Microtubule reorganization during mitotic cell division in the dinoflagellate Ostreospis cf. ovata.

Dinoflagellates are marine organisms that undergo seasonal proliferation events known as algal blooms. Vegetative cell proliferation is a main contributing factor in these events. However, mechanistical understanding of mitosis and cytokinesis in dinoflagellates remains rudimentary. Using an optimized immunofluorescence protocol, we analysed changes in microtubule organization occurring during the mitotic cycle of the toxic dinoflagellate Ostreopsis cf. ovata. We find that the flagella and the cortical microtubule array persist throughout the mitotic cycle. Two cytoplasmic microtubule bundles originate from the ventral area, where the basal bodies are located - a cortical bundle and a cytoplasmic bundle. The latter associates with the nucleus in the cell centre before mitosis and with the acentrosomal extranuclear spindle during mitosis. Analysis of tubulin post-translational modifications identifies two populations of spindle microtubules - polar acetylated microtubules, whose length is constant, and central tyrosinated microtubules, which elongate during chromosome segregation. During cell division a microtubule-rich structure forms along the dorsal-ventral axis, associated with the site of cytokinesis, consistent with a cytokinetic mechanism that is independent of the actomyosin ring typical of animal and yeast cells.

Rapid Biotic and Abiotic Transformation of Toxins produced by Ostreopsis. cf. ovata.

The dinoflagellate Ostreopsis cf. ovata produces several families of toxic polyketides. Despite only a few field measurements of these phycotoxins in seawater and aerosols, they are believed to be responsible for dermatitis and the toxic inhalations reported during blooms of this species. Therefore, the stability of these compounds in seawater is essential to understanding the causes of these symptoms, however, this has never been assessed. In the current study, the optimization of a solid phase extraction (SPE) procedure was first performed to ensure the most efficient extraction of all phycotoxins known to be produced by this strain, including the recently described liguriatoxins. The SPE cartridge SDBL® under non acidified conditions offered the best option. The stability of the ovatoxins and the liguriatoxins under biotic and abiotic stress was assessed by exposing the spent medium of a culture of Ostreopsis cf. ovata to its bacterial consortium and natural sunlight. A rapid biotic transformation was detected for both families of compounds. When exposed to bacteria, the half-lives of the ovatoxins were reached before 10 h and at 36 h, 97% of these toxins had been transformed. The half-lives of the liguriatoxins were 10 h under these conditions. Photolysis (abiotic degradation) of the ovatoxins (T1/2 < 36 h) was faster than for the liguriatoxins (T1/2 > 62 h). Although none of the catabolites of these phycotoxins were thoroughly identified, an untargeted metabolomics approach combined with molecular networking highlighted the presence of several compounds exhibiting structural similarities with the ovatoxins. Additional work should confirm the preliminary findings on these potential ovatoxins' catabolites and their biological properties. The rapid transformation of O. cf. ovata's phycotoxins introduces questions concerning their presence in seawater and their dispersion in the sea spray aerosols. The compounds involved in the toxic inhalations and dermatitis often experienced by beachgoers may stem from the catabolites of these toxins or even unrelated and as yet unidentified compounds.

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.

In a future scenario of increasing temperatures in North-Atlantic waters, the risk associated with the expansion of the harmful, benthic dinoflagellate Ostreopsis cf. siamensis has to be evaluated and monitored. Microscopy observations and spatio-temporal surveys of environmental DNA (eDNA) were associated with Lagrangian particle dispersal simulations to: (i) establish the current colonization of the species in the Bay of Biscay, (ii) assess the spatial connectivity among sampling zones that explain this distribution, and (iii) identify the sentinel zones to monitor future expansion. Throughout a sampling campaign carried out in August to September 2018, microscope analysis showed that the species develops in the south-east of the bay where optimal temperatures foster blooms. Quantitative PCR analyses revealed its presence across almost the whole bay to the western English Channel. An eDNA time-series collected on plastic samplers showed that the species occurs in the bay from April to September. Due to the water circulation, colonization of the whole bay from the southern blooming zones is explained by inter-site connectivity. Key areas in the middle of the bay permit continuous dispersal connectivity towards the north. These key areas are proposed as sentinel zones to monitor O. cf. siamensis invasions towards the presumably warming water of the North-East Atlantic.

First Evidence of the Presence of Anatoxin-A in Sea Figs Associated with Human Food Poisonings in France.

