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On August 12, 2014, an Anchorage hospital notified the Alaska Section of Epidemiology (SOE) that a middle-aged male resident of Anchorage (patient A) had arrived in the emergency department with possible palytoxin exposure. Patient A complained of a bitter metallic taste, fever, weakness, cough, and muscle pain 7-8 hours after introduction of live zoanthid coral into his home aquarium. Palytoxin, a potent toxin known to produce the reported effects, is contained in zoanthid marine corals.

Benthic dinoflagellates Ostreopsis spp. are known as producers of palytoxin and its analogs, resulting occasionally in human health problems worldwide. Although distribution of Ostreopsis spp. along the Thai coasts has been reported, little is known about their growth characteristics. To discuss the bloom dynamics of Ostreopsis spp. in Thailand, first we tested four kinds of media to optimize growth conditions and then clarified the effects of temperature, salinity and temperature–salinity interaction on the growth of strains of the O. cf. ovata Thailand subclade, O. cf. ovata South China Sea subclade, Ostreopsis sp. 6 and Ostreopsis sp. 7. We showed that the f/2 medium was a suitable medium which gave the highest cell yields for all the strains tested. The strains of the O. cf. ovata Thailand subclade, O. cf. ovata South China Sea subclade and Ostreopsis sp. 6 grew in the temperature range 20–32.5 °C, whereas the strain of Ostreopsis sp. 7 grew in 20–30 °C. The semi-optimal temperature ranges (≧80% of the maximal growth rate) for the former three strains were 22.7–27.4 °C, 27.9–30.8 °C and 23.5–26.4 °C, respectively, whereas that of the latter strain was 23–27.2 °C. The optimal temperature for the O. cf. ovata South China Sea subclade was 30 °C, whereas for the others it was 25 °C. All the Ostreopsis strains tested could grow in a salinity range of 20–40. The semi-optimal salinities for the O. cf. ovata Thailand subclade, O. cf. ovata South China Sea subclade Ostreopsis sp. 6 and Ostreopsis sp. 7 were 28.7–35, 23.8–30.8, 29.8–36 and 28–36, respectively. The optimal salinities for the O. cf. ovata Thailand subclade and O. cf. ovata South China Sea subclade were 30 and 25, respectively, whereas for Ostreopsis sp. 6 and Ostreopsis sp. 7 it was 35. In this study, our results suggested that the optimal and tolerable temperature–salinity conditions differ among the Thai Ostreopsis species/clades/subclades. Tolerances of the O. cf. ovata Thailand subclade, O. cf. ovata South China Sea subclade and Ostreopsis sp. 6 to the high temperature of 32.5 °C may allow these organisms to be distributed in the tropical areas, where the water temperature often reaches >30 °C.

The presence of Ostreopsis cf. ovata on the Mediterranean coast represents a serious concern to human health due to production of toxins–putative palytoxin and ovatoxins (ovatoxin-a, -b, -c, -d, -e, -f and -g). However, purified ovatoxins are not widely available and their toxicities are still unknown. In the present study, we report on HR LC-MS/MS analysis of a French Ostreopsis cf. ovata strain (IFR-OST-0.3 V) collected at Villefranche-sur-Mer (France) during a bloom in 2011. Investigation of this strain of Ostreopsis cf. ovata cultivated in our laboratory by ultra-high performance liquid chromatography coupled to high resolution mass spectrometry (UHPLC-HRMS) confirmed the production of ovatoxins -a to–e and revealed the presence of a new ovatoxin analogue, named ovatoxin-h. Ostreopsis cf. ovata extracts were pre-purified by Sephadex LH-20 to obtain a concentrated fraction of ovatoxins (OVTXs). This method provided a recovery of about 85% of OVTXs and a cleanup efficiency of 93%. Different stationary phases were tested with this fraction of interest to elucidate the structure of the new OVTX congener and to obtain purified ovatoxins. Eight reversed phase sorbents were evaluated for their capacity to separate and purify ovatoxins. Among them Kinetex C18, Kinetex PFP and Uptisphere C18-TF allowed for best separations almost achieving baseline resolution. Kinetex C18 is able to sufficiently separate these toxins, allowing us to identify the toxins present in the extract purified by Sephadex LH-20, and to partly elucidate the structure of the new ovatoxin congener. This toxin possesses one oxygen atom less and two hydrogens more than ovatoxin-a. Investigations using liquid chromatography coupled to high resolution tandem mass spectrometry suggest that the part of the molecule where ovatoxin-h differs from ovatoxin-a is situated between C42 and C49. Uptisphere C18-TF was proposed as a first step preparative chromatography as it is able to separate a higher number of ovatoxins (especially ovatoxin-d and ovatoxin-e) and because it separates ovatoxins from unknown compounds, identified using full scan single quadrupole mass spectrometry. After pre-purification with Sephadex LH-20, purification and separation of individual ovatoxins was attempted using an Uptisphere C18-TF column. During recovery of purified toxins, problems of stability of OVTXs were observed, leading us to investigate experimental conditions responsible for this degradation.

