savonitto | Addressing reproductive stochasticity and grazing impacts in the restoration of a canopyforming brown alga by implementing mitigation solutions[END_REF]. In the case of smaller invertebrates the herbivory deterrens may not be sufficient and when considering restoration actions the use of more expensive ex-situ cultivation techniques could be evaluated, as larger recruits could be less susceptible to the herbivory effects of smaller invertebrates [START_REF] Cebrian | A Roadmap for the Restoration of Mediterranean Macroalgal Forests[END_REF]. Despite that, these measures will not address the underlying cause of the proliferation of herbivores and are unlikely to provide a long-term solution to conserve and restore forests and ecosystem function on its own [START_REF] Miller | Sea Urchin Removal as a Tool for Macroalgal Restoration: A Review on Removing "the Spiny Enemies[END_REF]. Due to the growing number of restoration projects the topic of herbivory is more up to date than ever for being considered as one of the main causes for unsuccessful restoration actions of marine forests [START_REF] Gianni | Conservation and restoration of marine forests in the Mediterranean Sea and the potential role of Marine Protected Areas[END_REF][START_REF] Tamburello | Enhanced nutrient loading and herbivory do not depress the resilience of subtidal canopy forests in Mediterranean oligotrophic waters[END_REF][START_REF] Savonitto | Addressing reproductive stochasticity and grazing impacts in the restoration of a canopyforming brown alga by implementing mitigation solutions[END_REF]. The regulation of populations of herbivores could also be considered as tested by [START_REF] Medrano | From marine deserts to algal beds: Treptacantha elegans revegetation to reverse stable degraded ecosystems inside and outside a No-Take marine reserve[END_REF][START_REF] Miller | Sea Urchin Removal as a Tool for Macroalgal Restoration: A Review on Removing "the Spiny Enemies[END_REF]. Therefore, when considering or performing restoration actions it is critical to include herbivory management actions to limit its effects on the survival of recruits and adults transplanted [START_REF] Cebrian | A Roadmap for the Restoration of Mediterranean Macroalgal Forests[END_REF]. As an example, the proliferation of sea urchins has been largely related with the creation of barren grounds or marine deserts [START_REF] Giakoumi | Relationships between fish, sea urchins and macroalgae: The structure of shallow rocky sublittoral communities in the Cyclades, Eastern Mediterranean[END_REF][START_REF] Sala | The Structure of Mediterranean Rocky Reef Ecosystems across Environmental and Human Gradients, and Conservation Implications[END_REF][START_REF] Azzarello | Linking habitat characteristics to abundance patterns of canopy-forming macroalgae and sea urchins in southwest Australia[END_REF][START_REF] Agnetta | Role of two co-occurring Mediterranean sea urchins in the formation of barren from Cystoseira canopy[END_REF] and their removal is considered a tool for restoring macroalgal forests [START_REF] Miller | Sea Urchin Removal as a Tool for Macroalgal Restoration: A Review on Removing "the Spiny Enemies[END_REF]. Assessing the herbivory pressure in marine forests is essential for their conservation and restoration and for the conception of efficient herbivory deterrents. Further studies could be: (i) better identifying species grazing on Cystoseira s.l. populations in function of the life stage, (ii) performing feeding preference experiments to elucidate the real impact that potentiall herbivores have in locations where marine forest populations are present, and (iii) to elucidate if the feeding preferences and behaviours of these species can change under global change [START_REF] Barrientos | Feeding preferences of range-shifting and native herbivorous fishes in temperate ecosystems[END_REF].

In particular, climate change can affect herbivores either by altering their densities, changing their feeding behaviour, their food preferences or their consumption rates [START_REF] Gutow | Ocean acidification affects growth but not nutritional quality of the seaweed Fucus vesiculosus (Phaeophyceae, Fucales)[END_REF][START_REF] Barrientos | Feeding preferences of range-shifting and native herbivorous fishes in temperate ecosystems[END_REF][START_REF] Mitterwallner | Exposure to moderately elevated temperatures changes food preferences in the tropical marine herbivore Haliotis squamata[END_REF]. On one hand, sea urchins have been reported to decrease in abundance near natural CO2 seeps, possibly decreasing its herbivory rate and being less competitive under OA [START_REF] Porzio | Effects of ocean acidification on macroalgal communities[END_REF][START_REF] Asnaghi | Cascading Effects of Ocean Acidification in a Rocky Subtidal Community[END_REF][START_REF] Asnaghi | Decreased pH impairs sea urchin resistance to predatory fish: A combined laboratory-field study to understand the fate of top-down processes in future oceans[END_REF][START_REF] Baggini | Seasonality Affects Macroalgal Community Response to Increases in pCO2[END_REF]. On the other hand, the proliferation of non-indigenous warm water herbivorous fish, such as Siganus spp., is an indirect effect of OW on macroalgal communities, potentially leading to higher herbivory pressure and regime shifts (Vergés et al., 2014a(Vergés et al., , 2014b;;Bennett et al., 2015b;[START_REF] Barrientos | Feeding preferences of range-shifting and native herbivorous fishes in temperate ecosystems[END_REF]. It is suggested that range-shifting herbivorous fish can also affect recruits by consuming microscopic stages present in the turf matrix (Bennett et al., 2015b;[START_REF] Barrientos | Feeding preferences of range-shifting and native herbivorous fishes in temperate ecosystems[END_REF]. In addition, metabolic rates of herbivores could increase due to OW and OA increasing also their consumption rates [START_REF] Rich | Physiological and biochemical responses of a coralline alga and a sea urchin to climate change: Implications for herbivory[END_REF]. For example, according to [START_REF] Barrientos | Feeding preferences of range-shifting and native herbivorous fishes in temperate ecosystems[END_REF] the feeding rates on early-life stages of Eklonia radiata were higher under most tropicalized scenarios. Also, the carbon content of macroalgae could increase under OA altering, in some cases, the consumption rates by herbivores [START_REF] Gutow | Ocean acidification affects growth but not nutritional quality of the seaweed Fucus vesiculosus (Phaeophyceae, Fucales)[END_REF][START_REF] Leung | Boosted nutritional quality of food by CO2 enrichment fails to offset energy demand of herbivores under ocean warming, causing energy depletion and mortality[END_REF].

MARINE FORESTS LOSS AND FACILITATION OF BLOOMS OF TOXIC BENTHIC DINOFLAGELLATES

The degradation and losses that marine forests are experiencing and will experience under global change, will have profound consequences for the ecosystems [START_REF] Álvarez-Losada | A regime shift in intertidal assemblages triggered by loss of algal canopies: A multidecadal survey[END_REF][START_REF] Smale | Climate-driven substitution of foundation species causes breakdown of a facilitation cascade with potential implications for higher trophic levels[END_REF]. We have already discussed some causes responsible for marine forests loss: the abiotic interactive effect of OW and OA and the biotic effects of species facilitation and herbivory on recruits of C. compressa. In the framework of interspecific interactions, in the two last chapters of this thesis, we investigated a possible effect of marine forests loss, in particular the blooms of Ostreopsis spp. The results from the third chapter of this thesis, in agreement with the results of the latest bibliographic reviews on the ecology of toxic benthic dinoflagellates [START_REF] Berdalet | Key Questions and Recent Research Advances on Harmful Algal Blooms in Benthic Systems[END_REF][START_REF] Tester | Climate change and harmful benthic microalgae[END_REF], clearly demonstrate the current proliferation of blooms of Ostreopsis spp. in temperate zones, including the Mediterranean Sea. The observed increase in harmful algal blooms in the last decades relates to more effective detection through improved observation and monitoring capacities [START_REF] Hallegraeff | Global harmful algal bloom status reporting[END_REF], but also to the increasing anthropogenic impacts and global climate change effects [START_REF] Berdalet | Harmful Algal Blooms in Benthic Systems Recent Progress and Future Research[END_REF][START_REF] Tester | Climate change and harmful benthic microalgae[END_REF].