From January 2011 to March 2018, 26 patients aged from 20 to 80 years old reported being sick in France after eating sea figs of the genus Microcosmus. The patients had symptoms evoking a cerebellar syndrome: blurred or double vision, ataxia and dizziness, asthenia, headache, muscle cramps, paresthesia and digestive disorders (nausea, vomiting and diarrhea). Three of the 18 food poisoning events recorded by the Poison Control Center in Marseille and involving four patients were further investigated as the meal leftovers were collected and analyzed. A previous study ruled out the presence of the regulated lipophilic marine toxins after high-resolution mass spectrometry, but further analyses were required to look for hydrophilic cyanotoxins. The sea fig leftovers from food poisoning case Numbers 1 (January 2011), 6 (December 2012) and 17 (March 2018) of this published case series were analyzed by hydrophilic interaction liquid chromatography coupled to low- and high-resolution mass spectrometry to investigate the presence of hydrophilic cyanotoxins. The sea fig samples showed anatoxin-a (ATX-a) concentrations ranging from 193.7 to 1240.2 µg/kg. The sea fig control sample analyzed was also contaminated with ATX-a but in a much smaller concentration (22.5 µg/kg). To the best of our knowledge, this is the first report of human food poisoning involving ATX-a as the possible causative toxin where the cyanotoxin could be unequivocally identified.

Passive Sampling and High Resolution Mass Spectrometry for Chemical Profiling of French Coastal Areas with a Focus on Marine Biotoxins.

Passive samplers (solid phase adsorption toxin tracking: SPATT) are able to accumulate biotoxins produced by microalgae directly from seawater, thus providing useful information for monitoring of the marine environment. SPATTs containing 0.3, 3, and 10 g of resin were deployed at four different coastal areas in France and analyzed using liquid chromatography coupled to high resolution mass spectrometry. Quantitative targeted screening provided insights into toxin profiles and showed that toxin concentrations and profiles in SPATTs were dependent on the amount of resin used. Between the three amounts of resin tested, SPATTs containing 3 g of resin appeared to be the best compromise, which is consistent with the use of 3 g of resin in SPATTs by previous studies. MassHunter and Mass Profiler Professional softwares were used for data reprocessing and statistical analyses. A differential profiling approach was developed to investigate and compare the overall chemical diversity of dissolved substances in different coastal water bodies. Principal component analysis (PCA) allowed for spatial differentiation between areas. Similarly, SPATTs retrieved from the same location at early, medium, and late deployment periods were also differentiated by PCA, reflecting seasonal variations in chemical profiles and in the microalgal community. This study used an untargeted metabolomic approach for spatial and temporal differentiation of marine environmental chemical profiles using SPATTs, and we propose this approach as a step forward in the discovery of chemical markers of short- or long-term changes in the microbial community structure.

Hunt for Palytoxins in a Wide Variety of Marine Organisms Harvested in 2010 on the French Mediterranean Coast.

During the summer of 2010, 31 species including fish, echinoderms, gastropods, crustaceans, cephalopods and sponges were sampled in the Bay of Villefranche on the French Mediterranean coast and screened for the presence of PLTX-group toxins using the haemolytic assay. Liquid chromatography tandem mass spectrometry (LC-MS/MS) was used for confirmatory purposes and to determine the toxin profile. The mean toxin concentration in the whole flesh of all sampled marine organisms, determined using the lower- (LB) and upper-bound (UB) approach was 4.3 and 5.1 µg·kg(-1), respectively, with less than 1% of the results exceeding the European Food Safety Authority (EFSA) threshold of 30 µg·kg(-1)and the highest values being reported for sea urchins (107.6 and 108.0 µg·kg(-1)). Toxins accumulated almost exclusively in the digestive tube of the tested species, with the exception of octopus, in which there were detectable toxin amounts in the remaining tissues (RT). The mean toxin concentration in the RT of the sampled organisms (fishes, echinoderms and cephalopods) was 0.7 and 1.7 µg·kg(-1) (LB and UB, respectively), with a maximum value of 19.9 µg·kg(-1) for octopus RT. The herbivorous and omnivorous organisms were the most contaminated species, indicating that diet influences the contamination process, and the LC-MS/MS revealed that ovatoxin-a was the only toxin detected.

Complex toxin profile of French Mediterranean Ostreopsis cf. ovata strains, seafood accumulation and ovatoxins prepurification.