Palytoxin (PlTX) induces a stress response in MCF-7 cells that involves the phosphorylation of HSP 27 at serines 15, 78, and 82 by an as yet undetermined mechanism. We have studied the involvement of major groups of the mitogen-activated protein kinase (MAPK) family in this molecular response and focused our analyses on the ERK1/2, JNK, p38 protein kinase (p38K), and ERK5 pathways. The results show that PlTX induces the activation of JNK and p38 kinase but not ERK1/2 and 5 in MCF-7 cells. Through the use of protein kinase inhibitors, we established that blocking p38K, but not JNK, prevents the phosphorylation of HSP 27 induced by PlTX and that MAPKAPK2 participates in the response induced by the toxin under our experimental conditions. The cell death response induced by PlTX was inhibited by preventing JNK phosphorylation but not by blocking p38K/MAPKAPK2 and HSP 27 phosphorylation. Sucrose density gradient centrifugation revealed that MCF-7 cell extracts contain a heterodisperse population of HSP 27, including oligomers and smaller forms. Treating MCF-7 cells with PlTX caused the dissociation of HSP 27 oligomers, and using inhibitors of the JNK and p38K pathways showed that the dissociation of HSP 27 oligomers induced by PlTX involves a p38K-dependent process. We conclude that the changes induced by PlTX in the HSP 27 stress response protein system proceed through a molecular mechanism involving the activation of the p38 kinase pathway and its substrate, MAPKAK2, leading to dissociation of HSP 27 oligomers and the stabilization of a cellular pool of monomers phosphorylated at serines 15, 78 and 82, which could play a protective role against the death response induced by PlTX.

The hazardous substance from Procambarus clarkii could cause rhabdomyolysis. Mice were administered mashed muscles and gills, intestines, and glands (GIG) obtained from cooked crayfish by gavage . Three of seventy-two mice administered with GIG (with an incidence rate of 4.17 %) appeared to have rhabdomyolysis symptoms with the serum creatine kinase (CK) content increased by more than 5 times, as well as the pathological changes in livers, spleens, kidneys, and skeletal muscles. Furthermore, the cytotoxicity and hemolysis analysis of the palytoxin revealed that the methanol/water extracts from GIG of crayfish were not inhibited by ouabain effectively, which pointed out the hazardous substance causing rhabdomyolysis was not allergen or palytoxin. These research results could be valuable in further comprehensive experiments and lay a foundation for basic rhabdomyolysis research.

The dinoflagellates of the genus Ostreopsis Schmidt are toxic species involved in the occurrence of massive blooms. These dinoflagellates are known to produce palytoxin and ovatoxins, which are considered amongst the most poisonous phycotoxins in the world. In this work, aqueous and methanolic extracts of one Ostreopsis cf. ovata Ionian strain were tested on the human-derived HeLa cell line, and cytotoxicity, cytoskeletal rearrangement, and apoptosis induction were recorded by morphological and molecular analysis. The 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide (MTT) test revealed high toxicity for both extracts, which activity was inhibited by ouabain, which suggests the main involvement of palytoxin-like molecules in the observed toxic effects of this strain. Overall, the effects induced by O. cf. ovata on HeLa cells were dependent on the type of adopted extracts, being both toxic, but with the methanolic extract more toxic than the aqueous one and able to induce cell death by apoptosis. This process was evidenced by both phosphatidylserine exposition and upregulation of caspase-3 gene expression. The different effect of the two extracts on cell death is indicative of their different composition and/or activity. Our findings represent the starting step for the characterization of novel bioactive molecules.

Introduction Palytoxin, produced by unicellular microalgae, is one of the most potent toxins known. Human exposure results most prominently through oral, dermal, and inhalational contact. There is no current data on the prevalence of cases in the United States. Methods This study reviewed human inhalational and dermal exposures to palytoxin via records from the National Poison Data System from 2000 to 2014. Results There were 171 cases reported, the majority occurred at a residence and experienced minor respiratory or dermal symptoms and recovered without any major sequelae or fatalities. Ten patients required admission to an intensive care unit. Conclusion The incidence of palytoxin exposure is low but significant symptoms can occur.

Palytoxin (PLTX) and its analogs are toxic marine polyethers detected in Palythoa corals, Ostreopsis dinoflagellates and Thricodesmium cyanobacteria. The main concern for human health related to these toxins is their accumulation in marine edible organisms and the possible entrance in the food chain up to humans. In tropical areas, human cases of even fatal foodborne poisonings had been ascribed to consumption of PLTXs contaminated seafood. On the other hand, in the Mediterranean Sea, human poisonings ascribed to PLTXs were associated to inhalation of marine aerosol and/or cutaneous exposure to seawater during Ostreopsis blooms and no foodborne poisoning associated with these toxins were documented in this area, so far. Although PLTXs are not regulated, the European Food Safety Authority (EFSA) suggested a maximum limit of 30 µg PLTXs/kg of shellfish meat, recommending the development of suitable, specific and rapid detection methods in seafood. Moreover, given the growing cases of adverse effects during Ostreopsis blooms in the Mediterranean Sea, the development of methods for PLTXs detection also in microalgae is suggested. Thus, two methods for PLTXs quantitation were developed and characterized: a hemolytic assay, based on PLTXs ability to convert the Na+/K+ ATPase into an unspecific cationic channel leading to erythrocytes lysis, and a cell-based immunoenzymatic assay (ELISA), measuring the binding of PLTXs to Na+/K+ ATPase of HaCaT cells by a mouse monoclonal anti-PLTX antibody targeted by an enzyme-conjugated anti-mouse detection antibody. The limits of PLTX detection (LOD) and quantitation (LOQ) by hemolytic assay were 1.4x10-10 M and 3.4x10-10 M, respectively, while those by the cell-based ELISA were 1.2x10-11 M and 2.8x10-11 M, respectively. The two methods were assessed for their suitability to quantify PLTXs in mussels (Mytilus galloprovincialis) and microalgae. The LOQ for PLTX in mussels by the hemolytic assay was 650 µg/kg meat, a value about 20 times higher than the limit suggested by EFSA (30 µg PLTXs/kg meat). On the other hand, the LOQ for PLTX in mussels by the cell-based ELISA was 9.3 µg/kg meat, a value below the safety limit suggested by EFSA. The high sensitivity of the cell-based ELISA, make this method suitable for PLTX detection in monitoring program. The LOQ by the hemolytic assay and the cell-based ELISA in microalgae were 0.03 pg/cell and 0.007 pg/cell, respectively. The second part of this thesis was focalized on the investigation of the sensitivity of different cell models to PLTX on which subsequent studies could be carried out to further characterize PLTX mechanism of action and to develop a new functional detection method for PLTXs. The sensitivity was evaluated by means of cytotoxicity and binding assay. The MTT assay showed Caco-2 cells as the most sensitive to PLTX cytotoxic effect with an EC50 = 3.1x10-11 M (95% C.I. = 5.9x10-12 - 3.4x10-11), while HaCaT cell line was assumed as the most sensitive cell line for PLTX binding, as confirmed by the Bmax and Kd values, equal to 0.043 and 1.4x10-10 M, respectively. The high sensitivity to PLTX demonstrated for Caco-2 cell line by the MTT assay and for HaCaT cell line by the binding assay, was not related to the expression of the Na+/K+ ATPase, suggesting that other factors, such as the expression of specific Na+/K+ ATPase isoforms may play an important role in cells sensitivity to PLTX.