It is expected that the ongoing OW can facilitate the expansion and proliferation of blooms of the genus Ostreopsis, as their seasonal variation is often linked to temperature fluctuations, especially in the warm season [START_REF] Ingarao | Harmful Algae along Abruzzo coast from 2007 to 2010: correlations with environmental factors and new reports[END_REF]Accoroni and Totti, 2016). The effects of OA on Ostreopsis spp. still require further investigations, but Di Cioccio et al. ( 2014) found bloom abundances of Ostreopsis cf. ovata in CO2 seeps in Ischia Island (Mediterranean Sea) where pH values are similar to those predicted for the end of the century. Furthermore, an indirect effect of OW and OA on the proliferation of blooms in tropical areas could be the substitution of corals for macroalgae, that are a preferred substrate for the development of Ostreopsis spp. [START_REF] Berdalet | Harmful Algal Blooms in Benthic Systems Recent Progress and Future Research[END_REF]. Our results, predicting larger abundances of Ostreopsis spp. on shrub and turf-forming macroalgal communities; also foresee climate change to indirectly promote the development of blooms of Ostreopsis spp. through regime shifts, either from corals to macroalgae [START_REF] Jouffray | Identifying multiple coral reef regimes and their drivers across the Hawaiian archipelago[END_REF][START_REF] Rains | Gambierdiscus species exhibit different epiphytic behaviors toward a variety of macroalgal hosts[END_REF][START_REF] Rains | Gambierdiscus species exhibit different epiphytic behaviors toward a variety of macroalgal hosts[END_REF][START_REF] Johns | Macroalgal feedbacks and substrate properties maintain a coral reef regime shift[END_REF] or from complex macroalgal communities to less complex ones [START_REF] Bulleri | Facilitation and the niche: implications for coexistence, range shifts and ecosystem functioning[END_REF][START_REF] Filbee-Dexter | The present is the key to the past: linking regime shifts in kelp beds to the distribution of deep-living sea urchins[END_REF][START_REF] Pessarrodona | Homogenization and miniaturization of habitat structure in temperate marine forests[END_REF], that can trigger this species development [START_REF] Fraga | Review of the main ecological features affecting benthic dinoflagellate blooms[END_REF]. Despite that, our bibliographic review makes patent that the ecological aspects involved in the proliferation of blooms of Ostreopsis spp. are still largely understudied and that most of the studies do not focus on the ecology of these species [START_REF] Pavaux | Chemical Ecology of the Benthic Dinoflagellate Genus Ostreopsis : Review of Progress and Future Directions[END_REF] and in particular on the description of the meso-and macrohabitat of the sampled locations.

Even if the growth, abundance, and distribution of benthic dinoflagellates are controlled by chemical (nutrients and salinity) and physical (temperature, irradiation and hydrodynamics) factors, biological (competition, predation, facilitation) factors are likely to influence Ostreopsis spp. [START_REF] Pavaux | Chemical Ecology of the Benthic Dinoflagellate Genus Ostreopsis : Review of Progress and Future Directions[END_REF][START_REF] Tester | Climate change and harmful benthic microalgae[END_REF]. It has been recently suggested that the characteristics of the macroalgal substrate, such as the thallus architecture and the production of allelopathic compounds, could facilitate or regulate the development of epibenthic dinoflagellates (Totti et al., 2010;Accoroni et al., 2015;[START_REF] Ternon | Allelopathic interactions between the benthic toxic dinoflagellate Ostreopsis cf. ovata and a cooccurring diatom[END_REF][START_REF] Pavaux | Chemical Ecology of the Benthic Dinoflagellate Genus Ostreopsis : Review of Progress and Future Directions[END_REF][START_REF] Tester | Using artificial substrates to quantify Gambierdiscus and other toxic benthic dinoflagellates for monitoring purposes[END_REF].

Allelochemicals produced by many macrophytes can inhibit or stimulate epibionts (Accoroni et al., 2015;[START_REF] Ternon | Allelopathic interactions between the benthic toxic dinoflagellate Ostreopsis cf. ovata and a cooccurring diatom[END_REF]Ternon et al., , 2020) ) and consequently affect the epibenthic community. Following the trend highlighted in the third chapter, where certain macroalgal groups are more commonly sampled in Ostreopsis spp. studies, we performed a preference experiment in the field. In the last chapter, we tested the effects of the micro-and mesohabitat on the facilitation of Ostreopsis spp. proliferation in the field, during an annual bloom. The effects at large scale were, in some cases, more important than the effects at small scale for the proliferation of Ostreopsis spp. The high variability on cell abundances and the contradictory results among macroalgal species and habitats in function of the sampling and methodology used for the quantification of the cells, has already been reported in other studies [START_REF] Berdalet | Harmful Algal Blooms in Benthic Systems Recent Progress and Future Research[END_REF]Jauzein et al., 2018;[START_REF] Tester | Using artificial substrates to quantify Gambierdiscus and other toxic benthic dinoflagellates for monitoring purposes[END_REF]. At the small scale, the variability in the abundance of cells can be high, and linked to local habitat characteristics such as hydrodynamics, light exposure, predation, nutrients, available surface, etc. [START_REF] Berdalet | Harmful Algal Blooms in Benthic Systems Recent Progress and Future Research[END_REF][START_REF] Tester | Climate change and harmful benthic microalgae[END_REF]. However, our results indicate that the physical structure of macroalgae alone does not explain the preferences of Ostreopsis spp. Overall, according to our results, Dictyota spp. seems to be a good substrate for the development of Ostreopsis spp. (this thesis Chapter 4; Park et al., 2020;[START_REF] Ibghi | Occurrence of three dominant epibenthic dinoflagellates (Ostreopsis spp., Coolia monotis and Prorocentrum lima) in relation to biotic substrates and environmental factors in a highly dynamic ecosystem, the Strait of Gibraltar (Southwestern Mediterranean)[END_REF], contrarly to the results from studies on allelopathic interactions (Accoroni et al., 2015;Ternon et al., 2020) that suggest a strong negative effect of some Dictyotales on Ostreopsis cf. ovata. The preferences could be due to physical characteristics of the macroalgae, to their allelochemical proprieties, to the presence of other organisms from the epyphitic community or to environmental factors acting at larger scale [START_REF] Ternon | Allelopathic interactions between the benthic toxic dinoflagellate Ostreopsis cf. ovata and a cooccurring diatom[END_REF][START_REF] Pavaux | Chemical Ecology of the Benthic Dinoflagellate Genus Ostreopsis : Review of Progress and Future Directions[END_REF].

As reported, in the last two chapters, abundances of Ostreopsis spp. can change in function of the methodology used in the sampling and quantification of the cells (Mangialajo et al., 2017;Jauzein et al., 2018;Vassalli et al., 2018). Therefore, the highly used quantification of cells of Ostreopsis spp. per gram FW of macroalgae may not be representative of what is really happening in the macroalgal community, as it is dependent on the specific weight of the macroalgae. The methodologies and quantifications that are best representing the reality of the ecosystem should be further investigated and better implemented in future studies. Finding the ideal sampling procedure for Ostreopsis spp. will not be an easy task, and should reflect the real risk of bloom for the sampled location. Also, the method should consider the structural complexity and characteristics of the benthic habitats sampled, the diversity of available substrates and the spatial and temporal scales of variability [START_REF] Berdalet | Harmful Algal Blooms in Benthic Systems Recent Progress and Future Research[END_REF]. The high variability on preferences of Ostreopsis spp. at the small scale (micro-and mesohabitat) could be influenced by conditions acting at the large scale, at the ecosystem level (abundance of patches of favourable mesohabitat, hydrodynamics, water quality temperature…), and for this reason so far has not been possible to reach clear conclusions (Accoroni and Totti, 2016;Fricke et al., 2018).

