# Furthering knowledge on seaweed growth and development to facilitate sustainable aquaculture

Bénédicte Charrier, Maria Helena Abreu, Rita Araujo, Annette Bruhn, Juliet C. Coates, Olivier de Clerck, Christos Katsaros, Rafael Romero Robaina, Thomas Wichard

► To cite this version:

Bénédicte Charrier, Maria Helena Abreu, Rita Araujo, Annette Bruhn, Juliet C. Coates, et al. Furthering knowledge on seaweed growth and development to facilitate sustainable aquaculture. New Phytologist, 2017, 216 (4), pp.697-975. 10.1111/nph.14728. hal-02183661

HAL Id: hal-02183661

https://hal.science/hal-02183661

Submitted on 17 Jul 2019

HAL is a multi-disciplinary open access
archive for the deposit and dissemination of sci-
entific research documents, whether they are pub-
lished or not. The documents may come from
teaching and research institutions in France or
abroad, or from public or private research centers.

L'archive ouverte pluridisciplinaire HAL, est
destinée au dépôt et à la diffusion de documents
scientifiques de niveau recherche, publiés ou non,
émanant des établissements d'enseignement et de
recherche français ou étrangers, des laboratoires
publics ou privés.

<figure><img src="image_1.png" /></figure>

Distributed under a Creative Commons Attribution 4.0 International License
Furthering knowledge on seaweed growth and development
to facilitate sustainable aquaculture.

<table><tr><td>Journal:</td><td>New Phytologist</td></tr><tr><td>Manuscript ID</td><td>NPH-V-2017-23688.R1</td></tr><tr><td>Manuscript Type:</td><td>V - Viewpoint</td></tr><tr><td>Date Submitted by the Author:</td><td>n/a</td></tr><tr><td>Complete List of Authors:</td><td>Charrier, Benedicte; Centre National de la Recherche Scientifique, Station Biologique Abreu, Helena; ALGAPLUS Lda, SME Araújo, Rita; European Commission, Joint Research Centre, Directorate for Sustainable Resources, Water and Marine Resources Unit; Universidade do Porto Centro Interdisciplinar de Investigação Marinha e Ambiental, CIIMAR Bruhn, Annette; Aarhus Universitet, Bioscience Coates, Juliet; University of Birmingham, School of Biosciences; De Clerck, Olivier; Ghent University, Onderzoeksgroep Algologie Katsaros, Christos; National and Kapodistrian University of Athens, Department of Biology Robaina, Rafael; Universidad de Las Palmas de Gran Canaria, Ciencias del Mar Wichard, Thomas; Friedrich Schiller University of Jena, Institute for Inorganic and Analytical Chemistry</td></tr><tr><td>Key Words:</td><td>aquaculture, development, macroalga, marine, reproduction, seaweed, morphogenesis, biomass</td></tr></table>
1 **Viewpoint**

2 **Title :**

3 **Furthering knowledge on seaweed growth and development to facilitate sustainable aquaculture.**

4

5 **Authors:**

6 Bénédicte Charrier¹, Maria Helena Abreu², Rita Araujo³, Annette Bruhn⁴, Juliet C. Coates⁵, Olivier

7 De Clerck⁶, Christos Katsaros⁷, Rafael R. Robaina⁸, Thomas Wichard⁹

8 **Affiliations:**

9 ¹ Morphogenesis of Macroalgae, UMR8227, CNRS-UPMC, Station Biologique, 29680 Roscoff, France. ² ALGAplus Lda, Travessa Alexandre da Conceição 3830-196 Ilhavo, Portugal. ³ European Comission, Joint Research Centre - Directorate for Sustainable Resources, Water and Marine Resources Unit, Ispra, Italy. ⁴ Aarhus University, Department of BioScience, Vejlsøvej 25, 8600 Silkeborg, Denmark. ⁵ School of Biosciences, University of Birmingham, Birmingham B15 2TT, United-Kingdom. ⁶ Department of Biology, Ghent University, Krijgslaan 281 S8, 9000 Ghent, Belgium. ⁷ Morphogenesis of Macroalgae, National and Kapodistrian University of Athens, Department of Biology, Athens, 157 84, Greece. ⁸ Facultad de Ciencias del Mar. Universidad de Las Palmas de Gran Canaria. ⁹ Friedrich Schiller University Jena, Institute for Inorganic and Analytical Chemistry, Lessingstr. 8, 07743 Jena, Germany.

19 Author for correspondence:

20 *Bénédicte Charrier*
21 Tel: 33-2-98-29-56-53

22 email: benedicte.charrier@sb-roscoff.fr

23 **Keywords:** development, macroalgae, life cycle, morphogenesis, reproduction, seaweeds.

<table><tr><td>Total word count (excluding summary)</td><td>3080</td><td>No. of Figures</td><td>4 (all in colour)</td></tr><tr><td>Summary</td><td>167</td><td>No. of Table</td><td>0</td></tr><tr><td>Paragraph 1: Ecological and societal position of macroalgae</td><td>609</td><td>No. of Supporting Information Files</td><td>0</td></tr><tr><td>Paragraph 2: Mastering genetics through the control of the life cycle</td><td>798</td><td>Open Access</td><td>Yes</td></tr><tr><td>Paragraph 3: Early and microscopic stages of development</td><td>388</td><td></td><td></td></tr><tr><td>Paragraph 4: Controlled growth and organogenesis factors: towards biomass production monitoring</td><td>332</td><td></td><td></td></tr><tr><td>Paragraph 5: Current technological requirements</td><td>655</td><td></td><td></td></tr><tr><td>Paragraph 6: Conclusion</td><td>267</td><td></td><td></td></tr><tr><td>Acknowledgements</td><td>20</td><td></td><td></td></tr></table>
24 Abstract

Macroalgae (seaweeds) are the subject of increasing interest for their potential as a source of valuable, sustainable biomass in the food, feed, chemical and pharmaceutical industries. Compared to microalgae, the pace of knowledge acquisition in seaweeds is slower despite the availability of whole-genome sequences and model organisms for the major seaweed groups. This is partly due to specific hurdles related to the large size of these organisms and their slow growth. As a result, this basic scientific field is falling behind, despite the societal and economic importance of these organisms. Here, we argue that sustainable management of seaweed aquaculture requires fundamental understanding of the underlying biological mechanisms controlling macroalgal life cycles - from the production of germ cells to the growth and fertility of the adult organisms - using diverse approaches requiring a broad range of technological tools. This viewpoint highlights several examples of basic research on macroalgal developmental biology that could enable the step-changes which are required to adequately meet the demands of the aquaculture sector.

