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Toxin production in marine microalgae was previously shown to be tightly coupled with cellular stoichiometry. The highest values of cellular toxin are in fact mainly associated with a high carbon to nutrient cellular ratio. In particular, the cellular accumulation of C-rich toxins (i.e., with C:N > 6.6) can be stimulated by both N and P deficiency. Dinoflagellates are the main producers of C-rich toxins and may represent a serious threat for human health and the marine ecosystem. As such, the development of a numerical model able to predict how toxin production is stimulated by nutrient supply/deficiency is of primary utility for both scientific and management purposes. In this work we have developed a mechanistic model describing the stoichiometric regulation of C-rich toxins in marine dinoflagellates. To this purpose, a new formulation describing toxin production and fate was embedded in the European Regional Seas Ecosystem Model (ERSEM), here simplified to describe a monospecific batch culture. Toxin production was assumed to be composed by two distinct additive terms; the first is a constant fraction of algal production and is assumed to take place at any physiological conditions. The second term is assumed to be dependent on algal biomass and to be stimulated by internal nutrient deficiency. By using these assumptions, the model reproduced the concentrations and temporal evolution of toxins observed in cultures of Ostreopsis cf. ovata, a benthic/epiphytic dinoflagellate producing C-rich toxins named ovatoxins. The analysis of simulations and their comparison with experimental data provided a conceptual model linking toxin production and nutritional status in this species. The model was also qualitatively validated by using independent literature data, and the results indicate that our formulation can be also used to simulate toxin dynamics in other dinoflagellates. Our model represents an important step towards the simulation and prediction of marine algal toxicity.

Blooms of the benthic toxic dinoflagellate genus Ostreopsis have been recorded more frequently during the last two decades, particularly in warm temperate areas such as the Mediterranean Sea. The proliferation of Ostreopsis species may cause deleterious effects on ecosystems and can impact human health through skin contact or aerosol inhalation. In the eastern Atlantic Ocean, the toxic O. cf. ovata has not yet been reported to the north of Portugal, and the only species present further north was O. cf. siamensis, for which the toxic risk is considered low. During summer blooms of unidentified Ostreopsis species on the French Basque coast (Atlantic) in 2020 and 2021, people suffered from irritations and respiratory disorders, and the number of analyzed cases reached 674 in 2021. In order to investigate the causes, sampling was carried out during summer 2021 to (i) taxonomically identify Ostreopsis species present using a molecular approach, (ii) isolate strains from the bloom and culture them, and (iii) characterize the presence of known toxins which may be involved. For the first time, this study reports the presence of both O. cf. siamensis and O. cf. ovata, for which the French Basque coast is a new upper distribution limit. Furthermore, the presence of ovatoxins a, b, c, and d in the environmental sample and in a cultivated strain in culture confirmed the toxic nature of the bloom and allowed identifying O. cf. ovata as the producer. The present data identify a new health risk in the area and highlight the extended distribution of some harmful dinoflagellates, presumably in relation to climate change.

The marine dinoflagellate genera Coolia Meunier and Ostreopsis Schmidt have been reported in the Western Indian Ocean and include potentially harmful species. However, no comprehensive observations have been reported in Mauritian waters. The primary aim of this study was to isolate, identify and characterize potentially toxic epiphytic Coolia and Ostreopsis species from the coastal waters of Mauritius. Morphological characteristics were examined using light/fluorescence microscopy and scanning electron microscopy. The morphologies of the Coolia and Ostreopsis strains were similar to those of Coolia canariensis and Ostreopsis ovata. The phylogenetic analyses (large subunit ribosomal [LSU] rDNA D1/D2) revealed that the Mauritian strains of Coolia canariensis and Ostreopsis ovata clustered within the clades of these species complexes with other isolates from different areas. This represents the first record of Coolia canariensis in Mauritius, ascribed to C. canariensis phylogroups I and V, with the latter representing a new lineage of this species complex. The findings broaden the current body of knowledge of Coolia canariensis lineages, while the additional information of Ostreopsis ovata further supports the presence of an Indo-Pacific lineage.