Phycotoxins, which are produced by harmful microalgae and bioaccumulate in the marine food web, are of growing concern for Australia. These harmful algae pose a threat to ecosystem and human health, as well as constraining the progress of aquaculture, one of the fastest growing food sectors in the world. With better monitoring, advanced analytical skills and an increase in microalgal expertise, many phycotoxins have been identified in Australian coastal waters in recent years. The most concerning of these toxins are ciguatoxin, paralytic shellfish toxins, okadaic acid and domoic acid, with palytoxin and karlotoxin increasing in significance. The potential for tetrodotoxin, maitotoxin and palytoxin to contaminate seafood is also of concern, warranting future investigation. The largest and most significant toxic bloom in Tasmania in 2012 resulted in an estimated total economic loss of~AUD$23M, indicating that there is an imperative to improve toxin and organism detection methods, clarify the toxin profiles of species of phytoplankton and carry out both intra‐ and inter‐species toxicity comparisons. Future work also includes the application of rapid, real‐time molecular assays for the detection of harmful species and toxin genes. This information, in conjunction with a better understanding of the life histories and ecology of harmful bloom species, may lead to more appropriate management of environmental, health and economic resources.

© 2017 by the authors; licensee MDPI, Basel, Switzerland. Phycotoxins, which are produced by harmful microalgae and bioaccumulate in the marine food web, are of growing concern for Australia. These harmful algae pose a threat to ecosystem and human health, as well as constraining the progress of aquaculture, one of the fastest growing food sectors in the world. With better monitoring, advanced analytical skills and an increase in microalgal expertise, many phycotoxins have been identified in Australian coastal waters in recent years. The most concerning of these toxins are ciguatoxin, paralytic shellfish toxins, okadaic acid and domoic acid, with palytoxin and karlotoxin increasing in significance. The potential for tetrodotoxin, maitotoxin and palytoxin to contaminate seafood is also of concern, warranting future investigation. The largest and most significant toxic bloom in Tasmania in 2012 resulted in an estimated total economic loss of ∼AUD$23M, indicating that there is an imperative to improve toxin and organism detection methods, clarify the toxin profiles of species of phytoplankton and carry out both intra- and inter-species toxicity comparisons. Future work also includes the application of rapid, real-time molecular assays for the detection of harmful species and toxin genes. This information, in conjunction with a better understanding of the life histories and ecology of harmful bloom species, may lead to more appropriate management of environmental, health and economic resources.

Intense blooms of the toxic dinoflagellate Ostreopsis have been a recurrent phenomenon along several Mediterranean coasts. Blooms have been associated with noxious effects on human health and mortality of marine organisms, due to the production of palytoxin-like compounds. We analyzed the toxin concentrations throughout an O. cf. ovata bloom to highlight their relationships with environmental parameters in the Conero Riviera, northern Adriatic Sea. High temperature and balanced nutrient conditions were the optimal environmental conditions to start and sustain blooms as well as to maximize toxin production. Ostreopsis showed a gradual decrease of toxin content throughout the bloom ascribed to the occurring of the same non-optimal conditions that led to the bloom decline. Moreover, our results suggest that toxin fraction released during bloom could be higher than that released in batch culture. Results from this study pointed out that the first bloom phase is potentially the most dangerous to human health.

Many instrumental techniques are currently employed to identify marine toxins and characterize their structures, with nuclear magnetic resonance (NMR) spectroscopy and mass spectrometry (MS) being the most popular and powerful. Although NMR is the technique of choice to fully elucidate stereostructure of new molecules, its applicability depends on availability of g to mg amounts of pure compound. This is often difficult to achieve in the field of marine toxins, which are usually present at sub-mg levels in complex matrices in the form of a wide array of different congeners. So recently, several MS based approaches have been developed to disclose the presence of toxins and to elucidate their structures based on the interpretation of their fragmentation pattern. This chapter provides with an overview of the different experimental strategies used to discover and elucidate the structure of marine toxins. Practical examples of how some structurally complex toxins - palytoxins, yessotoxins, and spirolides - have been studied until final characterization are included.