37 Ecological and societal position of macroalgae

Macroalgae are macroscopic aquatic organisms belonging to three distinct and distantly-related eukaryotic lineages (commonly named green, red, and brown algae). Their unicellular ancestors diverged more than 1.6 billion years ago (Parfrey et al., 2011) implying independent acquisitions of multicellularity, and leading to a bewildering diversity of life cycles, fertilization processes and morphogenetic strategies. At the ecological level, macroalgae fulfil important roles as key habitat-structuring agents and primary producers in coastal ecosystems. The goods and services seaweeds (marine macroalgae) support are varied (Figure 1), and include elevated secondary production,
45 nutrient cycling, energy capture and flow, and coastal defence (Steneck et al., 2002). They can also
46 significantly contribute to carbon sequestration at a level exceeding that of angiosperm marine
47 coastal vegetation (up to 1.5 times as much as seagrass meadows, salt marshes and mangroves and
48 up to 2% of the annual anthropogenic emission; Krause-Jensen & Duarte, 2016 and references
49 therein). In addition, macroalgae support complex food webs in coastal zones and provide habitats
50 and food for associated organisms, from apex predators to invertebrates (Reisewitz et al., 2006).

51 Macroalgal communities also enable transfer of biomass between ecosystems (Krumhansl &
52 Scheibling, 2012), removal of dissolved nutrients from coastal waters and coastal protection from
53 erosion (Arkema et al., 2013). De Groot et al. (2012) estimated the value of coastal ecosystem
54 services provided by macroalgae to be over 28,000 intL.$·ha⁻¹·year⁻¹.

55 Seaweeds are also an alternative/additional source of food, feed, fuel, biomolecules and livelihood
56 for humans. Over 80% of macroalgal production and harvesting is at present destined for human
57 consumption directly (Abreu et al., 2014) or as hydrocolloids (thickeners, gelling agents, etc)
58 (Rebours et al., 2014). Macroalgae are also used as fertilizers and animal feed (Makkar et al.,
59 2016). In addition, the industrial sector uses seaweed biomass for nutraceuticals, cosmetics,
60 biotechnological and pharmaceutical applications, thus propelling the growth of seaweed
61 biotechnology (Mazarrasa et al., 2013). Currently, ~28 million tonnes of seaweeds per year (wet
62 weight) are produced and, as a proxy for the growth of the biotechnology-market of seaweed-
63 derived products, seaweed-related patent applications increased at a rate of 11% per year since
64 1990 (Mazarrasa et al., 2014).

65 While in Asia 99% of seaweed production is sourced from cultivation (accounting for 93% of the
66 global production in 2013) (FAO, 2016), the dominant practice of non-Asian countries is still
67 harvesting natural stocks. However, the availability of wild stocks under the current scenario of
68 global change needs to be assessed, while management plans for seaweed exploitation must be
69 adapted to the natural population dynamics of commercially important species. Increasing demands
70 for high-quality seaweed biomass may therefore affect the long-term sustainability of seaweed
71 exploitation. Seaweed cultivation is the alternative to cope with industry's demand for biomass,
72 concomitantly protecting natural resources (Fig. 1). Unlike terrestrial crops, they do not compete
73 for arable land, fertilizer and freshwater resources. Furthermore, the development of Integrated
74 Multi-Trophic Aquaculture (IMTA: co-cultivation of seaweeds with fin/shell fishes) enables
75 recapture of excessive inorganic nutrients released in coastal areas by fish farms, thereby
76 improving their sustainability (Holdt & Edwards, 2014). Beyond aquaculture proper, seaweed
77 cultivation could also function as a general instrument for circular resource management (Seghetta
78 et al., 2016), treatment of waste-water produced by land-based farming and municipal treatment
79 plants (Neveux et al., 2016), heavy metal biosorption (He & Chen, 2014) and recolonisation of
80 artificial reefs (Fig. 1). As a response to this assessment, the European seaweed aquaculture sector
81 has progressively expanded, accounting for 12% of total European biomass production in 2013
82 (FAO, 2016). Further expansion calls for advances in seaweed production technology, which rely
83 on a better knowledge of both the environmental and the intrinsic factors controlling the
84 development of macroalgae.

85 How could developmental biology help solve bottlenecks in seaweed aquaculture?

86 *Mastering genetics through the control of the life cycle*

87 Most seaweeds have complex, biphasic life cycles, involving free-living haploid gametophyte and
diploid sporophyte generations (Coelho *et al.*, 2007) (Box 1). Either phase of the life cycle can be
88 exploited, depending on the seaweed species. The harvestable biomass of kelps consists of
89 sporophytes up to several meters long (45 m in *Macrocystis*), while in nori (*Pyropia and*
90 *Porphyra*), the life stage of interest is the haploid gametophyte. Other exploited seaweeds e.g.
91 *Gracilaria* and *Chondrus* (red algae) have isomorphic life-cycles, with both sporophyte and
92 gametophyte developing macroscopic exploitable thalli. Currently, clonal propagation (e.g. red alga
93
Kappaphycus) and recourse to a limited number of parent genotypes (kelp) account for the
production of most commonly cultivated seaweeds. The resulting impoverishment of genetic
diversity increases seaweed susceptibility to diseases and decreases their fitness within their
cultivation environment (Loureiro et al., 2015). For example, the continuous vegetative
propagation of the carrageenophyte Kappaphycus in intensively cultivated areas has increased its
vulnerability to diseases (e.g. bacterial mediated "ice-ice" disease), thereby dramatically impacting
the production in various countries (Largo et al., 1995). This problem requires counteraction by the
selection of new breeding strains, potentially through artificial hybrids (Gupta et al., 2015), but
more optimally through crossings, as somatic hybridisation usually results in severe and unstable
phenotypic alteration (Charrier et al., 2015). However, whilst in some seaweeds the promotion of
sexual reproduction still requires development (e.g. Gracilariopsis; Zhou et al., 2013), the loss of
the genetic patrimony resulting from cross-fertilisation might be detrimental to maintaining specific
and valuable genotypes resulting from decades of selection. Therefore, manipulating the different
steps of the seaweed life cycles would allow a balance between the maintenance of given
genotypes of interest and controlled breeding. Progress in basic research opens possible paths to
bypass steps of the life cycle, thereby allowing to reach this goal (Box 1).

Manipulating the sexual life cycle.

Most cultivated seaweeds reproduce sexually (kelps, red algae *Porphyra* ssp.), placing both time
and genetic constraints on seaweed farmers. Physiological studies have long been establishing
protocols for maintaining seaweeds in a vegetative stage or shifting them to the next phase using
specific temperature and light conditions, or even by tissue ablation. This allows year-round
production of juveniles and increases the cultivated net biomass (Pang & Lüning, 2004). Several
illustrations of these practices applied to exploited seaweeds are displayed in Box 1. Recent
fundamental studies propose potential alternatives. Treatments with algal phytohormones could be
used to control the vegetative-to-reproductive transition and speed up reproduction, as illustrated in
119 the red alga *Grateloupia imbricata* upon addition of methyl jasmonate (García-Jiménez *et al*.,
120 2016).

121 *Promoting parthenogenesis*.