Ostreopsis cf. ovata produces palytoxin analogues including ovatoxins (OVTXs) and a putative palytoxin (p-PLTX), which can accumulate in marine organisms and may possibly lead to food intoxication. However, purified ovatoxins are not widely available and their toxicities are still unknown. The aim of this study was to improve understanding of the ecophysiology of Ostreopsis cf. ovata and its toxin production as well as to optimize the purification process for ovatoxin. During Ostreopsis blooms in 2011 and 2012 in Villefranche-sur-Mer (France, NW Mediterranean Sea), microalgae epiphytic cells and marine organisms were collected and analyzed both by LC-MS/MS and hemolysis assay. Results obtained with these two methods were comparable, suggesting ovatoxins have hemolytic properties. An average of 223 µg·kg-1 of palytoxin equivalent of whole flesh was found, thus exceeding the threshold of 30 µg·kg-1 in shellfish recommended by the European Food Safety Authority (EFSA). Ostreopsis cells showed the same toxin profile both in situ and in laboratory culture, with ovatoxin-a (OVTX-a) being the most abundant analogue (~50%), followed by OVTX-b (~15%), p-PLTX (12%), OVTX-d (8%), OVTX-c (5%) and OVTX-e (4%). Ostreopsis cf. ovata produced up to 2 g of biomass per L of culture, with a maximum concentration of 300 pg PLTX equivalent cell-1. Thus, an approximate amount of 10 mg of PLTX-group toxins may be produced with 10 L of this strain. Toxin extracts obtained from collected biomass were purified using different techniques such as liquid-liquid partition or size exclusion. Among these methods, open-column chromatography with Sephadex LH20 phase yielded the best results with a cleanup efficiency of 93% and recovery of about 85%, representing an increase of toxin percentage by 13 fold. Hence, this purification step should be incorporated into future isolation exercises.

Basin scale variability of Ostreopsis spp. blooms provides evidence of effectiveness of an integrated sampling approach.

Ostreopsis spp. blooms have been occurring in the last two decades in the Mediterranean Sea in association with a variety of biotic and abiotic substrata (macroalgae, seagrasses, benthic invertebrates, sand, pebbles and rocks). Cells proliferate attached to the surfaces through mucilaginous trichocysts, which lump together microalgal cells, and can also be found in the plankton and on floating aggregates: such tychoplanktonic behavior makes the quantitative assessment of blooms more difficult than planktonic or benthic ones. Different techniques have been so far applied for quantifying cell abundances of benthic microalgae for research, monitoring and risk assessment purposes. In this context, the Benthic Dinoflagellates Integrator (BEDI), a non-destructive quantification method for benthic dinoflagellate abundances, was developed and tested within the EU ENPI-CBCMED project M3-HABs. This device allows mechanical detachment of cells without collecting the benthic substrate, providing an integrated assessment of both epiphytic and planktonic cells, i.e. of the number of cells potentially made available in the water volume from "resuspension" which could have harmful effects on other organisms (including humans). The present study confirms the effectiveness of the BEDI sampling device across different environments across the Mediterranean Sea and constitutes the first large-scale study of Ostreopsis spp. blooms magnitude in function of different macro- and meso­habitat features across the basin.

Ovatoxin-a and palytoxin accumulation in seafood in relation to Ostreopsis cf. ovata blooms on the French Mediterranean coast.

Dinoflagellates of the genus Ostreopsis are known to cause (often fatal) food poisoning in tropical coastal areas following the accumulation of palytoxin (PLTX) and/or its analogues (PLTX group) in crabs, sea urchins or fish. Ostreopsis spp. occurrence is presently increasing in the northern to north western Mediterranean Sea (Italy, Spain, Greece and France), probably in response to climate change. In France, Ostreopsis. cf. ovata has been associated with toxic events during summer 2006, at Morgiret, off the coast of Marseille, and a specific monitoring has been designed and implemented since 2007. Results from 2008 and 2009 showed that there is a real danger of human poisoning, as these demonstrated bioaccumulation of the PLTX group (PLTX and ovatoxin-a) in both filter-feeding bivalve molluscs (mussels) and herbivorous echinoderms (sea urchins). The total content accumulated in urchins reached 450 µg PLTX eq/kg total flesh (summer 2008). In mussels, the maximum was 230 µg eq PLTX/kg (summer 2009) compared with a maximum of 360 µg found in sea urchins during the same period at the same site. This publication brings together scientific knowledge obtained about the summer development of Ostreopsis spp. in France during 2007, 2008 and 2009.