Fifty-five strains of Ostreopsis were collected in the Mediterranean Sea and analyzed to characterize their toxin profiles. All the strains were grown in culture under the same experimental conditions and identified by molecular PCR assay based on the ITS-5.8S rDNA. A liquid chromatography-high resolution multiple stage mass spectrometry (LC-HRMSn) approach was used to analyze toxin profiles and to structurally characterize the detected toxins. Despite morphological and molecular characterization being consistent within the species O. cf. ovata, a certain degree of toxin variability was observed. All the strains produced ovatoxins (OVTXs), with the exception of only one strain. Toxin profiles were quite different from both qualitative and quantitative standpoints: 67% of the strains contained OVTX-a to -e, OVTX-g, and isobaric PLTX, in 25% of them only OVTX-a, -d, -e and isobaric PLTX were present, while 4% produced only OVTX-b and -c. None of the strains showed a previously identified profile, featuring OVTX-f as dominant toxin, whereas OVTX-f was a minor component of very few strains. Toxin content was mostly in the range 4-70 pg/cell with higher levels (up to 238 pg/cell) being found in strains from the Ligurian and South Adriatic Sea. Structural insights into OVTX-b, -c, -d, and -e were gained, and the new OVTX-l was detected in 36 strains.

13 páginas, 6 figuras, 7 tablas, The toxin profile and hemolytic activity of a strain of Ostreopsis cf. ovata (UFBA013) isolated from Todos os Santos Bay (northeastern Brazil) were evaluated under different levels of N and P. Phylogenetic analyses based on ITS rDNA region (ITS1-5.8S-ITS2) placed UFBA013 within the Atlantic/Mediterranean/Pacific clade of O. cf. ovata. Growth experiments were conducted in f/2 medium modified by adding N and P (P: 0–36 μM; N: 0–882 μM). The growth kinetics was adequately described by logistic equations. The best growth (highest Gm) was recorded under levels of N/P = 0/18, 129/5 and 441/36, while one of the lowest Gm was obtained under P-depletion. The maximum and specific maximum growth rates (as vm; cells mL−1 d−1 and μm; d−1) were achieved with N limitation (N/P = 441/36) and P-limitation/depletion (753/5.3 and 441/0) and are the highest values reported in the literature, most similar to isolates from Pacific and Mediterranean areas. The control experiment (N/P = 441/18) also yielded similar values to those from some Mediterranean isolates, but higher than formerly reported for Brazilian isolates. In all conditions assayed, no palytoxin (PLTX) was detected. The ovatoxins (OVTXs) a, b, c, d and e did not show significant differences in cell quota between exponential and stationary phases. A significant relationship was detected between OVTXs concentration and hemolytic activity, This study was supported by FAPESB - RED0006/2012 and CNPq (Productivity Scholarship in Research # 07368/2015-7) granted to J.M.C. Nunes; CNPq (research grant # 471303/2013-5) to M. Menezes; CAPES (PhD scholarship # 9947-14-14-0/2014) and FAPESB (PhD scholarship Bol. # 10685/2013) to M.C.Q. Mendes

Benthic dinoflagellates of the genus Ostreopsis are found all over the world in temperate, subtropical, and tropical coastal regions. Our recent studies revealed that a putative “cryptic” species of Ostreopsis ovata is present widely along Japanese coasts. This organism, Ostreopsis sp. 1, possesses palytoxin analogs and thus its toxic blooms may be responsible for potential toxification of marine organisms. To evaluate the bloom dynamics of Ostreopsis sp. 1, the present study examined the growth responses of Ostreopsis sp. 1 strain s0716 to various light intensities (photon flux densities: μmol photons m −2 s −1 ) using a newly devised photoirradiation-culture system. This novel system has white light-emitting diodes (LEDs) capable of more closely simulating the wavelength spectrum of light entering the oceanic water column than do fluorescent tubes and halogen lamps. In this system, the light intensity of the white LEDs was reduced through two polarizing filters by varying the rotation angles of the filters. Thereby, the new system was capable of culturing microalgae under well-controlled light intensity conditions. Ostreopsis sp. 1 grew proportionally when light intensity was increased from 49.5 to 199 μmol photons m −2 s −1 , but its growth appeared to be inhibited slightly at ≥263 μmol photons m −2 s −1 . The relationship between observed growth rates and light intensity was calculated at R > 0.99 ( P Ostreopsis sp. 1 and the organism's growth potential as follows: (1) the threshold light intensity for growth: 29.8 μmol photons m −2 s −1 ; (2) the optimum light intensity ( L m ) giving the maximum growth rate ( μ max = 0.659 divisions day −1 ): 196 μmol photons m −2 s −1 ; (3) the optimum light intensity range ( L opt ) giving ≥95% μ max : 130–330 μmol photons m −2 s −1 ; (4) the semi-optimum range ( L sopt ) giving ≥80% μ max : 90 to over 460 μmol photons m −2 s −1 . The L sopt represents 4.5–23% ambient light intensity present in surface waters off of a temperate region of the Japanese coast, Tosa Bay; putatively, this semi-optimum range of light intensity appears at depth of 12.9–27.8 m. Considering these issues, our data indicate that Ostreopsis sp. 1 in coastal environments may form blooms at ca. ∼28 m depth in regions along Japanese coasts.

Palytoxin (PTX), a marine toxin, represents an increasing hazard for human health. Despite its high toxicity for biological systems, the mechanisms triggered by PTX, are not well understood. The high affinity of PTX for erythrocyte Na(+)/K(+)-ATPase pump is largely known, and it indicates PTX as a sensitive tool to characterize the signal transducer role for Na(+)/K(+)-ATPase pump. Previously, it has been reported that in red blood cells (RBC), probably via a signal transduction generated by the formation of a PTX-Na(+)/K(+)-ATPase complex, PTX alters band 3 functions and glucose metabolism. The present study addresses the question of which other signaling pathways are regulated by Na(+)/K(+)-ATPase in RBC. Here it has been evidenced that PTX following its interaction with Na(+)/K(+)-ATPase pump, alters RBC morphology and this event is correlated to decreases by 30% in nitrites and nitrates levels, known as markers of plasma membrane eNOS activity. Orthovanadate (OV), an antagonist of PTX binding to Na(+)/K(+)-ATPase pump, was able to reverse the effects elicited by PTX. Finally, current investigation firstly suggests that Na(+)/K(+)-ATPase pump, following its interaction with PTX, triggers a signal transduction involved in NO metabolism regulation.