122 Other seaweeds propagate vegetatively from a single life phase through parthenogenesis, mainly by
123 apogamy but also by apomeiosis. The flexibility is high and is a valuable feature for aquaculture, as
124 it allows the maintenance of a specific genotype in potentially morphologically different organisms
125 (Box 1, left side). Parthenogenesis can be induced by hybridisation (e.g. *Caloglossa*
126 tetrasporophytes; Kamiya & West, 2008) or through chemical treatments preventing gamete
127 motility (e.g. formaldehyde in brown algae Ectocarpales; Gwo & Chen, 1999). The lab-based
128 identification of endogenous factors controlling seaweed parthenogenesis might provide more
129 natural alternatives to regulate or manipulate parthenogenesis in aquaculture. Recently, Han *et al.*
130 (2014) identified three mitochondrial proteins involved in the control of parthenogenesis in
131 *Scytosiphon lomentaria* (brown alga Ectocarpales). In parallel, Arun *et al.* (2013) showed that algal
132 chemical factors (so far unidentified) secreted by the parthenosporophyte of *Ectocarpus siliculosus*
133 (brown alga Ectocarpales) control the fate of the released zoospores (Box 1). Coelho *et al.* (2011)
134 showed that the whole parthenosporophytic stage itself was controlled by a single genetic locus.
135 The characterisation of these factors could lead to the development of additional strategies to
136 control parthenogenesis.

137 Finally, Li *et al*., (2014) produced *Undaria pinnatifida* (brown alga) gametophytes that made only
138 male gametes from both oogonia and antheridia (Shan *et al*., 2015). These gametes are able to self-
139 cross and to produce homozygous male diploid sporophytes. This example illustrates that crosses
140 are controlled by the morphological identity of the reproductive organs rather than by their
141 genotypes, emphasizing the importance of a control over morphogenesis.

142 In parallel to these improvements for seaweeds cultivated off-shore (Fernand et al., 2017),
143 standardized protocols should also be developed specifically for not-yet cultivated, high-value
144 seaweeds amenable to on-shore cultivation. This includes seaweeds producing high-value
145 chemicals, or seaweeds in high demand on the food market, such as *Ulva*, *Palmaria*, *Porphyra*,
146 *Cystoseira*, *Himanthalia*, *Codium*, *Polysiphonia* and *Asparagopsis* (Abreu *et al.*, 2014), as well as
147 the red macroalgae *Ochtodes* and *Portieria* cultivated in photobioreactors (Rorrer & Cheney,
148 2004).

149 Altogether, basic research into the development and reproduction of macroalgae will likely provide
150 alternative means of manipulating seaweed reproduction, which will be very valuable for future
151 breeding programmes and aquaculture practices (Cottier-Cook *et al.*, 2016).

152 *Early and microscopic stages of development*

153 Seaweed growth starts with the formation and development of juveniles, which originate from the
154 release and germination of single cells (zygotes or spores). They subsequently attach to marine
155 substrata to initiate their sessile development (bloom-forming algae are usually free-living).
156 Deciphering the early and microscopic developmental stages of seaweeds is an important
157 requirement for future integrative management of their cultivation (Fig. 2). Exploitation of seaweed
158 biomass concentrates on the macroscopic life-cycle stage, which is the sporophyte in the most
159 predominantly exploited brown algae (*Ecklonia*, *Laminaria*, *Saccharina*, *Undaria*), together with
160 the gametophyte in red seaweeds (*Gracilaria*, *Kappaphycus*, *Euchema*) and in some isomorphic
161 green (*Ulva*) seaweeds. Optimizing fertilisation success could help control the rate of production of
162 seaweed embryos in hatcheries, which, when too high, impedes the quality of sporophyte juveniles
163 (Fig. 2 and 3). Environmental cues inducing fertility and spore/gamete release have been
164 determined for tens of seaweed species (photoperiod, irradiance, temperature and nutrient
165 concentration; previous section and Box 1). However, the paucity of molecular studies regarding
166 e.g. the periodicity of gamete release, attraction of gametes to opposite sex or mating type, and
167 cell-cell recognition (Fig. 3) stands in a stark contrast to the wealth of eco-physiological and
168 biochemical studies that predate the molecular era. As an illustration, in certain *Ulva* species,
gametogenesis and subsequent gamete release can be artificially induced by removal of sporulation and swarming inhibitors (Vesty et al., 2015 and references therein), but so far, neither these inhibitors nor the signalling pathways inducing gametogenesis have been characterised. Similar cases could be made for pheromone signalling in brown seaweeds (Boland, 1995) and glycoprotein recognition between opposite-sex gametes (Schmid et al., 1994).

Many macroalgal zygotes experience polarisation prior to the growth and development of the embryo (Fig. 3), similarly to land plants and metazoans. Whether polarisation is necessary for proper development, and the identity of polarisation cues and regulatory factors, are unknown for most macroalgae: only Fucales and Dictyotales (brown algae) zygotes have allowed the identification of detailed polarisation cues (light direction and location of sperm entry; Brownlee et al., 2001; Bogaert et al., 2017) and of specific cell cycle checkpoints (Bothwell et al., 2008). Bogaert et al. (2017) recently described in Dictyota a unique two-phase polarisation mechanism, thereby illustrating the importance of seaweeds to decipher fundamental developmental processes in the tree of life.

*Controlled growth and organogenesis factors: towards biomass production monitoring,*

Production of large seaweed biomass with specific features of industrial interest (polysaccharides, proteins and pigments) depends both on seaweed net growth and seaweed capacity to grow organs and tissues with specific structures and compositions. Indeed, the quantity and quality of key compounds vary within the algal body (beta-glucan in *Durvillaea*: Bobadilla et al., 2013; phytohormones in *Sargassum*: Li et al., 2016), and cells with thicker walls, storage organelles and vacuoles might be more resistant to dehydration, chemical exposure, eutrophication, and pathogen attacks, and hence be of high interest. Unfortunately, macroalgal cell fate specification is one of the least-understood areas of macroalgal biology. Undoubtedly, both endogenous (e.g. bacteria: Spoerner et al., 2012; circadian rhythm: Cunningham & Guiry, 1989) and abiotic environmental
factors (light, temperature, sea currents) are required (Fig. 3), but the intrinsic signalling pathways
are largely unknown. To understand how to manipulate hatchery culture conditions to give
juveniles the best start in life in tune with aquaculture demands, additional studies assessing the
molecular impact of the surrounding physical and chemical environment (light, nutrients, salinity,
water movement) are required. In some seaweeds, complex interactions with bacteria are a
prerequisite for proper cell growth and differentiation into specific tissues (Goecke et al., 2010).
This has been well-illustrated in green seaweeds (Ulva and Monostroma - Matsuo et al., 2005;
Spoerner et al., 2012), as well as in brown algal species where bacteria might control their life
cycle (Tapia et al., 2016) and their morphology in waters with different salinities (Dittami et al.,
2014). It is tempting to hypothesize that controlling macroalgal development with bacteria will
direct the chemical composition of the macroalga and its value as cash crop. This is mainly relevant
for land-based aquaculture starting with a defined seed-stock (axenic germlings) and a synthetic
microbiome, which could influence the production of primary and secondary metabolites.
However, further work determining macroalgal-bacterial interactions throughout algal life-cycles is
necessary to discriminate between mutualistic, beneficial or pathogenic interactions.