Harmful benthic dinoflagellates, usually developing in tropical areas, are expanding to temperate ecosystems facing water warming. Reports on harmful benthic species are particularly scarce in the Southern Mediterranean Sea. For the first time, three thermophilic benthic dinoflagellates (Ostreopsis cf. ovata, Prorocentrum lima and Coolia monotis) were isolated from Bizerte Bay (Tunisia, Mediterranean) and monoclonal cultures established. The ribotyping confirmed the morphological identification of the three species. Maximum growth rates were 0.59 ± 0.08 d−1 for O. cf. ovata, 0.35 ± 0.01 d−1 for C. monotis and 0.33 ± 0.04 d−1 for P. lima. Toxin analyses revealed the presence of ovatoxin-a and ovatoxin-b in O. cf. ovata cells. Okadaic acid and dinophysistoxin-1 were detected in P. lima cultures. For C. monotis, a chromatographic peak at 5.6 min with a mass m/z = 1061.768 was observed, but did not correspond to a mono-sulfated analogue of the yessotoxin. A comparison of the toxicity and growth characteristics of these dinoflagellates, distributed worldwide, is proposed.

The genus of benthic dinoflagellates Ostreopsis is of particular interest because some species negatively impact human health and coastal marine ecosystems. Ostreopsis populations from a remote area, such as the Galapagos Marine Reserve with its unique biodiversity, can provide significant data. Samples of epibionthic dinoflagellates were collected from two islands (Santa Cruz and Santa Fé) in 2017. Species of the genera Gambierdiscus, Amphidinium, Coolia and Ostreopsis were found. Ostreopsis strains were isolated to characterize their morphology, molecular biology and toxicity. Three dif­ferent morphotypes of Ostreopsis based on dorsoventral and width diameters (n=369) were distinguished. The small cell morphotype was dominant in ten samples, with abundances of up to 33405 cells g-1 fresh weight of macroalgae. A total of 16 strains were isolated from field samples with subsequent polymerase chain reaction amplifications of rDNA, 5.8S rDNA and internal transcribed space regions; 13 strains (small cell morphotype) clustered in the O. cf. ovata Atlantic/Indian/Pacific clade; and 3 strains (large cell morphotype) clustered in the Ostreopsis lenticularis genotype from the type locality. The strains proved to be non-toxic. The presence of these genera/species represents a potential threat to marine ecosystems, and it is thus important to consider benthic species in the surveillance of harmful algae blooms in the reserve.

This study aimed to improve the understanding of the nutrient modulation of Ostreopsis cf. ovata toxin content. During the 2018 natural bloom in the NW Mediterranean, the total toxin content (up to ca. 57.6 ± 7.0 pg toxin cell−1) varied markedly. The highest values often coincided with elevated O. cf. ovata cell abundance and with low inorganic nutrient concentrations. The first culture experiment with a strain isolated from that bloom showed that cell toxin content was higher in the stationary than in the exponential phase of the cultures; phosphate- and nitrate-deficient cells exhibited similar cell toxin variability patterns. The second experiment with different conditions of nitrogen concentration and source (nitrate, urea, ammonium, and fertilizer) presented the highest cellular toxin content in the high-nitrogen cultures; among these, urea induced a significantly lower cellular toxin content than the other nutrient sources. Under both high- and low-nitrogen concentrations, cell toxin content was also higher in the stationary than in the exponential phase. The toxin profile of the field and cultured cells included ovatoxin (OVTX) analogues -a to -g and isobaric PLTX (isoPLTX). OVTX-a and -b were dominant while OVTX-f, -g, and isoPLTX contributed less than 1-2%. Overall, the data suggest that although nutrients determine the intensity of the O. cf. ovata bloom, the relationship of major nutrient concentrations, sources and stoichiometry with cellular toxin production is not straightforward.