The toxic benthic dinoflagellate genus Ostreopsis has been connected to the production of palytoxin and its analogs in many tropical and temperate areas. Although the type species, O. siamensis, was originally described from the Gulf of Thailand in 1901, little is known about the species composition and distribution of the genus Ostreopsis in Thailand. In this study, a total of 64 Ostreopsis strains isolated from the Andaman Sea as well as the Gulf of Thailand were investigated by analyzing the nucleotide sequences of the LSU rDNA D1/D2, D8/D10 and ITS-5.8S rDNA regions. Phylogenetic analyses (BI and ML) resulted in some of the strains being assigned to previously described clades, O. cf. ovata and Ostreopsis sp. 6, and revealed the existence of a novel clade named Ostreopsis sp. 7, which exhibited large genetic distances from the other clades. Among O. cf. ovata, several strains from Thailand were formed into a new subclade, the Thailand subclade, whereas a few strains belonged to the South China Sea subclade. Morphometric characteristics such as the cell sizes of the two O. cf. ovata subclades and those of Ostreopsis sp. 7 were not significantly different from each other (p > 0.05). Their characteristics were similar but slightly different from those of O. ovata and were significantly different from those of Ostreopsis sp. 6 (p < 0.05). Toxicities of Ostreopsis from Thailand were evaluated using mouse bioassay. Strains of Ostreopsis sp. 6 and Ostreopsis sp. 7 tested were highly toxic, while the two subclades of O. cf. ovata strains seemed to be nontoxic. This study suggests that toxic Ostreopsis sp. 7 is distributed in the Andaman Sea, whereas the two subclades of O. cf. ovata and toxic Ostreopsis sp. 6 are distributed in the Gulf of Thailand.

In the last decade Ostreopsis cf. ovata blooms have been among the most intense along the entire Mediterranean coast, leading to ecological and human health problems, that are associated with the toxins (palytoxin-like compounds) produced by these algal cells. These compounds are secondary metabolites, whose rates of synthesis depend on the metabolism of their precursors. In general, growth dynamics and toxicity of dinoflagellates reflect the physiological status of the organism. The aim of the present study was to investigate the cellular production of the main biochemical compounds likely involved in the growth and toxicity dynamics of O. cf. ovata during exponential to the late stationary phase in batch cultures of an Adriatic strain. Removal of major nutrients from the medium was monitored along with concentration, biovolume and production of the main cellular components (e.g. polysaccharides, proteins, lipids and toxins). Nutrient uptake, as well as toxin production rates were calculated in the different growth periods. Nutrients (N and P) were completely depleted when cells entered stationary phase and the greatest net toxin production rate (RTOX) occurred during the first days of growth. The various palytoxins reported a relative abundance quite stable during the different growth phases, while the total toxin cellular amount increased along the growth curve. Total and extracellular released polysaccharides, as well as the lipid content increased greatly during the stationary phase, while proteins were mainly produced by cells during the exponential phase. The continuous release of polysaccharides could facilitate cell aggregation and the formation of the benthic community during algal blooms. The trend of production of the main cellular compounds in O. cf. ovata and the growth dynamics of this species lead us to hypothesize that the fast growth of this dinoflagellate, associated with the rapid use of environmental resources (nutrients, and phosphates in particular), may be an ecological/ adaptive strategy which could favor this organism in competition with other species.

International audience; 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 OPEN ACCESS Mar. Drugs 2014, 12 2852 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.

Palytoxin ranks among the most potent marine biotoxins. Its lethality was well known to native Hawaiians that used to smear a "moss" containing the toxin on their spears to cause instant death to their victims. Human intoxications due to exposure to palytoxin and to its many congeners have been reported worldwide. Currently, palytoxins constitute the main threat to public health across the Mediterranean Sea. In the present work we report on the isolation and stereostructural determination of a new palytoxin analogue from a Hawaiian Palythoa tuberculosa sample. This new toxin is a stereoisomer of 42-hydroxypalytoxin isolated from Palythoa toxica. The whole absolute configuration of this latter toxin is also reported in the paper. Interestingly, the two 42-hydroxypalytoxins do not share the same biological activity. The stereoisomer from P. tuberculosa showed cytotoxicity toward skin HaCaT keratinocytes approximately 1 order of magnitude lower than that of 42-hydroxypalytoxin from P. toxica and about 2 orders of magnitude lower than that of palytoxin itself. This finding holds the prospect of interesting structure-activity relationship evaluations in the future.

Phycotoxins, which are produced by harmful microalgae and bioaccumulate in the marine food web, are of growing concern for Australia. These harmful algae pose a threat to ecosystem and human health, as well as constraining the progress of aquaculture, one of the fastest growing food sectors in the world. With better monitoring, advanced analytical skills and an increase in microalgal expertise, many phycotoxins have been identified in Australian coastal waters in recent years. The most concerning of these toxins are ciguatoxin, paralytic shellfish toxins, okadaic acid and domoic acid, with palytoxin and karlotoxin increasing in significance. The potential for tetrodotoxin, maitotoxin and palytoxin to contaminate seafood is also of concern, warranting future investigation. The largest and most significant toxic bloom in Tasmania in 2012 resulted in an estimated total economic loss of~AUD$23M, indicating that there is an imperative to improve toxin and organism detection methods, clarify the toxin profiles of species of phytoplankton and carry out both intra- and inter-species toxicity comparisons. Future work also includes the application of rapid, real-time molecular assays for the detection of harmful species and toxin genes. This information, in conjunction with a better understanding of the life histories and ecology of harmful bloom species, may lead to more appropriate management of environmental, health and economic resources.