208 Current technological requirements

Reliable, cost-effective and long-term maintenance of genetic resources is a major requirement to ensure the sustainability of the quality of the exploited traits (biomass yield, quality of extracted polysaccharides, texture and taste of species for human consumption; Chapman *et al.*, 2015). Both sub-culturing of macroalgal explants and cryopreservation of macroalgal omnipotent cells are current techniques to vegetatively propagate macroalgae over time. However, sub-cultivation is time-consuming and re-iteration of the protocol over years is a source of bacterial or fungal contamination. Long-term preservation (through refrigeration or liquid-nitrogen freezing) of commercially important seaweed explants has therefore received increasing attention and several
217 protocols are now available. Techniques depend on the species (e.g. gametophytic filaments of
218 *Macrocystis*; Barrento *et al.*, 2016; pieces of *Ulva* thalli; Lee & Nam, 2016; and apical meristems
219 of *Gracilaria*: Lalrinsanga *et al.*, 2009) and a better knowledge of both the mitotic activities within
220 the thallus and the underlying molecular mechanisms governing cell proliferation versus cell
221 differentiation would accelerate the assessment of the regenerative potential of these seaweeds and
222 the necessary development of adequate protocols (Stacey & Day, 2014) (Fig. 3). Basic research has
223 revealed specificities in brown seaweeds, specifically in the *Fucus* embryo, where cell division is
224 subject to distinct control mechanisms compared to other eukaryotes (Corellou *et al.*, 2001). As
225 bacteria play a crucial role in many algal developmental processes (Goecke *et al.*, 2010),
226 macroalgal preservation should also consider cryopreservation of algae with their natural
227 microbiome rather than axenic explants. Therefore, development of seaweed biobanking
228 procedures may be pivotal to meet future aquaculture demands.

229 Beyond cryopreservation, while some techniques are easily transferable from land plants to
230 macroalgae, others require species-specific optimization. The impact of the sea water medium on
231 the ionic concentration of buffers used in standard lab protocols and the different polysaccharide
232 compositions of red and brown algal cell walls (Deniaud-Bouët et al., 2014; Popper et al., 2011)
233 require different cell wall enzymolytic treatments in cytology protocols (Joubert & Fleurence,
234 2008). At the genetic level, the sequence of reporter genes commonly used in other organisms
235 require modification for transgene expression, because of differing codon usages, as shown in red
236 and green seaweeds (Uji et al., 2014; Oertel et al., 2015). The growing interest of the evolutionary
237 developmental biology (“evo-devo”) community in macroalgae would help phycologists develop
238 these techniques further.

239 In addition to the requirement for cell biology and genetic adjustments, ‘OMICS’ technology must
240 be adapted to the level of analysis required to tackle developmental mechanisms taking place at the
241 microscopic and early developmental stages (Fig. 2 and 3). Several transcriptomic (Wang et al.,
242 2015), proteomic (Qian et al., 2016) and metabolomic (Kumar et al., 2016 and references therein)
studies have been reported in both model and exploited macroalgae. In addition, exo-metabolomic profiling in standardized *Ulva* cultures with a designed microbiome have shown growth phase-dependent biomarkers that might be relevant for aquaculture (Alsufyani *et al.*, 2017). Such analyses are assisted by an increasing number of sequenced macroalgal genomes. Currently 18 public algal nuclear genomes have been sequenced, including four seaweeds. However, "-OMICS" studies at early developmental stages are hampered by a scarcity of tissue. While proteomics and metabolomics still require a significant biomass, transcriptomics can bypass this handicap through RNA amplification. Cell-specific expression patterns were thereby obtained using laser microdissection prior to RNA amplification on the model brown seaweed *Ectocarpus* (Saint-Marcoux *et al.*, 2015), and this technology is easily transferable to larger seaweeds.

Finally, transgenesis will be a highly valuable tool to discover how molecular processes are regulated in seaweeds, and to interfere with these processes by knocking down/upregulating endogenous genes. So far, only four multicellular algae, namely *Ulva*, *Pyropia* (*Porphyra*), *Volvox* and *Gonium* are genetically transformable (Schiedlmeier *et al.*, 1994; Oertel *et al.*, 2015; Mikami, 2014; Lerche & Hallmann, 2009), and *Ulva* is the only stably transformable seaweed (Oertel *et al.*, 2015). These first successes must now be replicated in additional, diverse species, via investment of time and expertise.

## Conclusion

A range of protocols are available to cultivate seaweeds, thanks to previous physiological studies carried out in an applied phycological context. Building on this key achievement, practices must be refined and developed with a more focused and on-demand approach. Indeed, demand from end-users is rising for new, high-commercial potential (mainly for food) seaweeds. However, because of their low production level, these seaweeds have not received high investment so far, and as a result, no standardised cultivation and preservation protocols exist. This second big step is much
267 more delicate, because of the greater number of species and of their reluctance to respond to the
268 simplest, classical protocols. The time has come, now that the first empirical studies have been
269 carried out, to engage the community in an in-depth study of the biological processes driving the
270 whole macroalgal life-cycle, from fertilization to the production of organisms. This must respond
271 to end-users' expectations of robustness against environmental constraints (e.g. climate, infection,
272 mechanical strain), biochemical composition and also natural and nature-friendly production
273 increasingly favoured by the consumers. This is even more necessary since, despite the benefit that
274 the development of cutting-edge technologies in animals and plants can bring to the sector, many of
275 these technologies need to be adapted to macroalgae because of their specific ecological niche
276 (highly saline) and their biology (in part due to their phylogenetic distance from better-known
277 organisms). Therefore, efforts must be intensified to fill the gaps in our fundamental knowledge of
278 macroalgal developmental mechanisms. We also believe that the scientific community of land plant
279 researchers will benefit from a deeper understanding of seaweed developmental biology.

280 Acknowledgement

281 This work is carried out in the frame of “Phycomorph” (http://www.phycomorph.org/), a network
282 funded by the COST Association (http://www cost.eu/COST_Actions/fa/FA1406).

283 Author contributions

284 All authors contributed to the writing of the manuscript.
285 **References**

286 **Abreu M, Pereira R, Sassi J-F. 2014.** Marine algae and the global food industry. In: Pereira L, In: Neto J, eds. Marine Algae. CRC Press, 300–319.

288 **Alsufyani T, Weiss A, Wichard T. 2017.** Time course exo-metabolomic profiling in the green marine macroalga *Ulva* (Chlorophyta) for identification of growth phase-dependent biomarkers. *Marine Drugs*, **15**: 14.

291 **Arkema KK, Guannel G, Verutes G, Wood SA, Guerry A, Ruckelshaus M, Kareiva P, Lacayo M, Silver JM. 2013.** Coastal habitats shield people and property from sea-level rise and storms. *Nature Climate Change* **3**: 913–918.

294 **Arun A, Peters NT, Scornet D, Peters AF, Cock JM, Coelho SM. 2013.** Non-cell autonomous regulation of life cycle transitions in the model brown alga *Ectocarpus*. *New Phytologist* **197**: 503–510.

297 **Barrento S, Camus C, Sousa-Pinto I, Buschmann AH. 2016.** Germplasm banking of the giant kelp: Our biological insurance in a changing environment. *Algal Research-Biomass Biofuels and Bioproducts* **13**: 134–140.