Macroalgal species can affect benthic dinoflagellates abundances, but epiphytic dinoflagellates in turn can also affect the macroalgal substrates [START_REF] Iveša | Increased ammonium levels occurring during benthic algal blooms are potentially toxic to fucalean algae[END_REF]. For example, O. cf. ovata cells show a high affinity for ammonia [START_REF] Berdalet | Harmful Algal Blooms in Benthic Systems Recent Progress and Future Research[END_REF][START_REF] Jauzein | Uptake of dissolved inorganic and organic nitrogen by the benthic toxic dinoflagellate Ostreopsis cf. ovata[END_REF] which can lead to thallus necrosis in Cystoseira s.l. populations due to the ammonium concentrations present on the mucilaginous aggregates of the microalgae during blooms [START_REF] Iveša | Increased ammonium levels occurring during benthic algal blooms are potentially toxic to fucalean algae[END_REF]. Further studies are needed to investigate the effects of Ostreopsis spp. biofilm on macroalgae, therefore we cannot exclude that recurrent blooms of benthic dinoflagellates could constitute another factor favouring the local loss of fucoids [START_REF] Iveša | Increased ammonium levels occurring during benthic algal blooms are potentially toxic to fucalean algae[END_REF]. To better understand the link between the disappearance of marine forests and the proliferation of blooms of Ostreopsis spp., we should collect information about the deterioration of the environmental parameters in locations with annual bloom of Ostreopsis spp., and historical and present data about marine forest presence.

Managing harmful algal blooms is extremely important, and with the increasing number of coastal urban areas, mitigating the impacts of blooms of Ostreopsis spp. on human health is becoming of major interest [START_REF] Berdalet | Marine harmful algal blooms, human health and wellbeing: challenges and opportunities in the 21st century[END_REF][START_REF] Berdalet | Environmental, human health and socioeconomic impacts of Ostreopsis spp. Blooms in the NW Mediterranean[END_REF]. The contact with contaminated seawater or the inhalation of aerosols can result in human intoxications and with ecological (mass mortalities of invertebrates) and socio-economic consequences in coastal touristic locations, such as the Mediterranean coast [START_REF] Berdalet | Marine harmful algal blooms, human health and wellbeing: challenges and opportunities in the 21st century[END_REF][START_REF] Berdalet | Harmful Algal Blooms in Benthic Systems Recent Progress and Future Research[END_REF][START_REF] Berdalet | Environmental, human health and socioeconomic impacts of Ostreopsis spp. Blooms in the NW Mediterranean[END_REF]. It is estimated that in the Provence-Alpes-Côte-d'Azur region, N-W Mediterranean Sea, an increase in Ostreopsis spp. outbreaks could have an economic cost from several hundred thousand to several million euros from loss tourism [START_REF] Lemee | Interactions between scientists, managers and policy makers in the framework of the French MediOs project on Ostreopsis (2008-2010)[END_REF]. The results obtained in this thesis highlight the high variability of Ostreopsis spp. abundances on macroalgal species and communities. However, the comparisons of cell abundances of Ostreopsis spp. on macroalgal species is controversial and makes it difficult to find clear patterns for the development of blooms of Ostreopsis spp. Next steps to better elucidate the relationship between Ostreopsis spp. blooms and macroalgal communities should involve: (i) setting similar studies at large scale, avoiding transplantation of missing macroalgal communities [START_REF] Bravo | Life cycle stages of the benthic palytoxin-producing dinoflagellate Ostreopsis cf. ovata (Dinophyceae)[END_REF](Bravo et al., , 2020;;[START_REF] Accoroni | New insights on the life cycle stages of the toxic benthic dinoflagellate Ostreopsis cf. ovata[END_REF]Yong et al., 2018;Lee et al., 2020), (ii) performing allelopathic experiments on different species of Ostreopsis and macroalgae (Accoroni et al., 2015;Ternon et al., 2020), and (iii) extending these experiments to other species of toxic benthic dinoflagellates [START_REF] Mustapa | Growth and epiphytic behavior of three Gambierdiscus species (Dinophyceae) associated with various macroalgal substrates[END_REF]Bravo et al., 2020).

GENERAL CONCLUSIONS

Globally, the results of this thesis allowed a step forward in the understanding of the causes and effects of Cystoseira s.l. forests loss in the Mediterranean Sea in the framework of BHABs proliferation. The findings provide important insights into the causes and effects of Cystoseira s.l.

spp. loss and are of major interest in the conservation of Mediterranean marine forests, contributing to the development of effective management measures. The early-life stages of Cystoseira compressa are negatively affected by ocean warming and acidification, and by the presence of Neogoniolithon brassica-florida. Recruits of C. compressa can also be affected by the herbivory pressure of several non-strict herbivorous species. Blooms of Ostreopsis spp. seem to prefer shrub and turf-forming macroalgae, such as Dictyota spp. and Halopteris scoparia, even if, in some locations, the high variability of cells of Ostreopsis spp. among substrats and communities could hide other potential patterns of Ostreopsis spp. preferences. Larger scale studies would be needed to confirm these results.

The restoration of ecosystems is essential in slowing biodiversity and ecosystem services loss.

Marine forests, due to their ecosystem services, offering nursery, habitat, protection and food for many invertebrates and fishes, have to be considered when restoring coastal locations (Wernberg and Filbee-Dexter, 2019). It is important to know the causes driving marine forest loss and prevent the effects of their decline, such as BHABs. It is not always possible to remove environmental causes of marine forests loss (e.g. herbivory), or even virtually impossible (e.g. OW and OA).

Future restoration actions could limit the effects of marine forests loss, especially through joint efforts, selecting the most adequate techniques prioritizing resistant species and/or populations and the less impacted areas for restoration. Such restoration actions will not only increase the productivity and biodiversity of coastal ecosystems but could potentially mitigate the public health, forests, (ii) in the conception of future restoration actions and guidelines in a changing sea, and (iii) in better assessing and eventually regulating Ostreopsis spp. blooms. -There is a need to collect more information at the mesohabitat scale to better understand the preferences of Ostreopsis spp. Chapter 4

Key highlights by chapter

-Ostreopsis spp. seem to prefer shrub and turf-forming macroalgae even if, in Rochambeau, the high variability of cells among substrats and communities could hide other potential patterns of Ostreopsis spp. preferences. Larger scale studies would be needed to confirm these results. -Dictyota spp. are potentially a good substrate for Ostreopsis spp. proliferation in Vernazzola.

-The morphology of the macroalgae alone, excluding allelopathic interactions, is likely not enough to explain Ostreopsis spp. preferences. -The macroalgal preferences of Ostreopsis spp. can vary in function of the counting methodology and the type of quantification used for the sampling. Di Turi, L., Lo Caputo, S., Marzano, M. C., Pastorelli, A. M., Pompei, M., Rositani, L., et al. (2003).

Ostreopsidiaceae (Dinophyceae) presence along the coastal area of Bari. Biol. Mar. Mediterr. 10,[675][676][677][678].

  the financial support of the 3-year scholarship Emplois Jeunes Doctorants from the Région Sud Provence-Alpes-Côte d'Azur. The research conducted in this thesis has been supported by the European Union project AFRIMED, funded from the Executive Agency for Small and Medium Enterprise (EASME) and European Maritime and Fisheries fund (EMFF) under grant agreement n° 789059. VI RÉSUMÉ ……………….…...…….

1 Figure 1 :Figure 2 :

 112 Figure 1: Different types of marine macroalgal habitats in function of the structural characteristics of the dominant species: marine forest, shrub and turf. Modified from Thiriet (2014). .................................................................................................................................. 4 Figure 2: Distribution of marine forests of large brown macroalgae for different regions.