Few works have been carried out on benthic harmful algal blooms (BHAB) species in the southern Mediterranean and no data are available for the highly dynamic Strait of Gibraltar (western Mediterranean waters). For the first time, Ostreopsis sp. 9, Prorocentrum lima and Coolia monotis were isolated in this key region in terms of exchanges between the Atlantic Ocean and the Mediterranean and subject to intense maritime traffic. Ribotyping confirmed the morphological identification of these three dinoflagellates species. Monoclonal cultures were established and the maximum growth rate and cell yield were measured at a temperature of 24 °C and an irradiance of 90 µmol photons m−2 s−1, for each species: 0.26 ± 0.02 d−1 (8.75 × 103 cell mL−1 after 28 days) for Ostreopsis sp. 9, 0.21 ± 0.01 d−1 (49 × 103 cell mL−1 after 145 days) for P. lima and 0.21 ± 0.01 d−1 (10.02 × 103 cell mL−1 after 28 days) for C. monotis. Only P. lima was toxic with concentrations of okadaic acid and dinophysistoxin-1 measured in optimal growth conditions ranging from 6.4 pg cell−1 to 26.97 pg cell−1 and from 5.19 to 25.27 pg cell−1, respectively. The toxin content of this species varied in function of the growth phase. Temperature influenced the growth and toxin content of P. lima. Results suggest that future warming of Mediterranean coastal waters may lead to higher growth rates and to increases in cellular toxin levels in P. lima. Nitrate and ammonia affected the toxin content of P. lima but no clear trend was noted. In further studies, we have to isolate other BHAB species and strains from Strait of Gibraltar waters to obtain more insight into their diversity and toxicity.

In September 2015, a massive occurrence of the Ostreopsis species was recorded in central Adriatic Kaštela Bay. In order to taxonomically identify the Ostreopsis species responsible for this event and determine their toxin profile, cells collected in seawater and from benthic macroalgae were analyzed. Conservative taxonomic methods (light microscopy and SEM) and molecular methods (PCR-based assay) allowed the identification of the species Ostreopsis cf. ovata associated with Coolia monotis. The abundance of O. cf. ovata reached 2.9 × 104 cells L−1 in seawater, while on macroalgae, it was estimated to be up to 2.67 × 106 cells g−1 of macroalgae fresh weight and 14.4 × 106 cells g−1 of macroalgae dry weight. An indirect sandwich immunoenzymatic assay (ELISA) and liquid chromatography–high-resolution mass spectrometry (LC-HRMS) were used to determine the toxin profile. The ELISA assay revealed the presence of 5.6 pg palytoxin (PLTX) equivalents per O. cf. ovata cell. LC-HRMS was used for further characterization of the toxin profile, which showed that there were 6.3 pg of the sum of ovatoxins (OVTXs) and isobaric PLTX per O. cf. ovata cell, with a prevalence of OVTXs (6.2 pg cell−1), while the isobaric PLTX concentration was very low (0.1 pg cell−1). Among OVTXs, the highest concentration was recorded for OVTX-a (3.6 pg cell−1), followed by OVTX-b (1.3 pg cell−1), OVTX-d (1.1 pg cell−1), and OVTX-c (0.2 pg cell−1).

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.

The genus Ostreopsis includes some species that produce high biomass blooms and/or synthesize toxic compounds that can be transferred through the marine food webs or aerosolized causing ecological, human health and socio-economic impacts. Ostreopsis species are increasing their biogeographic distribution from tropical to more temperate waters and causing recurrent blooms in certain coastal areas, thus constituting an emerging concern worldwide. The proliferation capacity of Ostreopsis is due to a complex and poorly understood combination of multiple factors, and may be a paradigm of chemical ecology reviewed here. A first section summarizes the basic knowledge on the different Ostreopsis species, the toxins they produce and the described foodborne and airborne effects of Ostreopsis toxins on humans. Secondly, direct and indirect interactions between Ostreopsis species and their environment are reviewed. Mucopolysaccharide substances produced by the cells to attach to different substrates appear to be a key element on the chemical ecology and requires further study. However, this research is challenged by technical limitations to conduct ecologically realistic and harmonized studies where organisms can be in direct contact with Ostreopsis cells, their mucus and/or the released extracellular toxic compounds. Understanding the transfer mechanisms of these substances within the food web, potentially affecting humans is critical and requires further study with new analytical tools. Still, the progress in knowledge achieved in the last years, combined with experimental and field studies using cutting edge methods will facilitate to address the open questions on the chemical ecology of Ostreopsis and understand its bloom dynamics now, and under future climate and anthropogenic change scenarios.