300 **Bobadilla F, Rodriguez-Tirado C, Imarai M, Jose Galotto M, Andersson R. 2013.** Soluble beta-1,3/1,6-glucan in seaweed from the southern hemisphere and its immunomodulatory effect. *Carbohydrate Polymers* **92**: 241–248.

303 **Bogaert KA, Beeckman T, De Clerck O. 2017.** Two-step cell polarization in algal zygotes. *Nature Plants* **3**: 16221.

305 **Boland W. 1995.** The chemistry of gamete attraction: chemical structures, biosynthesis, and (a)biotic degradation of algal pheromones. *Proceedings of the National Academy of Sciences of the United States of America* **92**: 37–43.
308 **Bothwell JHF, Kisielewska J, Genner MJ, McAinsh MR, Brownlee C. 2008.** Ca2+ signals 309 coordinate zygotic polarization and cell cycle progression in the brown alga *Fucus serratus*. 310 *Development* (*Cambridge, England*) **135**: 2173–2181.

311 **Brownlee C, Bouget FY, Corellou F. 2001.** Choosing sides: establishment of polarity in zygotes 312 of fucoid algae. *Seminars in Cell & Developmental Biology* **12**: 345–351.

313 **Chapman AS, Stevant P, Larssen WE. 2015.** Food or fad? Challenges and opportunities for 314 including seaweeds in a Nordic diet. *Botanica Marina* **58**: 423–433.

315 **Charrier B, Rolland E, Gupta V, Reddy CRK. 2015.** Production of genetically and 316 developmentally modified seaweeds: exploiting the potential of artificial selection techniques. 317 *Frontiers in Plant Science* **6**: 127.

318 **Coelho SM, Godfroy O, Arun A, Le Corguillé G, Peters AF, Cock JM. 2011.** *OUROBOROS* is 319 a master regulator of the gametophyte to sporophyte life cycle transition in the brown alga 320 *Ectocarpus*. *Proceedings of the National Academy of Sciences of the United States of America* **108**: 321 11518–11523.

322 **Coelho SM, Peters AF, Charrier B, Roze D, Destombe C, Valero M, Cock JM. 2007.** Complex 323 life cycles of multicellular eukaryotes: new approaches based on the use of model organisms. *Gene* 324 **406**: 152–170.

325 **Corellou F, Brownlee C, Detivaud L, Kloareg B, Bouget FY. 2001.** Cell cycle in the *Fucus* 326 zygote parallels a somatic cell cycle but displays a unique translational regulation of cyclin- 327 dependent kinases. *The Plant Cell* **13**: 585–598.

328 **Cottier-Cook E, Nagabhatla N, Badis Y, Campbell ML, Chopin T, Dai W, Fang J, He P, 329 Hewitt CL, Kim GH, et al. 2016.** Safeguarding the future of the global seaweed aquaculture 330 industry. United Nations University (INWEH) and Scottish Association for Marine Science Policy 331 Brief. ISBN 978-92-808-6080-1. 12pp.
332 **Cunningham EM, Guiry MD. 1989.** A Circadian Rhythm in the Long-Day Photoperiodic
333 Induction of Erect Axis Development in the Marine Red Alga *Nemalion Helminthoides*1,2.
334 *Journal of Phycology* 25: 705–712.

335 **Deniaud-Bouët E, Kervarec N, Michel G, Tonon T, Kloareg B, Hervé C. 2014.** Chemical and enzymatic fractionation of cell walls from Fucales: insights into the structure of the extracellular matrix of brown algae. *Annals of Botany* 114: 1203–1216.

338 **Dittami SM, Barbeyron T, Boyen C, Cambefort J, Collet G, Delage L, Gobet A, Groisillier A,**
339 **Leblanc C, Michel G, et al. 2014.** Genome and metabolic network of ‘Candidatus
340 *Phaeomarinobacter ectocarpi*’ Ec32, a new candidate genus of Alphaproteobacteria frequently
341 associated with brown algae. *Frontiers in Genetics* 5.

342 FAO. 2016. Fisheries & Aquaculture - Fisheries and Aquaculture Fact Sheets.

343 **Fernand F, Israel A, Skjermo J, Wichard T, Timmermans KR, Golberg A. 2017.** Offshore macroalgae biomass for bioenergy production: Environmental aspects, technological achievements and challenges. *Renewable and Sustainable Energy Reviews* 75: 35–45.

346 **García-Jiménez P, Brito-Romano O, Robaina RR. 2016.** Occurrence of jasmonates during cystocarp development in the red alga *Grateloupia imbricata*. *Journal of Phycology* 52: 1085–1093.

349 **Goecke F, Labes A, Wiese J, Imhoff JF. 2010.** Chemical interactions between marine macroalgae and bacteria. *Marine Ecology Progress Series* 409: 267–299.

351 **de Groot R, Brander L, van der Ploeg S, Costanza R, Bernard F, Braat L, Christie M,**
352 **Crossman N, Ghermandi A, Hein L, et al. 2012.** Global estimates of the value of ecosystems and
353 their services in monetary units. *Ecosystem Services* 1: 50–61.

354 **Gupta V, Kumari P, Reddy CRK. 2015.** Development and characterization of somatic hybrids of
355 *Ulva reticulata* Forsskal (x) *Monostroma oxyspermum* (Kutz.) Doty. *Frontiers in Plant Science* 6:
356 3.
357 **Gwo HH, Chen CS. 1999.** Evidence of fusion and artificially initialized parthenogenesis in 358 isogametes of *Endarachne binghamiae* (Scytosiphonaceae, Phaeophyta) from Taiwan. *Journal of 359 Applied Phycology*. **11**: 567–577.

360 **Han JW, Klochkova TA, Shim J, Nagasato C, Motomura T, Kim GH. 2014.** Identification of 361 three proteins involved in fertilization and parthenogenetic development of a brown alga, 362 *Scytosiphon lomentaria*. *Planta* **240**:1253–1267.

363 **He J, Chen JP. 2014.** A comprehensive review on biosorption of heavy metals by algal biomass: 364 Materials, performances, chemistry, and modeling simulation tools. *Bioresource Technology* **160**: 365 67–78.

366 **He P, Yarish C. 2006.** The developmental regulation of mass cultures of free-living conchocelis for 367 commercial net seeding of *Porphyra leucosticta* from Northeast America. *Aquaculture* **257**: 373– 381.

369 **Holdt SL, Edwards MD. 2014.** Cost-effective IMTA: a comparison of the production efficiencies 370 of mussels and seaweed. *Journal of Applied Phycology* **26**: 933–945.

371 **Joubert Y, Fleurence J. 2008.** Simultaneous extraction of proteins and DNA by an enzymatic 372 treatment of the cell wall of *Palmaria palmata* (Rhodophyta). *Journal of Applied Phycology* **20**: 373 55–61.

374 **Kamiya M, West JA. 2008.** Origin of apomictic red algae: outcrossing studies of different strains 375 in *Caloglossa monotricha* (Ceramiales, Rhodophyta). *Journal of Phycology* **44**: 977–984.