FigureCHAPTER 1 :Figure 1 :Figure 2 :

 112 Figure from Wernberg and Filbee-Dexter (2019). ............................................................... 5

Figure 3 :Figure 4 :Figure 5 :Figure 6 :Figure 1 :Figure 2 :Figure 3 :Figure 4 :Figure 5 :Figure 6 :Figure 7 :Figure 8 :Figure 9 :Figure 1 :Figure 2 :Figure 3 :Figure 4 :Figure 5 :Figure 6 :Figure 7 :Figure 8 :Figure 1 :Figure 2 :Figure 3 :Figure 4 :Figure 6 : 1 :Table 1 :Table 1 :Table 1 :Table 2 :

 3456123456789123456781234611112 Figure 3: Density (a) and size (b) of recruits of C. compressa as a function of temperature during the first experiment (96 days). The errors bars show the confidence intervals. ...... 30 Figure 4: Densities of recruits on living and dead Neogoniolithon brassica-florida and on artificial substrates, as a function of temperature and pH under the different treatments. The errors bars show the confidence intervals. .......................................................................... 31 Figure 5: Size of recruits on living and dead Neogoniolithon brassica-florida and on artificial substrates as a function of temperature and pH under the different treatments. The errors bars show the confidence intervals. .......................................................................... 32 Figure 6: Calcification rate of Neogoniolithon brassica-florida after 64 days under different treatments. Error bars show the confidence intervals. ........................................................ 33

  . To assess the ongoing impacts of climate change, the Intergovernmental Panel on Climate Change (IPCC) prepares Assessment Reports about the state of scientific, technical and socio-economic knowledge on climate change using the Shared Socioeconomic Pathways (SSPs). The SSPs are scenarios of projected socioeconomic global changes up to 2100 used to derive greenhouse gas emissions scenarios from different climate policies (IPCC; 2022). The last IPPC report is the Sixth Assessment Report (AR6 Climate Change 2022: Mitigation of Climate Change -IPCC). GENERAL INTRODUCTION AND OBJECTIVES …………………………………………..………………………………

Figure 1 :

 1 Figure 1: Different types of marine macroalgal habitats in function of the structural characteristics of the dominant species: marine forest, shrub and turf. Modified from Thiriet (2014).

Figure 2 :

 2 Figure 2: Distribution of marine forests of large brown macroalgae for different regions. Figure from Wernberg and Filbee-Dexter (2019).

  assigned three different genera to the clades based on their morphological characteristics: Cystoseira C. Agardh, 1820; Treptacanta Kützing, 1843; and Carpodesmia Greville, 1830. More recently, Molinar-Novoa and Guiry (2020) revised this proposition concluding that Gongolaria Boehmer, 1760 and Ericaria Stackhouse, 1890 have priority over the ones proposed by Orellana et al. (2019), and now Cystoseira, Ericaria and Gongolaria are the accepted names for the three clades (Molinari -Novoa and Guiry, 2020). Shallow Cystoseira s.l. forest in Saint Honorat Island (Lérins Islands, Cannes, France) composed by Ericaria amentacea, Ericaria brachycarpa and Cystoseira compressa. Cystoseira s.l. spp. form canopies up to 1 m height in exposed or sheltered locations from the upper littoral to the circalittoral zone, down to 50 m depth, and represent the highest level of Mediterranean macroalgae complexity

Chapter 1 .

 1 Climate change and species facilitation affect the recruitment of macroalgal marine forests. In this chapter we assess the effects of ocean warming, acidification and species facilitation on the recruitment of C. compressa in controlled experiments in the laboratory. We determine how future climate change scenarios could affect recruitment and growth of C. compressa and how the presence of crustose coralline algae can facilitate the recruitment under the different treatments of temperature and pH.

Figure 1 :

 1 Figure 1: Experimental set-up used to test the effects of temperature (28 and 32°C), pH (ambient 8.07, and low 7.8) and species facilitation (artificial, coralline and dead coralline substrates) on the recruits of C. compressa. The experimental set-up was repeated 4 times, resulting in 8 header tanks, and 48 experimental tanks in which the different types of substrates were randomly assigned.

(

  64 days). Surfaces of coralline substrates were determined on photographs using the software ImageJ (ImageJ, NIH US Department of Health and Human Services). All experiments were carried out according to relevant regulations and guidelines concerning Cystoseira compressa and Neogoniolithon brassica-florida sampling. The latter were collected under prefectoral order No. 277, delivered by the Interregional Directorate of the Mediterranean Sea, Regulatory/Control Service, authorizing the ECOSEAS Laboratory to sample fauna and flora for scientific purposes only.

Figure 2 :

 2 Figure 2: In-situ mean seawater temperature at the donor population site.

Figure 3 :

 3 Figure 3: Density (a) and size (b) of recruits of C. compressa as a function of temperature during the first experiment (96 days). The errors bars show the confidence intervals.

Figure 4 :

 4 Figure 4: Densities of recruits on living and dead Neogoniolithon brassica-florida and on artificial substrates, as a function of temperature and pH under the different treatments. The errors bars show the confidence intervals.

Figure 5 :

 5 Figure 5: Size of recruits on living and dead Neogoniolithon brassica-florida and on artificial substrates as a function of temperature and pH under the different treatments. The errors bars show the confidence intervals.

Figure 6 :

 6 Figure 6: Calcification rate of Neogoniolithon brassica-florida after 64 days under different treatments. Error bars show the confidence intervals.

  CHAPTER 2 …….….……………..…..… 43

  2 and 1.0 m depth) is composed mainly of C. compressa, Gongolaria barbata, Ericaria crinita and Ericaria brachycarpa. The two sites deprived of Cystoseira s.l. forests are located on the continental open coast in (i) Fourmis' Bay in Beaulieu-sur-Mer (from now Beaulieu), a semi-exposed site between 0.5 and 1 m depth, and (ii) Passable in Saint -Jean -Cap -Ferrat (from now Passable), a sheltered site around 0.5 m depth (Figure 1).

Figure 1 :

 1 Figure 1: Location of the three experimental sites: the rockpool system in Sainte Marguerite Island in Lérins Islands (43°30'57.6''N, 7°3'14.4''E), and the two open coast locations Beulieu (43°42'3.6''N, 7°19'48''E) and Passable (43°41'42''N, 7°16'33.6''E).

  Before the start of the experiments, the presence and density of known and potential herbivorous species, including sea urchins, decapods, gastropods and Sarpa salpa, were characterized at the three locations. The density of decapods and gastropod was determined using 20*20 cm quadrates in the rockpool system in Sainte Marguerite (n = 40) and in the open coast in Beaulieu (n = 20) and CHAPTER 2 …….….……………..…..… Passable (n = 20); while the density of sea urchins was assessed using 50*50 cm quadrates (n = 20 per site). The surveys were performed from summer to autumn 2020 and 2021 on five different days in Sainte Marguerite and in 2021 on three different days in Beaulieu and Passable. Herbivorous fish, mainly Sarpa salpa larger or equal to 5 cm length, were visually quantified in Sainte Marguerite, while in Beaulieu and Passable, herbivorous fish densities were quantified by recording stationary videos at random points and counting number of S. salpa individuals in a radius of 5 m during 5 min (n = 10; Sala and Ballesteros, 1997).

Figure 2 :

 2 Figure 2: Representation of the different experiments performed. The two herbivory experiments in the field, to asses the herbivory pressure on recruits of C. compressa; and the mesocosm experiment, to assess the potential herbivory on recruits of C. compressa of the decapod Clibanarius erythropus, the gastropod Cerithium vulgatum, the amphipod Gammarus sp. and the isopod Idotea balthica.