First insights on the diversity of the genus Ostreopsis (Dinophyceae, Gonyaulacales) in Guadeloupe Island, with emphasis on the phylogenetic position of O. heptagona.

The present study aims to identify epiphytic Ostreopsis cells collected in Guadeloupe between 2017 and 2018 using a morpho-molecular approach. This method combined microscopical observations of wild specimens (light and scanning electron microscopy) with a phylogenetic analysis inferred from concatenated sequences of ribosomal operon (SSU + ITS + LSU) of Ostreopsidoideae. Four distinct morphotypes were identified in our samples and studied by SEM. Molecular data obtained from single-cell PCR for the four morphotypes were consistent with observations and confirmed the presence of three Ostreopsis species resolved in well characterized genotypes (O. cf. ovata, O. lenticularis and O. siamensis) and an unidentified clade. Detailed morphological characters including sulcal plates confirmed the identification of the last morphotype as O. heptagona D.R. Norris, J.W. Bomber & Balech, which forms a new basal clade in the genus, not previously reported. Observations highlighted overlapping sizes for O. lenticularis, O. siamensis and O. heptagona. Direct sequencing of PCR products obtained for some cells of O. lenticularis and O. heptagona collected at one site revealed unexpectedly the presence of the parasitoid dinoflagellate Amoebophrya. Some Ostreopsis cells were found partially emptied and exhibiting a compact mass. Further analyses are needed to understand the ecological role of Amoebophrya on blooms of epiphytic Ostreopsis species.

On the Hunt for New Toxin Families Produced by a Mediterranean Strain of the Benthic Dinoflagellate Ostreopsis cf. ovata.

Ostreopsis cf. ovata is a benthic dinoflagellate known to produce palytoxin (PLTX) and its analogues. Recent investigations suggested the production of unknown toxins by a Mediterranean strain. In the present work, two new families of toxins, potentially novel in their structures, were purified from this same Mediterranean strain of Ostreopsis cf. ovata. The low amount of material isolated only allowed for acquisition of high-resolution mass spectrometry data and the evaluation of their cytotoxicity to human lung cancer cells. Based on their HRMS data, none of these new compounds appear to be close PLTX analogues, although their mass spectra suggest poly-hydroxylated long chain compounds of high molecular weight (1370-2143 Da). The cell cytotoxicity concentrations (CC50) of these new purified toxins ranged between 0.68 and 3.12 µg/mL, and this was enhanced when they were tested as mixtures, suggesting synergistic effects of Ostreopsis toxins. The two families of compounds were named the liguriatoxins (LGTX) and rivieratoxins (RVTX), with each family containing three members. Additional work on purification is needed to fully characterize the structures of these six new dinoflagellate toxins.

The role of habitat in the facilitation of Ostreopsis spp. blooms.

In recent decades, recurrent Ostreopsis spp. blooms have been recorded throughout the globe, causing public health issues and mass mortalities of invertebrates. Ostreopsis species are benthic and develop in shallow waters in close relation with a substrate, but possible substrate preferences are still ambiguous. Bloom develops on both living and dead substrates and several interacting biotic and abiotic factors acting at different spatial scales can potentially foster or regulate Ostreopsis spp. development. The objective of this review is to collect and summarize information on Ostreopsis spp. blooms related to the habitat at different spatial scales, in order to assess preferences and trends. References including Ostreopsis spp. samplings in the field were analysed in this review, as potentially including information about the micro- (substrate), meso­ (community) and macrohabitat (ecosystem) related to Ostreopsis spp. blooms. The sampled substrate and the ecosystem where Ostreopsis spp. were collected were generally reported and described in the studies, while the description of the mesohabitat was rarely reported. Ostreopsis spp. were generally described as attached to biotic substrates and in particular, macroalgae, even in studies conducted in coral reefs, where macroalgae are generally not dominant (but they can be in case of coral reef degradation). In both temperate and tropical areas, Ostreopsis spp. were mostly sampled on algal species usually forming medium or low complexity communities (erect or turf-forming algae), often characteristic from post-regime shift scenarios, and rarely on canopy-forming species (such as fucoids and kelps). This literature review highlights the need of collecting more information about the mesohabitat where important Ostreopsis spp. blooms develop, as much as of the underlying mechanisms driving eventual differences on Ostreopsis spp. abundances. This knowledge would allow a better risk assessment of Ostreopsis spp. blooms, identifying areas at high risk on the base of the benthic habitats.