376 **Krause-Jensen D, Duarte CM. 2016.** Substantial role of macroalgae in marine carbon 377 sequestration. *Nature Geoscience* **9**: 737–742.

378 **Krumhansl KA, Scheibling RE. 2012.** Detrital subsidy from subtidal kelp beds is altered by the 379 invasive green alga *Codium fragile ssp fragile*. *Marine Ecology Progress Series* **456**: 73–85.

380 **Kumar M, Kuzhiumparambil U, Pernice M, Jiang Z, Ralph PJ. 2016.** Metabolomics: an 381 emerging frontier of systems biology in marine macrophytes. *Algal Research-Biomass Biofuels 382 and Bioproducts* **16**: 76–92.
383 Lalrinsanga PL, Deshmukhe G, Chakraborty SK, Dwivedi A, Barman N, Kumar N. 2009.
384 Preliminary studies on cryopreservation of vegetative thalli of some economically important
385 seaweeds of India. *Journal of Applied Aquaculture* 21: 250–262.

386 Largo DB, Fukami K, Nishijima T, Ohno M. 1995. Laboratory-induced development of the ice-
387 ice disease of the farmed red algae *Kappaphycus alvarezii* and *Eucheuma denticulatum*
388 (Solieriaceae, Gigartinales, Rhodophyta). *Journal of Applied Phycology* 7: 539–543.

389 Lee YN, Nam KW. 2016. Cryopreservation of gametophytic thalli of *Ulva prolifera* (Ulvales, Chlorophyta) from Korea. *Journal of Applied Phycology* 28: 1207–1213.

391 Lerche K, Hallmann A. 2009. Stable nuclear transformation of *Gonium pectorale*. *BMC Biotechnology* 9: 64.

393 Li J, Pang S, Shan T, Liu F, Gao S. 2014. Zoospore-derived monoecious gametophytes in
394 *Undaria pinnatifida* (Phaeophyceae). *Chinese Journal of Oceanology and Limnology* 32: 365–371.

395 Li Y, Zhou C, Yan X, Zhang J, Xu J. 2016. Simultaneous analysis of ten phytohormones in
396 *Sargassum horneri* by high-performance liquid chromatography with electrospray ionization
397 tandem mass spectrometry. *Journal of Separation Science* 39: 1804–1813.

398 Loureiro R, Gachon CMM, Rebours C. 2015. Seaweed cultivation: potential and challenges of
399 crop domestication at an unprecedented pace. *New Phytologist* 206: 489–492.

400 Luning K, Dring MJ. 1975. Reproduction, growth and photosynthesis of gametophytes of
401 *Laminaria saccharina* grown in blue and red-light. *Marine Biology* 29: 195–200.

402 Makkar HPS, Tran G, Heuzé V, Giger-Reverdin S, Lessire M, Lebas F, Ankers P. 2016.
403 Seaweeds for livestock diets: A review. *Animal Feed Science and Technology* 212: 1–17.

404 Matsuo Y, Imagawa H, Nishizawa M, Shizuri Y. 2005. Isolation of an algal morphogenesis
405 inducer from a marine bacterium. Science (New York, N.Y.) 307: 1598.

406 Mazarrasa I, Olsen YS, Mayol E, Marbà N, Duarte CM. 2013. Rapid growth of seaweed
407 biotechnology provides opportunities for developing nations. *Nature Biotechnology* 31: 591–592.
408 **Mazarrasa I, Olsen YS, Mayol E, Marbà N, Duarte CM. 2014.** Global unbalance in seaweed production, research effort and biotechnology markets. *Biotechnology Advances* **32**: 1028–1036.

410 **Mikami K. 2014.** A technical breakthrough close at hand: feasible approaches toward establishing a gene-targeting genetic transformation system in seaweeds. *Frontiers in Plant Science* **5**: 498.

412 **Nakahara H. 1984.** Alternation of generations of some brown algae in unialgal and axenic cultures. 北海道大學理學部海藻研究所歐文報告 = Scientific papers of the Institute of Algological Research, Faculty of Science, Hokkaido University **7**: 77–194.

415 **Neveux N, Magnusson M, Mata L, Whelan A, de Nys R, Paul NA. 2016.** The treatment of municipal wastewater by the macroalga *Oedogonium* sp. and its potential for the production of biocrude. *Algal Research* **13**: 284–292.

418 **Oertel W, Wichard T, Weissgerber A. 2015.** Transformation of *Ulva Mutabilis* (chlorophyta) by Vector Plasmids Integrating into the Genome. *Journal of Phycology* **51**: 963–979.

420 **Pang SJ, Lüning K. 2004.** Breaking seasonal limitation: year-round sporogenesis in the brown alga *Laminaria saccharina* by blocking the transport of putative sporulation inhibitors. *Aquaculture* **240**(1-4):531–541.

423 **Pang SJ, Lüning K. 2006.** Tank cultivation of the red alga *Palmaria palmata*: Year-round induction of tetrasporangia, tetraspore release in darkness and mass cultivation of vegetative thalli. *Aquaculture* **252**: 20–30.

426 **Parfrey LW, Lahr DJG, Knoll AH, Katz LA. 2011.** Estimating the timing of early eukaryotic diversification with multigene molecular clocks. *Proceedings of the National Academy of Sciences of the United States of America* **108**: 13624–13629.

429 **Popper S, Michel G, Herve C, Domozych DS, Willats WGT, Tuohy MG, Kloareg B, Stengel DB. 2011.** Evolution and diversity of plant cell walls: from algae to flowering plants. In: Merchant SS, In: Briggs WR, In: Ort D, eds. Annual Review of Plant Biology, Vol 62. Palo Alto: *Annual Reviews*, 567–588.
433 **Qian W-G, Li N, Lin L-D, Xu T, Zhang X, Wang L-H, Zou H-X, Wu M-J, Yan X-F. 2016.**
Parallel analysis of proteins in brown seaweed *Sargassum fusiforme* responding to hyposalinity
stress. *Aquaculture* **465**: 189–197.

436 **Rebours C, Marinho-Soriano E, Zertuche-González JA, Hayashi L, Vásquez JA, Kradolfer P,**
**Soriano G, Ugarte R, Abreu MH, Bay-Larsen I, et al. 2014.** Seaweeds: an opportunity for
wealth and sustainable livelihood for coastal communities. *Journal of Applied Phycology* **26**:
1939–1951.

440 **Reisewitz SE, Estes JA, Simenstad CA. 2006.** Indirect food web interactions: sea otters and kelp
forest fishes in the Aleutian archipelago. *Oecologia* **146**: 623–631.

442 **Rorrer GL, Cheney DP. 2004.** Bioprocess engineering of cell and tissue cultures for marine
seaweeds. *Aquacultural Engineering* **32**: 11–41.

444 **Saint-Marcoux D, Billoud B, Langdale JA, Charrier B. 2015.** Laser capture microdissection in
*Ectocarpus siliculosus*: the pathway to cell-specific transcriptomics in brown algae. *Frontiers in
Plant Science* **6**: 54.