Field experiment 2 :

 2 Ex-situ recruitment on open coast in deforested sites Beaulieu and Passable Field experiment 1: In-situ recruitment in a forested rockpool Sainte

C

  . erythropus and C. vulgatum individuals were obtained in the Sainte Marguerite, within the marine forest, while I. balthica and Gammarus sp. were obtained from Anse des Fossés (Saint-Jean-Cap-Ferrat, France) within leaves of Posidonia oceanica detritus. Different numbers of individuals of each species were selected and placed in separated close system experimental tanks(2.5 L). In each experimental tank, there was one substrate with similar biomass of recruits of Cystoseira compressa (between 2-3 months old). Recruits were obtained in the laboratory following the same ex-situ cultivation technique as in section 2.4 and kept in 30 L tanks until the beginning of the mesocosm experiment. The substrates with C. compressa were cleaned from epiphytes and/or growing biofilm with tweezers and a brush before the start of the experiment. The selected densities of individuals for C. erythropus were 1, 2 and 3 individuals per tank (n = 5); for C. vulgatum were 1, 3 and 5 individuals per tank (n = 5); and for Gammarus sp. and I. balthica 1 individual per tank (n = 7). Controls with 0 herbivorous were included (n = 5). As the experiments with C. erythropus and C. vulgatum were performed simultaneously, the same controls (n = 5) were used for both species. The experiments with Gammarus sp. and I. balthica were performed later and, as were again performed simultaneously, both species share the control experimental tanks with 0 individuals (n = 5). This experiment consisted in a total of 50 experimental tanks. All the species were kept without food for 48h before the start of the experiment. Every two days the temperature was measured and the filtered seawater (200 µm mesh) of the tanks was changed, faeces were cleaned every day. C. erythropus and C. vulgatum were maintained in the experimental tanks 23 days and Gammarus sp. and I. balthica 4 days, until they consumed all or almost all of the available biomass of C. compressa. C. erythropus and C. vulgatum were only weighted once at the end of the experiment (mg FW) after extracting them from their shells. Gammarus sp. and I. balthica individuals where weighted (mg FW) every 24h. The mean biomass of Gammarus sp. individuals was 0.08 ± 0.02 g and the mean biomas of I. balthica individuals was 0.07 ± 0.01 g (mean ± SD) at the beginning of the experiment. The consumption of the different species was assessed as the consumed biomass (mg FW) of recruits and calculated from the number of C. compressa recruits.At the start of the experiment the mean biomass of C. compressa on each tank was 5.02 ± 0.53 mg FW for C. erythropus and C. vulgatum and 6.05 ± 0.91 mg FW (mean ± SD) for Gammarus sp. and I. balthica (Appendix B: Chapter 2 Supplementary material). The biomass of recruits was evaluated every 24h the first week and then twice a week.The biomass (mg FW) of the C. compressa recruits was extrapolated from a linear regression [Volume (mm 3 ) ~ biomass (mg FW)]. To do so, the volume of each recruit was calculated approximating its shape to a cone and semi sphere (Appendix B: Chapter 2 Supplementary material). The maximum length and maximum width of each recruit was measured under a stereomicroscope. Then this volume was transformed to g FW, using the previously adjusted volume (mm 3 ) ~ biomass (mg FW) linear regression using other recruits (R 2 = 0.994). To obtain the linear regression, the volume of 38 recruits was calculated by water displacement (mL). The volume of each recruit was measured six times and the mean of the measurements was used. Then each recruit was dried with absorbent paper before being weighed three times in mg (0.001), for obtaining the FW. Correlating the volume and the biomass of each recruit we adjusted the linear regression (Appendix B: Chapter 2 Supplementary material).A Linear Mixed-Effects Model (LMER) was used to test the cumulative biomass consumption of recruits of C. compressa by C. erythropus, C. vulgatum and I. balthica, with the density of herbivorous (four levels for C. erythropus, C. vulgatum and two levels for I. balthica) as a fixed factor, and time and the aquarium as a random factor as the LMER models can cope with repeated measures over time. Finally, a Linear Model (LM) was used to test the consumption rate (mg C. compressa mg herbivor -1 day -1 ) of the different species, with the species (three levels) as fixed factor. Gammarus sp. was exclude as no consumption was detected. CHAPTER 2 …….….……………..…..… 49

Figure 3 :

 3 Figure 3: Density of herbivores in the three studied locations. The errors bars show the standard error.

Figure 4 :

 4 Figure 4: Boxplot of the density of recruits of C. compressa per 0.04 m 2 of natural substrates in the rockpool in Sainte Marguerite 2 months after the starting of the experiment. The bold horizontal lines indicate the median value (Q2), the box marks the interquartile distances (Q1 and Q3), the whiskers mark the values that are less than Q3 + 1.5 * IQR but greater than Q1 -1.5 * IQR.

Figure 5 :

 5 Figure 5: Density of recruits of C. compressa per 0.04 m 2 of substrates in the rockpool in Sainte Marguerite. The natural population densities of C. compressa in Sainte Marguerite are represented in red considering the mean and the standard error (3.05 ± 0.14 individuals 0.04 m -2 , mean ± SD). The errors bars show the standard error.

Figure 6 :

 6 Figure 6: Size class distribution of the C. compressa recruits on Sainte Marguerite's substrates over time for each treatment. The X-axis represent the size classes in 1 cm intervals and the Y-axis is the proportion of each size class per treatment.

Figure 7 :

 7 Figure 7: Average evolution of the density of recruits of C. compressa (individuals per 0.04 m 2 )on the two open coast sites and in the laboratory controls. The density on the laboratory control substrates was lower as the most recruited substrates were selected to be transplanted to the field. The errors bars show the standard error.

Figure 8 :

 8 Figure 8: Cumulative consumption of C. compressa biomass (mg FW) by Clibanarius erythropus, Cerithium vulgatum, Gammarus sp. and I. balthica, and control substrates. Not that the biomass loss in the controls (0 individuals) it is not due to herbivory. The errors bars show the standard error.Regarding the results of the LM used to test the consumption rate of C. compressa by C. erythropus, C. vulgatum and I. balthica, the last one was the herbivore consuming more C. compressa, with no difference between the consumption rate of C. erythropus and C. vulgatum (Figure9and Table1).

Figure 9 :

 9 Figure9: Consumption rate (mg C. compressa mg herbivore -1 day -1 ) for the three herbivores for the whole duration of each experiment. Gammarus sp. is not represented as we could not observe consumption of C. compressa by this species. The error bars show the standard error.

  observed how individuals from the same species (fishes and decapods) exerted a stronger herbivory pressure on artificial habitats than on natural ones, and the same could have happened during our experiments in Beaulieu and Passable that are located in urbanized areas. The recruits obtained in the rockpool within a Cystoseira s.l. forest (Sainte Marguerite) could have been less affected by herbivores because of the lager abundance and variety of other more palatable species of macroalgae present in the site that were not present in the open coast location (Beaulieu and Passable), possibly explaining the rapid decrease in recruits observed in the urban open coast locations.

  from 1973 to 2021. Only 249 studies, published from 1981 to 2021, were based on field sampling of Ostreopsis spp. cells and were therefore considered as relevant for the review (Appendix C: Chapter 3 Supplementary material). The 249 considered studies comprise 957 sampling locations, involving a total of 1644 different substrates sampled between 1972 and 2019. 90% of the papers from this review (224) focused on Ostreopsis cf. ovata. CHAPTER 3 ………………….…………. The number of studies per year reveals a positive trend of ecological studies involving sample collection in the field (Figure 1) in both temperate and tropical areas. The first studies involving field sampling were performed in the tropics, but, since year 2000, studies performed in temperate areas have exceeded in number the tropical ones. Since 2018 tropical studies are again more numerous than in temperate areas, but this trend has to be verified in the next years. Nearly half of the 957 different locations investigated between 1972 and 2019 are in temperate areas (471), the other half (486) in tropical ones. The exact geographical coordinates were available for 595 (62.2%) sampling locations and it was possible to position 132 (13.8%) additional ones on the basis of the description of the location in the text. A distribution map of the 727 locations, out of the 957 sampled in the 249 relevant studies, is reported in Figure 2.