Environmental, human health and socioeconomic impacts of Ostreopsis spp. Blooms in the NW Mediterranean.

This paper summarizes the research conducted by the partners of the EU co-funded CoCliME project to ascertain the ecological, human health and economic impacts of Ostreopsis (mainly O. cf. ovata) blooms in the NW Mediterranean coasts of France, Monaco and Spain. This knowledge is necessary to design strategies to prevent, mitigate and, if necessary, adapt to the impacts of these events in the future and in other regions. Ostreopsis proliferations in the Mediterranean have been related to massive mortalities of benthic organisms and to symptoms of respiratory and cutaneous irritation in humans. A six-year epidemiologic study in a Ostreopsis hot spot in Catalonia and the accumulated experience of the French Mediterranean National Ostreopsis Surveillance Network confirm the main effects of these blooms on human health in the NW Mediterranean. The impacts are associated to direct exposure to seawater with high Ostreopsis cell concentrations and to inhalation of aerosols containing unknown irritative chemicals produced under certain circumstances during the blooms. A series of mild acute symptoms, affecting the entire body as well as the ophthalmic, digestive, respiratory and dermatologic systems have been identified. A main remaining challenge is to ascertain the effects of the chronic exposure to toxic Ostreopsis blooms. Still, the mechanisms involved in the deletereous effects of Ostreopsis blooms are poorly understood. Characterizing the chemical nature of the harmful compounds synthesized by Ostreopsis as well as the role of the mucus by which cells attach to benthic surfaces, requires new technical approaches (e.g., metabolomics) and realistic and standardized ecotoxicology tests. It is also necessary to investigate how palytoxin analogues produced by O. cf. ovata could be transferred through the marine food webs, and to evaluate the real risk of seafood poisonings in the area. On the other hand, the implementation of beach monitoring and surveillance systems in the summer constitutes an effective strategy to prevent the impacts of Ostreopsis on human health. In spite of the confirmed noxious effects, a survey of tourists and residents in Nice and Monaco to ascertain the socioeconomic costs of Ostreopsis blooms indicated that the occurrence of these events and their impacts are poorly known by the general public. In relationship with a plausible near future increase of Ostreopsis blooms in the NW Mediterranean coast, this survey showed that a substantial part of the population might continue to go to the beaches during Ostreopsis proliferations and thus could be exposed to health risks. In contrast, some people would not visit the affected areas, with the potential subsequent negative impacts on coastal recreational and touristic activities. However, at this stage, it is too early to accurately assess all the economic impacts that a potentially increasing frequency and biogeographic expansion of the events might cause in the future.

Daily variations of Ostreopsis cf. ovata abundances in NW Mediterranean Sea.

Ostreopsis cf. ovata is a benthic dinoflagellate very common in tropical and temperate coastal areas, particularly in the Mediterranean Sea. This species is also found in the plankton, i.e. swimming in the water column or in aggregates floating at the sea surface. The potential links between the planktonic and benthic populations influencing their relative distribution in the water column and attached to the benthic substrate are poorly understood. To shed light on this question, a high-frequency temporal monitoring was conducted in the Villefranche bay (France) to determine the abundance of (1) epibenthic cells attached to macroalgae, (2) planktonic cells in the water column and (3) cells in aggregates floating at the sea water surface (hereafter, referred to sea surface cells) . This monitoring was realized over 3 consecutive years (2018, 2019 and 2020) and at different phases of the bloom (exponential phase - 2020, peak - 2019 and decline phase - 2018). Strong variations in benthic and planktonic O. cf. ovata abundances were observed over the 24 h sampling cycles conducted in three consecutive years. The three populations, planktonic, benthic and sea surface cells, exhibited the highest numbers during the day (light) hours and lowest values at night in 2018 and 2019. In 2020, however, benthic abundances did not differ significantly between light and dark periods. Moreover, epibenthic cells abundances peaked in the morning, followed by the peak of the cells in the plankton and in the surface aggregates during the afternoon. Monitoring of O. cf. ovata is often based on a single sampling per day without precise indications of sampling time and shows great variability in O. cf. ovata abundances. Our observations of daily variations in cell abundances along the water column clearly indicate that time and water column depth of sampling constitute a great source of variability and have to be considered when designing new monitoring strategies to reduce variability and to harmonize data acquisition and international comparisons.