447 **Schiedlmeier B, Schmitt R, Muller W, Kirk M, Gruber H, Mages W, Kirk D. 1994.** Nuclear
Transformation of *Volvox carteri*. *Proceedings of the National Academy of Sciences of the United
States of America* **91**: 5080–5084.

450 **Schmid C, Schroer N, Muller D. 1994.** Female gamete membrane-glycoproteins potentially
involved in gamete recognition in *Ectocarpus siliculosus* (phaeophyceae). *Plant Science* **102**: 61–
67.

453 **Seghetta M, Tørring D, Bruhn A, Thomsen M. 2016.** Bioextraction potential of seaweed in
Denmark - An instrument for circular nutrient management. *The Science of the Total Environment*
**563–564**: 513–529.

456 **Shan T, Pang S, Li J, Li X, Su L. 2015.** Construction of a high-density genetic map and mapping
of a sex-linked locus for the brown alga *Undaria pinnatifida* (Phaeophyceae) based on large scale
458 marker development by specific length amplified fragment (SLAF) sequencing. BMC Genomics 16.

460 **Shan TF, Pang SJ, Gao, SQ. 2013.** Novel means for variety breeding and sporeling production in the brown seaweed *Undaria pinnatifida* (Phaeophyceae): Crossing female gametophytes from parthenosporophytes with male gametophyte clones. *Phycological Research* **61**: 154–61

463 **Spoerner M, Wichard T, Bachhuber T, Stratmann J, Oertel W. 2012.** *Ulva mutabilis* (Chlorophyta) depends on a combination of two bacterial species excreting regulatory factors. *Journal of Phycology* **48**: 1433–1447.

466 **Stacey GN, Day JG. 2014.** Putting cells to sleep for future science. *Nature Biotechnology* **32**: 320–322.

468 **Steneck RS, Graham MH, Bourque BJ, Corbett D, Erlandson JM, Estes JA, Tegner MJ. 2002.** Kelp forest ecosystems: biodiversity, stability, resilience and future. *Environmental Conservation* **29**: 436–459.

471 **Tapia JE, González B, Goulitquer S, Potin P, Correa JA. 2016.** Microbiota influences morphology and reproduction of the brown alga *Ectocarpus* sp. *Frontiers in Microbiology* 7.

473 **Uji T, Hirata R, Fukuda S, Mizuta H, Saga N. 2014.** A codon-optimized bacterial antibiotic gene used as selection marker for stable nuclear transformation in the marine red alga *Pyropia yezoensis*. *Marine Biotechnology* (New York, N.Y.) **16**: 251–255.

476 **Vesty EF, Kessler RW, Wichard T, Coates JC. 2015.** Regulation of gametogenesis and zoosporogenesis in *Ulva linza* (Chlorophyta): comparison with *Ulva mutabilis* and potential for laboratory culture. *Frontiers in Plant Science* **6**: 15.

479 **Wang L, Mao Y, Kong F, Cao M, Sun P. 2015.** Genome-wide expression profiles of *Pyropia haitanensis* in response to osmotic stress by using deep sequencing technology. *BMC Genomics* **16**: 1012.
482

Wichard T, Oertel W. 2010. Gametogenesis and gamete release of *Ulva mutabilis* and *Ulva lactuca* (Chlorophyta): Regulatory effects and chemical characterization of the 'swarming inhibitor'. *Journal of Phycology* 46: 248-259.

483

484

485

Zhou W, Sui Z, Wang J, Chang L. 2013. An orthogonal design for optimization of growth conditions for all life history stages of *Gracilariopsis lemaneiformis* (Rhodophyta). *Aquaculture* 392: 98-105.

486

487

488

Box and Figure legends

489

**Box 1: Life cycle stages in seaweeds and possible manipulations**

490

491

492

493

494

495

496

497

498

499

500

501

502

503

Seaweed life cycles comprise several (usually 4) multicellular phases, including vegetative and
fertile sporophytes and vegetative and fertile gametophytes (grey boxes). On the left, grey arrows
indicate the different natural alternatives that seaweeds can use to reproduce (either sexually or
asexually). On the right, brown, red and green horizontal lines represent the 3 groups of seaweeds.
Transition between two successive phases, and bypassing or maintenance of one phase (either by
delaying the maturation of the organism or by asexual looping) are ways to exert a tight control on
the life cycle. Straight arrows indicate controls over a given phase of the life cycle (maintenance,
induction or inhibition). Dashed arrows indicate asexual looping. A few specific examples are
represented by the numbers that follow. [1] vertical arrow: maintaining vegetative growth of the
brown seaweed *Saccharina latissima* gametophytes under red light or by sub-culturing (grinding)
filaments; horizontal arrow: induction of gametophyte fertility under blue light (Luning & Dring,
1975). [2] sporulation maintenance by removal of the basal meristem of *S. latissima* (Pang &
Lüning, 2004). [3] maintenance of the vegetative stage of the sporophyte: in *Porphyra conchocelis*
by temperature, photoperiod and irradiance (He & Yarish, 2006); of the reproductive stage of the
sporophyte: in *Palmaria* tetrasporophytes by short daylength (Pang & Lüning, 2006). [4] control of
the shift to the reproductive phase of the vegetatively propagated *Gracilariopsis* gametophyte by
511
512
temperature optimisation (Zhou *et al.*, 2013). [5] identification of sporulation-inhibiting factors
513
(Glycoprotein SP-1 and low molecular weight factor SP-2) from *Ulva* gametophytes and
514
sporophytes (Wichard & Oertel, 2010; Vesty *et al.*, 2015). [6] parthenogenesis in brown algae
515
(Nakahara, 1984) and red algae (*Undaria* female spore seeding; Shan *et al.*, 2013). [7] production
516
of gametophytes from gametes of the *Ectocarpus siliculosus* mutant *ouroboros* (Coelho *et al.*,
517
2011). [8] production of *Ulva* gametophytes from the germination of its own gametes when
518
separated from another mating type (Wichard & Oertel, 2010). [9] germination of
519
parthenosporophytes (instead of gametophytes in this strain) from *Ectocarpus* zoospores by an
520
inhibiting factor produced by the parthenosporophyte (Arun *et al.*, 2013).
521

522 **Figure 1: Position of macroalgae in the scientific and societal landscapes.**

Macroalgae grow rapidly in a wide range of temperatures, using only sunlight, atmospheric carbon
523
and naturally nutritious coastal waters. They are therefore valuable feedstock for the production of
524
food, feed, biofuel, hydrocolloids, fertilisers, cosmetics, probiotics, biodegradable packaging
525
through aquaculture and IMTA (see text for details). They provide curative ecological roles
526
necessitated by human activities (waste-water treatments and seabed recolonisation). Ecology also
527
benefits from a knowledge of macroalgal reproductive mechanisms via a better understanding of
528
dispersion and persistence of both natural and exotic populations. This also contributes to the
529
development of conservation protocols for threatened or susceptible populations. Because their life
530
histories differ from land plants, macroalgae also inspire molecular evo-devo studies involving the
531
whole green lineage.
532
**Figure 2: Importance of the microscopic early developmental stages in the life cycle of exploited seaweeds: Example of the kelp *Saccharina latissima*.