Figure 1 :

 1 Figure 1: Trend of the number of peer-reviewed studies involving field sampling of Ostreopsis spp. from 1981 to 2021.

Figure 2 :

 2 Figure 2: Map of the sampling locations. The colour corresponds to the year it was first sampled. The two red lines (35°N and 35°S) separate the tropical/subtropical from the temperate areas; (A) global distribution; (B) detailed map for the Mediterranean Sea.

Figure 3 :

 3 Figure 3: Macrohabitats where Ostreopsis spp. have been sampled in temperate and tropical areas.

  respectively). Seagrass meadows are also regularly sampled, especially in tropical areas (5.2%).

Figure 4 :

 4 Figure 4: Mesohabitats where Ostreopsis spp. have been sampled in tropical and temperate areas.

Figure 5 :

 5 Figure 5: Microhabitats (substrates) where Ostreopsis spp. have been sampled in temperate and tropical areas.

Figure 6 :

 6 Figure 6: Macroalgae sampled in function of the phylum.

Figure 7 :

 7 Figure 7: Macroalgae sampled in function of the type of community they can form.

Figure 8 :

 8 Figure 8: Maximum cell densities of Ostreopsis spp. (cells g -1 FW macroalgae) for each study grouped according to the morphological structure of the macroalgae.

Figure 1 :

 1 Figure 1: The two sampling locations: Rochambeau in the Bay of Villefranche-sur-Mer (France) and Vernazzola in Genova (Italy).

Figure 2 :

 2 Figure 2: Abundances of Ostreopsis spp. in seawater (1000 Cells Ostreopsis spp. / L) for Rochambeau (n = 4) and Vernazzola (n = 2) for the duration of the experiment. The confidence interval represents the standard error.

Figure 3 :

 3 Figure 3: Abundances of Ostreopsis spp. on natural macroalgae (cells Ostreopsis spp. / g FW macroalgae) in Rochambeau (A) and Vernazzola (B). The error bars show the standard error.

Figure 4 :

 4 Figure 4: Abundances of Ostreopsis spp. on artificial macroalgae (cells Ostreopsis spp. / cm 2 artificial macroalgae) in Rochambeau (A) and Vernazzola (B). The error bars show the standard error.

Figure 5 :

 5 Figure 5: Abundances of Ostreopsis spp. on natural macroalgal communities (cells Ostreopsis spp. / 0.04 m 2 macroalgal communities) sampled in Rochambeau with the adapted BEDI device. The error bars show the standard error.

Figure 6 :

 6 Figure 6: Concentration of Ostreopsis spp. on natural macroalgal communities (cells Ostreopsis spp. / 0.04 m 2 ) in Rochambeau (A) and Vernazzola (B). The error bars show the standard error.

  ecological and economic consequences of Ostreopsis spp. blooms. The results presented in this thesis follow the European Union Biodiversity Strategy for 2030, United Nations Decade on Ecosystem Restoration and the 2030 Agenda for Sustainable Development objectives, widening the view of the effects of global change (i.e. OW, OA and herbivores proliferation) on the disappearance of marine forests, potentially triggering the development of Ostreopsis spp. blooms in the Mediterranean Sea. Overall, the insights reported in this thesis can be useful across species and could help us (i) in the conservation of Cystoseira s.l.

Chapter 1 - 2 - 3 -

 123 High temperatures (32°C) and low pH(7.8) negatively affect the density of recruits of Cystoseira compressa and the calcification rate of its associate understory crustose coralline algae Neogoniolithon brassica-florida.-C. compressa recruit size is larger under lower temperatures (24°C and 28°C) and low pH(7.8).-The presence of N. brassica-florida negatively affect the recruitment of C. compressa, independently of the temperature and pH level. -OW and OA are expected to have a negative impact on C. compressa populations.-Clay substrates are a good support for the recruitment of C. compressa and recommended for restoration actions. Chapter Herbivory control has to be considered for the protection of Cystoseira s.l. forests and the success of restoration actions. -Herbivory pressure on recruits of C. compressa can differ among locations.-Different herbivorous and omnivorous species can graze on recruits of Cystoseira compressa compromising the population recruitment and future restoration actions. Chapter Studies involving field sampling of Ostreopsis spp. increased in the last decade in both temperate and tropical areas. -In both temperate and tropical areas, erect and turf-forming macroalgae are the most sampled substrate for Ostreopsis spp. sampling, independently of the mesohabitat or macrohabitat dominant in the sampling location. -Ostreopsis spp. are often sampled on macroalgal communities characteristic of post-regime shift scenarios. -The description of the mesohabitat where Ostreopsis spp. are sampled is rarely reported.

Figure 2 :

 2 Figure 2: Real size artificial macroalgae unit Dictyota type. The measures are in mm.

Figure 3 :

 3 Figure 3: Real size artificial macroalgae unit Halopteris type. The measures are in mm.

Figure 4 :

 4 Figure 4: Real size artificial macroalgae unit turf type. The measures are in mm.

BOX 3: Cystoseira compressa (Esper) Gerloff & Nizamuddin 1975

	GENERAL INTRODUCTION AND OBJECTIVES
	…………………………………………..………………………………
	Caespitose macroalgae, light brown colour, very variable in size, from several centimetres to 80 cm
	in maximum height according to the degree of wave action. Some specimens are perennial but often
	present an annual development. Cylindrical or flattened axes of several centimetres high (2-3 cm),
	almost inexistent in juveniles (A). Apices of the axes are smooth (B). Tophules are absent. Primary
	branches totally flattened, at least at their base. No spiny appendages. Aerocysts usually associated
	with the receptacles. Receptacles in the apices of the terminal branches, simple or more frequently
	branched, sometimes located on an aerocyst. Male and female gametangia in conceptacles grouped
	in receptacles (C and D). They present a monophasic diploid life cycle
	This thesis focuses on Cystoseira s.l. forests from shallow coastal areas, mainly inhabiting
	rockpools and shallow sheltered locations, usually very impacted by human activities (e.g.
	urbanisation, recreational activities, climate change) which could be magnified due to the low
	turnover rate conditions of these locations. In view of the new restoration techniques developed for
	Mediterranean Cystoseira s.l. forests (see Cebrian et al., 2021 for a review), we consider necessary
	to further study the causes that could negatively impact Cystoseira s.l. spp. in the future (e.g. climate
	change and species interactions) and compromise the success of such restoration actions. In
	addition, despite the efforts done in understanding the mechanisms compromising Cystoseira s.l.
	forests in the Mediterranean still little is known on how the interactive effects of climate change
	(i.e. ocean warming and acidification) affect this species, especially its early-life stages. We have
	selected Cystoseira compressa (Box 3) as a target species for several reasons. First, this macroalgal
	species is considered the most resistant Cystoseira s.l. spp. and, still at the present time, can be
	found in both cosmopolitan and pristine locations in many coastal areas (including the French
	Riviera; Thibaut et al., 2015; Verlaque et al., 2019). Second, there are currently no specific studies
	on the effect of climate change on adults and recruits of C. compressa, nor results of restoration
	attempts for this species. Thus, the first objective is to study the causes of loss, assessing how
	climate change and species interactions affect the recruits of C. compressa.

Table 1 :

 1 Measured (regular characters) and expected (bold characters) seawater physico-chemical parameters (temperature in °C, pHT in total scale, calculated pCO2 in µatm, and total alkalinity in µmol Kg -1 with mean ± SD) according to different treatments.