Atypical Membrane-Anchored Cytokine MIF in a Marine Dinoflagellate.

Macrophage Migration Inhibitory Factors (MIF) are pivotal cytokines/chemokines for vertebrate immune systems. MIFs are typically soluble single-domain proteins that are conserved across plant, fungal, protist, and metazoan kingdoms, but their functions have not been determined in most phylogenetic groups. Here, we describe an atypical multidomain MIF protein. The marine dinoflagellate Lingulodinium polyedra produces a transmembrane protein with an extra-cytoplasmic MIF domain, which localizes to cell-wall-associated membranes and vesicular bodies. This protein is also present in the membranes of extracellular vesicles accumulating at the secretory pores of the cells. Upon exposure to biotic stress, L. polyedra exhibits reduced expression of the MIF gene and reduced abundance of the surface-associated protein. The presence of LpMIF in the membranes of secreted extracellular vesicles evokes the fascinating possibility that LpMIF may participate in intercellular communication and/or interactions between free-living organisms in multispecies planktonic communities.

Efficient, fast and inexpensive bioassay to monitor benthic microalgae toxicity: Application to Ostreopsis species.

Even though HPLC-MS is commonly used to quantify the toxin content of Ostreopsis spp. cells, there is a need to develop easy-to-use toxicological tests to set thresholds during Ostreopsis spp. blooms. The crustacean Artemia has been widely used to evaluate the presence and toxicity of chemicals and biological contaminants and we anticipated that it could also be useful to test Ostreopsis spp. toxicity. Its relevance was first assessed by investigating the variability of the toxic effects among Ostreopsis spp. strains and throughout the dinoflagellate life cycle in combination with chemical analyses of the toxinic content by UHPLC-HRMS. After testing the toxicity of fractions prepared from Ostreopsis spp. cells, the known ova- and paly-toxins were not the only toxic metabolites to Artemia franciscana, indicating that other toxic compounds synthesized by Ostreopsis spp. still remain to be identified. To extend the bioassay to in situ monitoring, the toxicity of the benthic microalgal consortium was tested during a natural bloom of Ostreopsis cf. ovata in the NW Mediterranean Sea. The results highlight the accuracy and sensitivity of the ecotoxicological assay with Artemia franciscana to assess the toxicity of Ostreopsis spp. blooms.

Toxin content of Ostreopsis cf. ovata depends on bloom phases, depth and macroalgal substrate in the NW Mediterranean Sea.

Over the last fifteen years, blooms of the genus Ostreopsis have been reported more frequently and at higher abundances in the Mediterranean area. Ostreopsis cf. ovata is known to produce ovatoxins (OVTXs), structural analogues of palytoxin, which is one of the most potent non-polymeric toxins. However, the production of OVTXs is poorly characterized in situ. The present study focuses on toxin content and profile according to the bloom phase during summer 2017 in Villefranche-sur-Mer, France (NW Mediterranean Sea), depth (from 0.5 to 5 m) and three different macroalgal substrates of this epiphytic dinoflagellate (Padina pavonica, Dictyota spp. and Halopteris scoparia). Ovatoxin quantification of all samples was performed by liquid chromatography coupled to tandem mass spectrometry (LC-MS/MS). The bloom started at the end of June and declined in mid-July, showing the typical seasonal pattern of the NW Mediterranean Sea area. The peak was observed on the 10 July with 1.8 × 106 cells/g FW and 1.7 × 104 cells/L for benthic and planktonic cells, respectively. Total toxin content of cells, collected using artificial substrates, increased during the exponential and stationary growth phases. After reaching a maximum concentration of 9.2 pg/cell on 18 July, toxin concentration decreased and remained stable from 25 July until the end of monitoring. A decreasing trend of the abundance and of the associated total toxin content was noted with depth. Finally, the decreasing order of maximal epiphytic concentration of O. cf. ovata was: Dictyota spp. (8.3 × 105 cells/g FW), H. scoparia (3.1 × 105 cells/g FW) and P. pavonica (1.6 × 105 cells/g FW). Interestingly, the highest OVTX quota was obtained in cells present on Halopteris scoparia, then on Dictyota spp. and Padina pavonica. This suggests that the nature of the macroalgal substrate influences both growth and toxin production of O. cf. ovata and further work will be required to understand the underlying mechanisms (e.g., competition for nutrition, pH or allelopathic interaction). However, the toxin profiles (i.e., the proportion of each ovatoxin analogue) were not affected by any of the studied parameters (bloom phase, depth, macroalgae or artificial substrates).