Production of kelp (large brown macroalga) sporophyte juveniles takes place in hatcheries under controlled growth conditions. Cultures of microscopic male and female gametophytes are produced from spores of macroscopic, mature plants collected from the sea. Gametophyte cultures are grown to fertility under controlled temperature and light conditions (see Box 1 for details). Microscopic, fertile, recently fertilised gametophytes, or (in turn) juvenile sporophytes are spread onto cultivation support materials (ropes or 2D substrates), which are subsequently deployed into the sea. Photos kindly provided by Teis Boderskov (Aarhus University, Denmark) and Eric Tamigneaux (Merinov, Canada).

**Figure 3: Scope of beneficial outflow from basic research to seaweed aquaculture.**

Sexual reproduction (top right) gives rise to polarised embryos (left), which progressively grow and differentiate, giving tissues and organs with specific shape and cellular functions (e.g. blade, stipe, holdfast, reproductive organs). The study of the different steps of the life cycle (here simplified, with adult representing either the sporophyte or the gametophyte) at the basic level (in blue) can lead to the control and improvement of key processes in seaweed aquaculture (in green).

In hatcheries, density of juveniles on the cultivation support material depends on both the fertilisation rate and the adhesive potential of the embryos. Fertilisation rate itself depends on the physical interactions between the two gametes (taxis, specific recognition and membrane fusion).

Better knowledge of the cell cycle and characterisation of the pluripotent cells (zygotes, meristems) will both contribute to develop cryopreservation protocols. Metabolic patterning of seaweed organs and tissues, mediated by molecular, biochemical or cellular markers, will assist farmers in
555 monitoring seaweed growth and fitness both in hatcheries and in the field. All these processes are

556 under the control of abiotic and biotic factors (see text and Box 1 for references).
Box 1

<figure><img src="image_2.png" /><figcaption>Box 1: Life cycle stages in seaweeds and possible manipulations</figcaption></figure>

Seaweed life cycles comprise several (usually 4) multicellular phases, including vegetative and fertile
sporophytes and vegetative and fertile gametophytes (grey boxes). On the left, grey arrows indicate the
different natural alternatives that seaweeds can use to reproduce (either sexually or asexually). On the
right, brown, red and green horizontal lines represent the 3 groups of seaweeds. Transition between two
successive phases, and bypassing or maintenance of one phase (either by delaying the maturation of the
organism or by asexual looping) are ways to exert a tight control on the life cycle. Straight arrows indicate
controls over a given phase of the life cycle (maintenance, induction or inhibition). Dashed arrows indicate
asexual looping. A few specific examples are represented by the numbers that follow. [1] vertical arrow:
maintaining vegetative growth of the brown seaweed Saccharina latissima gametophytes under red light or
by sub-culturing (grinding) filaments; horizontal arrow: induction of gametophyte fertility under blue light
(Luning & Dring, 1975). [2] sporulation maintenance by removal of the basal meristem of S. latissima (Pang
& Lüning, 2004). [3] maintenance of the vegetative stage of the sporophyte: in Porphyra conchocelis by
temperature, photoperiod and irradiance (He & Yarish, 2006); of the reproductive stage of the sporophyte:
in Palmaria tetrasporophytes by short daylength (Pang & Lüning, 2006). [4] control of the shift to the
reproductive phase of the vegetatively propagated Gracilariopsis gametophyte by temperature optimisation
(Zhou et al., 2013). [5] identification of sporulation-inhibiting factors (Glycoprotein SP-1 and low molecular
weight factor SP-2) from Ulva gametophytes and sporophytes (Wichard & Oertel, 2010; Vesty et al., 2015).
[6] parthenogenesis in brown algae (Nakahara, 1984) and red algae (Undaria female spore seeding; Shan et
al., 2013). [7] production of gametophytes from gametes of the Ectocarpus siliculosus mutant ouroboros (Coelho et al., 2011). [8] production of Ulva gametophytes from the germination of its own gametes when separated from another mating type (Wichard & Oertel, 2010). [9] germination of parthenosporophytes (instead of gametophytes in this strain) from Ectocarpus zoospores by an inhibiting factor produced by the parthenosporophyte (Arun et al., 2013).

<figure><img src="image_3.png" /></figure>
<figure><img src="image_4.png" /><figcaption>Figure 1: Position of macroalgae in the scientific and societal landscapes.</figcaption></figure>

Macroalgae grow rapidly in a wide range of temperatures, using only sunlight, atmospheric carbon and naturally nutritious coastal waters. They are therefore valuable feedstock for the production of food, feed, biofuel, hydrocolloids, fertilisers, cosmetics, probiotics, biodegradable packaging through aquaculture and IMTA (see text for details). They provide curative ecological roles necessitated by human activities (wastewater treatments and seabed recolonisation). Ecology also benefits from a knowledge of macroalgal reproductive mechanisms via a better understanding of dispersion and persistence of both natural and exotic populations. This also contributes to the development of conservation protocols for threatened or susceptible populations. Because their life histories differ from land plants, macroalgae also inspire molecular evo-devo studies involving the whole green lineage.
<figure><img src="image_5.png" /><figcaption>Figure 2</figcaption></figure>

Figure 2: Importance of the microscopic early developmental stages in the life cycle of exploited seaweeds: Example of the kelp Saccharina latissima.

Production of kelp (large brown macroalga) sporophyte juveniles takes place in hatcheries under controlled growth conditions. Cultures of microscopic male and female gametophytes are produced from spores of macroscopic, mature plants collected from the sea. Gametophyte cultures are grown to fertility under controlled temperature and light conditions (see Box 1 for details). Microscopic, fertile, recently fertilised gametophytes, or (in turn) juvenile sporophytes are spread onto cultivation support materials (ropes or 2D substrates), which are subsequently deployed into the sea. Photos kindly provided by Teis Boderskov (Aarhus University, Denmark) and Eric Tamigneaux (Merinov, Canada).
<figure><img src="image_6.png" /><figcaption>Figure 3</figcaption></figure>

Figure 3: Scope of beneficial outflow from basic research to seaweed aquaculture.

Sexual reproduction (top right) gives rise to polarised embryos (left), which progressively grow and differentiate, giving tissues and organs with specific shape and cellular functions (e.g. blade, stipe, holdfast, reproductive organs). The study of the different steps of the life cycle (here simplified, with adult representing either the sporophyte or the gametophyte) at the basic level (in blue) can lead to the control and improvement of key processes in seaweed aquaculture (in green).

In hatcheries, density of juveniles on the cultivation support material depends on both the fertilisation rate and the adhesive potential of the embryos. Fertilisation rate itself depends on the physical interactions between the two gametes (taxis, specific recognition and membrane fusion). Better knowledge of the cell cycle and characterisation of the pluripotent cells (zygotes, meristems) will both contribute to develop cryopreservation protocols. Metabolic patterning of seaweed organs and tissues, mediated by molecular, biochemical or cellular markers, will assist farmers in monitoring seaweed growth and fitness both in hatcheries and in the field. All these processes are under the control of abiotic and biotic factors (see text and Box 1 for references).

<figure><img src="image_7.png" /></figure>