	Treatment	Temperature (°C)	pHT n = 15	pCO2 (µatm) n = 15	Total alkalinity (µmol kg -1 ) n = 8
	24°C	24.15 ± 0.43	-	-	-
		n = 23
	28°C	28.3±0.41	-	-	-
		n = 23
	32°C	31.47 ± 0.60	-	-	-
		n = 23
	28°C pH 8.07	28.22 ± 0.71 n = 15	8.04 ± 0.04	434.23 ± 42.63	2564.32 ± 5.80
	28°C pH 7.8	28.37 ± 0.74 n= 15	7.83 ± 0.12	795.89 ± 170.97	2568.73 ± 22.51
	32°C pH 8.07	31.56 ± 0.47 n = 15	8.01 ± 0.03	468.05 ± 45.53	2564.96 ± 6.23
	32°C pH 7.8	31.61 ± 0.70 n = 15	7.83 ± 0.063	788.17 ± 135.69	2563.26 ± 5.50

Table 1 :

 1 Results from the statistical analyses.

	2 )
	compressa (n°ind. / 0.04m	200 400 600 800
	Density C.	0	Protected Artefact control	Open	Protected Artefact control	Open
			Recruitment enhancement	No recruitment enhancement

Experiment I. balthica lmer (Biomass consumption (mg FW) ~ n° herbivores + (1 | Time) + (1 | Tank))

			N° herbivores (0 and 1)	1	45.562	< 0.001 ***
	All herbivores	lm (Consumption Rate mg FW C. ~ Species of herbivores) compressa + mg FW herbivores -1 * Day -1 )	Species of herbivores (4 levels)	2	27.666	< 0.001 ***

Table 1 :

 1 Maximum cell densities of Ostreopsis spp. (cells g -1 FW macrophyte) for each study on different microhabitats.

						Temperate
						Tropical
	Log maximum density	cells Ostreopsis spp. g -1 FW)
		(log
		Forest Shrub	Turf	Forest Shrub	Turf	Forest Shrub	Turf
		Major bloom		Bloom		No bloom

  that small filamentous macroalgae developing on dead or damaged coral surfaces could foster Ostreopsis spp. development, showing the role of macroalgae as a favourable substrate.

	CHAPTER 3
	………………….………….
	2018) suggested
	Benthic dinoflagellate development is affected by interacting biotic and abiotic factors linked to the
	habitat and acting at different scales (i.e. substrate, benthic community and seascape). Interestingly
	the present study highlighted that the sampled substrate (herein the microhabitat, e.g. coral
	fragment, macroalgal species) and the ecosystem (herein the macrohabitat, e.g. coral reefs,
	macroalgal-dominated rocky shores, sandy or muddy bottoms) where Ostreopsis spp. are sampled
	are generally reported and described. In 98% and 79% of the studies the sampled substrate and the
	seascape are respectively described or, alternatively, the text allows undoubtful extrapolation of
	this information. Concerning the microhabitat, and in particular organic substrates, most studies
	76

. The search performed in this study did not take into account non-indexed papers or grey literature likely providing a low estimation of old papers, in particular in developing countries. Since the 90s Ostreopsis spp. are reported in temperate areas; a rapid increase in publications in temperate areas after 2006 is likely due to the first recognised mass intoxication in Genoa (Italy) in 2005

[START_REF] Brescianini | Ostreopsis ovata algal blooms affecting human health in Genova, Italy, 2005 and 2006[END_REF]

) that encouraged several European research groups to focus on this emergent phenomenon. An increase in publications in tropical areas is registered after 2010, often in synergy with research performed in temperate areas. It is worth noting that the peak of Ostreopsis spp. publications in temperate areas registered in 2012 is due to the publication of 18 articles in a special issue (Cryptogamie Algologie) resulting from the proceedings of the International Congress on Ostreopsis Development (ICOD conference, 2011;

[START_REF] Lemee | Interactions between scientists, managers and policy makers in the framework of the French MediOs project on Ostreopsis (2008-2010)[END_REF]

. detail the species (56.8%), or at least the genus (20.1%) of the organisms sampled. Yet, even if the coral fragment or shell is identified at the species level, it is not clearly reported if 1) it is a living organism and 2) it is the direct Ostreopsis spp. host. Some studies

(Widiarti, 2008; Yong et al.,

Table 1 :

 1 Macroalgae species (microhabitat) and communities (mesohabitat scale) sampled in Rochambeau and Vernazzola.

	MACROALGAL SPECIES AND COMMUNITIES	SAMPLING SITE	BIOMASS (g mean ± SD) FW / 0.04 m 2 ,	COMPOSITION
	TURF -DICTYOTA
	SPP.	Rochambeau		Jania rubens (80%), Dictyota spp. (20%)
	TURF -FORMING
	TURF TURF -FORMING	Rochambeau	55.34 ± 16.45	Small Padina pavonica and Halopteris scoparia (80%), J. rubens (10%), crustose coralline (10%), Acetabularia acetabulum and Cladophora sp. (1%)
				Polysiphonia sp, Chondracantus acicularis and
	TURF TURF -FORMING	Vernazzola	133.50 ± 26.16	other filamentous macroalgae (50%), Dictyota spp. (20%), Ellisolandia elongata (20%), Caulerpa cylindracea (5%), Hypnea sp. (5%) and Ulva sp.
				(2%)
	DICTYOTA SPP. SHRUB -FORMING	Rochambeau	75.65 ± 22.30	Dictyota spp. (80%), J. rubens (10%) and crustose coralline (10%)
	DICTYOTA SPP. SHRUB -FORMING	Vernazzola	87.00 ± 25.45	Dictyota spp. (80%) and crustose coralline (20%)
				H. scoparia (90%), P. pavonica (10%), A.
	HALOPTERIS			acetabulum and Haliptilon virgatum (2%),
	SCOPARIA	Rochambeau	135.98 ± 34 .36	Caulerpa cylindracea, Peyssonnelia sp.,
	SHRUB -FORMING			Sphaerococcus coronopifolius and E. elongata
				(1%)
	HALOPTERIS SCOPARIA SHRUB -FORMING	Vernazzola	120.00 ± 45.25	H. scoparia (70%), Dictyota spp. (20%), Caulerpa cylindracea (5%), E. elongata (5%)
	CYSTOSEIRA COMPRESSA FOREST-FORMING	Rochambeau	150.75 ± 59.44	Cystoseira compressa (90%), crustose coralline (10%) *
	CYSTOSEIRA COMPRESSA FOREST-FORMING	Vernazzola	241.50 ± 34.65	C. compressa (85%), Sargassum vulgare (10%), crustose coralline (5%)
	GONGOLARIA BARBATA FOREST-FORMING	Rochambeau	53.79 ± 1.77	Gongolaria barbata (90%), crustose coralline (10%) *
	SARGASSUM VULGARE FOREST-FORMING	Vernazzola	96.50 ± 21.92	S. vulgare (90%), C. compressa (5%), crustose coralline (5%)

  . In Genova-Quarto on July 2017Meroni et al. (2018b) reported abundances reaching 2.89 × 10 6 cells / g FW on Halopteris scoparia.