In September 2015, a massive occurrence of the Ostreopsis species was recorded in central Adriatic Ka&scaron, tela 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 &times, 104 cells L&minus, 1 in seawater, while on macroalgae, it was estimated to be up to 2.67 &times, 106 cells g&minus, 1 of macroalgae fresh weight and 14.4 &times, 1 of macroalgae dry weight. An indirect sandwich immunoenzymatic assay (ELISA) and liquid chromatography&ndash, 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&minus, 1), while the isobaric PLTX concentration was very low (0.1 pg cell&minus, 1). Among OVTXs, the highest concentration was recorded for OVTX-a (3.6 pg cell&minus, 1), followed by OVTX-b (1.3 pg cell&minus, 1), OVTX-d (1.1 pg cell&minus, 1), and OVTX-c (0.2 pg cell&minus, 1).

Palytoxin (PLTX) is a highly toxic hydrophilic polyether detected in several edible marine organisms from intra-tropical areas, where seafood poisoning were reported. Symptoms usually start with gastro-intestinal malaise, often accompanied by myalgia, muscular cramps, dyspnea and, sometimes, arrhythmias. Monitoring programs in the Mediterranean Sea have detected PLTX-like molecules in edible mollusks and echinoderms. Despite the potential exposure of the human population and its high toxic potential, the toxicological profile of the molecule is still an issue. Thus, the effects of repeated oral administration of PLTX in mice were investigated. Seven days of PLTX administration caused lethality and toxic effects at doses ≥30 μg/kg/day. A NOAEL was estimated equal to 3 μg/kg/day, indicating a quite steep dose–response curve. This value, due to the limited number of animal tested, is provisional, although represents a sound basis for further testing. Macroscopic alterations at gastrointestinal level (gastric ulcers and intestinal fluid accumulation) were observed in mice dead during the treatment period. Histological analysis highlighted severe inflammation, locally associated with necrosis, at pulmonary level, as well as hyper-eosinophilia and fiber separation in myocardium. A cardiac damage was supported by the in vitro effect of the toxin on cardiomyocytes, indicating a severe and irreversible impairment of their electrical properties: electrophysiological recordings detected a progressive cell depolarization, arrest of action potentials and beating.

Dinoflagellates are prolific producers of polyketide secondary metabolites. Dinoflagellate polyketide synthases (PKSs) have sequence similarity to Type I PKSs, megasynthases that encode all catalytic domains on a single polypeptide. However, in dinoflagellate PKSs identified to date, each catalytic domain resides on a separate transcript, suggesting multiprotein complexes similar to Type II PKSs. Here, we provide evidence through coimmunoprecipitation that single-domain ketosynthase and ketoreductase proteins interact, suggesting a predicted multiprotein complex. In Karenia brevis (C.C. Davis) Gert Hansen & O. Moestrup, previously observed chloroplast localization of PKSs suggested that brevetoxin biosynthesis may take place in the chloroplast. Here, we report that PKSs are present in both cytosol and chloroplast. Furthermore, brevetoxin is not present in isolated chloroplasts, raising the question of what chloroplast-localized PKS enzymes might be doing. Antibodies to K. brevis PKSs recognize cytosolic and chloroplast proteins in Ostreopsis cf. ovata Fukuyo, and Coolia monotis Meunier, which produce different suites of polyketide toxins, suggesting that these PKSs may share common pathways. Since PKSs are closely related to fatty acid synthases (FAS), we sought to determine if fatty acid biosynthesis colocalizes with either chloroplast or cytosolic PKSs. [(3) H]acetate labeling showed fatty acids are synthesized in the cytosol, with little incorporation in chloroplasts, consistent with a Type I FAS system. However, although 29 sequences in a K. brevis expressed sequence tag database have similarity (BLASTx e-value

Here we summarize the current knowledge on the transfer and accumulation of harmful algal bloom (HAB)-related toxins in cephalopods (octopods, cuttlefishes and squids). These mollusks have been reported to accumulate several HAB-toxins, namely domoic acid (DA, and its isomers), saxitoxin (and its derivatives) and palytoxin (and palytoxin-like compounds) and, therefore, act as HAB-toxin vectors in marine food webs. Coastal octopods and cuttlefishes store considerably high levels of DA (amnesic shellfish toxin) in several tissues, but mainly in the digestive gland (DG)—the primary site of digestive absorption and intracellular digestion. Studies on the sub-cellular partitioning of DA in the soluble and insoluble fractions showed that nearly all DA (92.6%) is found in the cytosol. This favors the trophic transfer of the toxins since cytosolic substances can be absorbed by predators with greater efficiency. The available information on the accumulation and tissue distribution of DA in squids (e.g., in stranded Humboldt squids, Dosidicus gigas) is scarcer than in other cephalopod groups. Regarding paralytic shellfish toxins (PSTs), these organisms accumulate them at the greatest extent in DG >> kidneys > stomach > branchial hearts > posterior salivary glands > gills. Palytoxins are among the most toxic molecules identified and stranded octopods revealed high contamination levels, with ovatoxin (a palytoxin analogue) reaching 971 μg kg−1 and palytoxin reaching 115 μg kg−1 (the regulatory limit for PlTXs is 30 μg kg−1 in shellfish). Although the impacts of HAB-toxins in cephalopod physiology are not as well understood as in fish species, similar effects are expected since they possess a complex nervous system and highly developed brain comparable to that of the vertebrates. Compared to bivalves, cephalopods represent a lower risk of shellfish poisoning in humans, since they are usually consumed eviscerated, with exception of traditional dishes from the Mediterranean area.