Spatio-temporal dynamics and biotic substrate preferences of benthic dinoflagellates in the Lesser Antilles, Caribbean sea.

Epibenthic dinoflagellates were monitored monthly over an 18 month period in Guadeloupe and Martinique (Lesser Antilles, Caribbean Sea). These islands are located in the second most affected ciguatera fish poisoning (CFP) region of the world. Guadeloupe presented five times more total epibenthic dinoflagellates and two times less abundant Gambierdiscus spp. compared to Martinique, although the area of frequent CFP outbreaks covers Guadeloupe and not Martinique. Results did not show any clear seasonal variations of benthic dinoflagellates abundances. Temperature and salinity were not driving parameters in the evolution of total benthic dinoflagellate abundances. Preferential associations were found between macrophyte species and epibenthic dinoflagellates. The Phaeophyceae Dictyota spp. hosted the highest abundances of total epibenthic dinoflagellates, composed mainly of Ostrepsis and Prorocentrum genera. The seagrass Halophila stipulacea hosted the highest abundances of Gambierdiscus spp. and Sinophysis spp. whilst the highest abundance of Coolia was determined on Galaxaura spp. The pelagic Sargassum spp. hosted the lowest abundances of benthic dinoflagellates including the genus Gambierdiscus.

Allelopathic interactions between the benthic toxic dinoflagellate Ostreopsis cf. ovata and a co-occurring diatom.

For decades the microphytobenthos assemblage in the coastal Mediterranean Sea has been regularly colonized by the toxic benthic dinoflagellate Ostreopsis cf. ovata. This harmful algal species is a toxin producer and occupies the same ecological niche as various diatoms. Surprisingly, there are only few insights reported on the physiological responses of diatoms to blooms of O. cf. ovata The chemical interactions of O. cf. ovata with the co-occurring diatom Licmophora paradoxa was studied using a bioassay (measuring impact of cell-free culture filtrate) and a co-culture approach (separate by a membrane) to investigate the effects of the exometabolome and its mode of action. Bioassays highlighted a toxic effect of the exometabolome of O. cf. ovata on the diatom photosynthetic activity. However, the co-cultures revealed that these toxic effects do not occur through remote allelopathy. Contact or close interactions between cells of the two species is most likely needed to impair the diatom growth. Ovatoxins are suspected to be the toxic metabolites secreted by O. cf. ovata although the current set of data did not give confirmation of this assumption. Interestingly, the exometabolome of L. paradoxa impaired the growth and the photochemistry of O. cf. ovata in both bioassays and co-cultures. Some biomarkers possibly involved for the effect were identified using a metabolomic approach and may correspond to oxylipins, however a bacterial source of the bioactive metabolites is also considered.

Uptake of dissolved inorganic and organic nitrogen by the benthic toxic dinoflagellate Ostreopsis cf. ovata.

Environmental factors that shape dynamics of benthic toxic blooms are largely unknown. In particular, for the toxic dinoflagellate Ostreopsis cf. ovata, the importance of the availability of nutrients and the contribution of the inorganic and organic pools to growth need to be quantified in marine coastal environments. The present study aimed at characterizing N-uptake of dissolved inorganic and organic sources by O. cf. ovata cells, using the 15N-labelling technique. Experiments were conducted taking into account potential interactions between nutrient uptake systems as well as variations with the diel cycle. Uptake abilities of O. cf. ovata were parameterized for ammonium (NH4+), nitrate (NO3-) and N-urea, from the estimation of kinetic and inhibition parameters. In the range of 0 to 10µmolNL-1, kinetic curves showed a clear preference pattern following the ranking NH4+>NO3->N-urea, where the preferential uptake of NH4+ relative to NO3- was accentuated by an inhibitory effect of NH4+ concentration on NO3- uptake capabilities. Conversely, under high nutrient concentrations, the preference for NH4+ relative to NO3- was largely reduced, probably because of the existence of a low-affinity high capacity inducible NO3- uptake system. Ability to take up nutrients in darkness could not be defined as a competitive advantage for O. cf. ovata. Species competitiveness can also be defined from nutrient uptake kinetic parameters. A strong affinity for NH4+ was observed for O. cf. ovata cells that may partly explain the success of this toxic species during the summer season in the Bay of Villefranche-sur-mer (France).