	CHAPTER 4
	………………….………….
	0,04 m 2 -Scrapings Cells Ostreopsis spp./	4e+08 6e+08 0e+00 2e+08 0e+00 2e+08 4e+08 6e+08	A 07/07/2021 D1 23/06/2021 B	11/07/2021 D2 12/07/2021	15/07/2021 D3 21/07/2021 30/07/2021	C. compressa Dictyota spp. Turf H. scoparia G. barbata C. compressa Dictyota spp. Turf H. scoparia S. vulgare 05/08/2021 D5 22/08/2022 21/07/2021 D4 01/09/2021
			D1	D2	D3	D4	D5	D6

28,25 0,79 8,05 0,05 429,42 57,51 2562,06 5,79 Coralline 28,19 0,66 8,04 0,04 439,46 50,41 2563,77 3,60 Dead Coralline 28,21 0,69 8,04 0,04 435,52 50,13 2566,66 7,20 28°C pH 7.8 Artificial 28,29 0,75 7,83 0,14 807,15 192,03 2564,20 3,52 Coralline 28,38 0,73 7,84 0,07 784,99 154,70 2564,79 15,41 Dead Coralline 28,43 0,73 7,84 0,13 793,41 166,83 2561,56 3,59 32°C pH 8.07 Artificial 31,57 0,44 8,01 0,04 468,46 47,74 2566,97 5,36 Coralline 31,58 0,48 8,00 0,03 477,15 39,59 2565,73 6,77 Dead Coralline 31,56 0,48 8,02 0,04 459,05 47,13 2562,03 6,51 32°C pH 7.8 Artificial 31,61 0,76 7,82 0,07 805,88 155,87 2561,92 4,89 Coralline 31,54 0,70 7,83 0,06 783,92 135,27 2562,58 6,06 Dead Coralline 31,67 0,64 7,84 0,05 774,72 112,88 2565,43 5,90

Photo 1: Artificial macroalgae Cystoseira type. Photo 2: Artificial macroalgae Dictyota type.

ACKNOWLEDGEMENTS

This work is supported by a PhD grant funded by the Région Provence-Alpes-Côte d'Azur (contract Emplois Jeunes Doctorants 2019-2022) and the AFRIMED project funded by the Executive Agency for Small and Medium Enterprise (EASME) and European Maritime and Fisheries Fund (EMFF) under grant agreement N° 789059. We thank C. Carbonne for the fruitful exchanges and support and to all the internship students that helped during the project: M. Ortolani, B. Brunet, M. Gillio, K. Vigreux and A. Muret. We are grateful to Sandra from Atelier Terracotta (Nice, France) who helped us with the clay substrates. Thanks to M. Drake for the English proofreading of the article AUTHOR'S CONTRIBUTION ACKNOWLEDGEMENTS This work is supported by a PhD grant funded by the Région Provence-Alpes-Côte d'Azur (contract Emplois Jeunes Doctorants 2019-2022), CoReFOs and the project CONVOST (UCA JEDI Investments ANR-15-IDEX-01). We thank G. Romero, F. Priouzeau and G. Spennato for their help constructing and installing the cages in the field and for helping in the Volume ~ Biomass regression. ACKNOWLEDGEMENTS This work is supported by a PhD grant funded by the Région Provence-Alpes-Côte d'Azur (contract Emplois Jeunes Doctorants 2019-2022), CoReFOs and the project CONVOST (UCA JEDI Investments ANR-15-IDEX-01). ACKNOWLEDGEMENTS This work is supported by a PhD grant funded by the Région Provence-Alpes-Côte d'Azur (contract Emplois Jeunes Doctorants 2019-2022) and the AFRIMED project funded by the Executive Agency for Small and Medium Enterprise (EASME) and European Maritime and Fisheries Fund (EMFF) under grant agreement N° 789059. We thank F. Priouzeau and G. Romero for their help in the confection of the artificial macroalgae and in the setting up of the experiment. We also thank all the internship students that helped during the project: B. Brunet, M. Gillio, K. Vigreux and A. Muret. CHAPTER 4 ………………….………….

my sincere thanks to the organizations that have made it possible to finance this research. This thesis has been realised thanks to the doctoral grand Emplois Jeunes Doctorants 2019 -2022 from the Région Provence-Alpes-Côte d'Azur and in the framework of the European project AFRIMED on the restoration of the degraded Mediterranean macroalgal forests, funded from the Executive Agency for Small and Medium Enterprise (EASME) and European Maritime and Fisheries fund (EMFF) under grant agreement n° 789059. I would also like to express my thanks to the members of my thesis committee, Pr.

DATA AVAILABILITY

Data Availability Statement: Data Availability Statement: The datasets generated and analysed during the current study are available in the Knowledge Network for Biocomplexity repository, https://knb.ecoinformatics.org/view/urn:uuid:daa9cc97-47eb-48a3-addc-ed0047f0f3c4

ABSTRACT

Grazing is one of the most important biological factors controlling the mortality of early-life stages of fucoids and one of the major problems when restoring marine forests. Benthic macroinvertebrates (e.g. sea urchins) and fish shape and regulate benthic macroalgal communities from temperate to tropical regions, being responsible for the formation of turfs or even barren grounds.

However, other smaller invertebrates associated with marine forests could significantly participate in the grazing, especially on early-life stages. Knowing which invertebrates could participate in the grazing of recruits of forest-forming macroalgae could be crucial for their conservation. We performed several experiments in the field and in mesocosms in order to investigate the herbivory pressure and the effects of different grazers on recruits of Cystoseira compressa. The results highlight that non-strict herbivorous species, such as Clibanarius erythropus, Cerithium vulgatum and Idotea balthica, can potentially graze on recruits of Cystoseira s.l. spp. probably affecting the success of marine restoration actions. From the different invertebrates studied in the mesocosms, I. balthica had the higher consumption rates of recruits of C. compressa. We conclude that biotic factors such as herbivory need to be well understood when planning restoration actions and that anti-herbivory devices adapted to the local diversity of potential herbivores should be included when restoring marine forests.

CHAPTER 4: Ostreopsis spp. BLOOMS IN RELATION TO MACROALGAL COMMUNITIES

ABSTRACT

There has been an increasing occurrence of benthic harmful algal blooms (BHAB) in temperate locations during the last decades. Moreover, the expansion and growth rates of benthic harmful dinoflagellates of the genus Ostreopsis could be favoured under future ocean warming scenarios.

Ostreopsis spp. are known to develop on the surface of macroalgae in both temperate and tropical areas. There is evidence that the morphology of the macrophytes could play a role in the abundance of dinoflagellates, being the filamentous morphotypes suggested to host higher abundances of the genus Ostreopsis. The fact that Ostreopsis spp. could be associated with structurally less complex macroalgal communities is of special interest, due to the generalized phenomenon of habitat miniaturization as the result of regime shifts in temperate ecosystems. In the present study samples for Ostreopsis spp. were taken at different spatial scales and on different macroalgal species and communities, using different sampling techniques and methodologies to quantify abundances. The objective was to understand the role that different macroalgal species (microhabitat) and communities (mesohabitat) can play in controlling the distribution and abundance of Ostreopsis spp. In general, the results obtained highlight the high variability and wide distribution of Ostreopsis spp. among macroalgal species and communities during a bloom, and the suitability of less complex macroalgal communities in hosting Ostreopsis spp. The high variability of Ostreopsis spp. abundances on bloom locations could be due to the stronger effects at the large scale, that in turn, could influence the dominance of specific macroalgal communities in the location. Larger scale studies would be needed to confirm these results.

Abundances of Ostreopsis spp. on artificial macroalgae (microhabitat scale)

The ANOVA from the GLMM model did not find differences in abundances of Ostreopsis spp.

among the different types of artificial macroalgae, neither with the control in Rochambeau (Figure 5A and Table 2). In Rochambeau, abundances of Ostreopsis spp. on artificial macroalgae remained high until day 3 (15/07) when they started decreasing (Figure 4A). The highest abundances on artificial macroalgae reached 1.38 × 10 5 cells / cm 2 on turf the day 3 (15/07), mean abundances on this sampling day were also the highest (mean abundance of 5.57 × 10 4 cells / cm 2 ).

APPENDIX A: CHAPTER 1 SUPPLEMENTARY MATERIAL