Since 1953, a total of 27 human poisoning cases caused by the consumption of blue humphead parrotfish, Scarus ovifrons, have been reported in Japan. Characteristic symptoms are severe muscle pain associated with rhabdomyolysis. Although it is believed that palytoxin, which is one of the most potent non-protein marine biotoxins, is the most likely causative toxin in blue humphead parrotfish poisoning, palytoxin has not been proven conclusively as the causative toxin because of lack of a reliable and sensitive analytical method for palytoxin. In 2011, human poisoning cases caused by the consumption of blue humphead parrotfish occurred in Miyazaki and Tokyo. In our present study, an LC-MS/MS method for palytoxin and its analogues in the blue humphead parrotfish samples causing the human poisoning cases in 2011 was developed and the samples were analysed by using the newly developed LC-MS/MS method. Palytoxin and its analogues were not detected in the samples from the food poisoning cases. The LC-MS/MS findings therefore do not support the recently accepted hypothesis that palytoxin is the causative agent in blue humphead parrotfish poisoning in Japan.

RATIONALE The ovatoxins are palytoxin analogs of a dinoflagellate origin implicated in human intoxication. The structures of ovatoxin-a, ovatoxin-d, and ovatoxin-e produced by the IK2 strain of Ostreopsis ovata collected in Japan were elucidated using liquid chromatography/quadrupole time-of-flight mass spectrometry (LC/QTOFMS). The novel structures and a new insight into the spectral data are presented. METHODS The structural elucidations were carried out by complementary use of positive and negative ion LC/QTOFMS. Ostreocin-D (C127H219N3O53), another palytoxin congener previously elucidated by negative fast-atom bombardment collision-induced tandem mass spectrometry (FAB CID MS/MS), was used as a reference. RESULTS Positive ion spectra allowed deduction of hydroxyl positions based on the conjugated polyene structures produced, while the negative ion spectra allowed assignments of cleavage sites of C–C bonds. The analysis could be performed using a small sample without extensive purification. CONCLUSIONS Ovatoxin-a IK2 (C129H223N3O52), ovatoxin-d IK2 (C129H223N3O53), and ovatoxin-e IK2 (C129H223N3O53) were tentatively assigned to 42-hydroxy-17,44,70-trideoxypalytoxin, 42-hydroxy-17,70-dideoxypalytoxin and 42,82-dihydroxy-17,44,70-trideoxypalytoxin, respectively. The wide applicability of the method was suggested. Copyright © 2013 John Wiley & Sons, Ltd.

Palytoxin is one of the most toxic marine toxins known. Distributed worldwide, it poses a potential human health risk linked to the consumption of contaminated seafood. Despite its high parenteral toxicity, the lethal oral dose of palytoxin is several times higher than the intraperitoneal lethal dose. In the present study, we investigated the passage of palytoxin through the human intestinal barrier by employing a well-characterized and accepted in vitro model of intestinal permeability that uses differentiated Caco-2 cell monolayers. Trans-epithelial electric resistance measurements showed that palytoxin disrupts the integrity of Caco-2 monolayers at concentrations > 0.135 nm. However, confocal microscopy imaging showed that the tight-junction protein occludin was not affected by palytoxin in the nanomolar range. This finding was supported by transmission electron microscopy imaging, where tight-junctions appeared to be unaffected by palytoxin treatment. In addition, the nuclear envelope does not appear to be altered by high concentrations of palytoxin. However, palytoxin-treated cells showed electron-dense and damaged mitochondria. Toxin exposure also induced the disappearance of the differentiated Caco-2 microvilli and organelles, as well as chromatin de-condensation. Permeability assays showed that palytoxin could not significantly pass the Caco-2 monolayer, despite the lack of epithelium integrity, suggesting that palytoxins would be poorly transported to blood, which may explain its lower oral toxicity. These data can help to achieve a better understanding of palytoxin poisoning. However, more studies regarding its repeated administration and chronic effects are needed.

Palytoxin (PTX) is considered a severe marine toxin. Although rare, reports of human exposure from consumption of PTX have described significant morbidity and mortality. PTX is the suspected agent in Haff disease, in which rhabdomyolysis occurs within 24 h of eating contaminated fish such as buffalo fish. PTX is primarily present in soft corals or in dinoflagellates, and it can contaminate crustaceans and other fish as it bioaccumulates up the food chain. Only 23 cases have been reported in the USA, including two recent cases in New York City. Reports of inhalational exposure to PTX are uncommon.We describe a case series of six patients, including four adults and two children, with inhalational exposure to PTX aerosolized from Palythoa corals. Their symptoms included some degree of respiratory involvement, myalgias, paresthesias, low-grade fevers, and gastrointestinal symptoms. Fortunately, there were no serious outcomes and all patients survived without sequelae.Although rare, exposure to palytoxin is not restricted to people visiting marine environments because of Palythoa coral in some home aquariums. Routes of exposure go beyond consumption of fish that feed on the coral and include dermal as well as inhalational exposure. Palytoxin exposure should be considered in the differential diagnosis of patients who own or work with fish tanks and present with symptoms that include respiratory complaints, myalgias, neuromuscular dysfunction, hemolysis, and cardiac toxicity. There is no known antidotal therapy and treatment should focus on meticulous supportive care.

Ciguatera fish poisoning (CFP), a disease caused by consuming fish that have accumulated ciguatoxins (CTXs) in their tissue, is regarded as the most prevalent form of intoxication in French Polynesia. Recently, the Australes, one of the least affected archipelago until the early 1980s, has shown a dramatic increase in its incidence rates in 2009 with unusual CFP cases. In the present work, potential health hazards associated with the proliferation of various marine phytoplankton species and the consumption of fish and marine invertebrates highly popular among local population were assessed in three Australes islands: Raivavae, Rurutu and Rapa. Extracts from the marine dinoflagellates Gambierdiscus, Ostreospis and mat-forming cyanobacteria as well as fish, giant clams and sea urchin samples were examined for the presence of CTXs and palytoxin (PLTX) by using the neuroblastoma cell-based assay (CBA-N2a). Cytotoxic responses observed with both standards (Pacific CTX-3C and PLTX) and targeted marine products indicate that CBA-N2a is a robust screening tool, with high sensitivity and good repeatability and reproducibility. In Rurutu and Raivavae islands, our main findings concern the presence of CTX-like compounds in giant clams and sea urchins, suggesting a second bio-accumulation route for CFP toxins in the ciguatera food chain. In Rapa, the potential CFP risk from Gambierdiscus bloom and fish was confirmed for the first time, with levels of CTXs found above the consumer advisory level of 0.01 ng Pacific CTX-1B g(-1) of flesh in three fish samples. However, despite the presence of trace level of PLTX in Ostreopsis natural assemblages of Rapa, no sign of PLTX accumulation is yet observed in tested fish samples. Because this multi-toxinic context is likely to emerge in most French Polynesian islands, CBA-N2a shows great potential for future applications in the algal- and toxin-based field monitoring programmes currently on hand locally.

Ostreopsis ovata is a benthic dinoflagellate that produces palytoxin and ovatoxins. Blooms of O. ovata causing human health problems and mortality of benthic fauna have been reported from many trop ...

The dinoflagellate genus Ostreopsis is known as a potential producer of palytoxin analogues. Recently a surprisingly diverse and widespread distribution of Ostreopsis along coastal waters in Japan was reported in our previous paper. In the present study, detailed toxin profiles of palytoxin analogues in several Japanese Ostreopsis strains were clarified for the first time by liquid chromatography–mass spectrometry (LC–MS). An Italian Ostreopsis strain (KAC 85) was also analyzed by LC–MS as a reference material of Ostreopsis and the presence of ovatoxin-a, -b, -c, -d, and -e was re-confirmed. The most dominant palytoxin analogue in Japanese Ostreopsis was a novel palytoxin analogue, which was identified as an isomer of ovatoxin-a. Novel isomers of ovatoxin-b, -d, and -e were also found in Japanese Ostreopsis. The isomers of ovatoxin-a, -b, -d, and -e detected in Japanese Ostreopsis were tentatively named ovatoxin-a AC, -b AC, -d AC and -e AC. LC–MS analysis suggests that ovatoxin-a AC is dideoxy-42-hydroxy-44-deoxy-palytoxin. The unidentified deoxy-positions can be assigned both between C16 and C20 and between C53 and C73. Toxin profiles of Ostreopsis collected in the middle to southern coastal area in Japan were similar irrespective of clades classified by the phylogenetic analysis reported in our previous study. The toxin contents of some strains collected in Japan were comparable to that obtained in a strain (KAC 85) collected in the Italian coast where human health problems occurred, suggesting a potential risk of Japanese Ostreopsis to human health.

Purpose To report a series of patients who developed corneal toxicity after exposure to aquarium coral palytoxin. Design Multicenter retrospective case series. Methods Retrospective review. Results Seven patients presented with corneal findings ranging from superficial punctate epitheliopathy to bilateral corneal melt with subsequent perforation. Among those with mild corneal findings, resolution was achieved with topical steroids and lubrication, whereas some patients who developed progressive corneal melt required therapeutic penetrating keratoplasty. The history in all patients revealed exposure to aquarium zoanthid corals shortly before disease onset. A review of the literature revealed that there are few prior reports of coral-associated corneal toxicity and that some species of coral secrete a substance known as palytoxin, a potent vasoconstrictor that inhibits the membranous sodium-potassium ATPase pump across cell types and can cause rapid death if inhaled or ingested. Conclusions This is the largest case series to date demonstrating patients with aquarium coral palytoxin–associated corneal toxicity, and is the first to provide details of related histopathologic findings. Similar to other forms of toxic keratoconjunctivitis, a detailed history and careful clinical assessment are required, as well as timely removal of the offending agent from the patients' ocular milieu and environment. Mild ocular surface and corneal disease may be treated effectively with aggressive topical steroid therapy and lubrication. Given the potential severity of ocular as well as systemic adverse effects, there should be increased awareness of this entity among eye care professionals, aquarium enthusiasts, and the general public.

Palytoxin (PLTX) is one of the most harmful marine toxins known so far. Although the ingestion of contaminated seafood is the most dangerous exposure route for humans, cutaneous and inhalational exposures are far more frequent, and can cause strong inflammatory reactions. However, little is known about the inflammatory events that follow the cutaneous exposure to the toxin. In this study, we investigated (1) the effects of both short (2 h) and long (24 h) term exposures of HaCaT keratinocytes to a sub-cytotoxic PLTX concentration on pro-inflammatory mediator gene expression and release and (2) the effect of PLTX-conditioned HaCaT cell media on undifferentiated (monocytes) and differentiated (macrophages; immature dendritic cells, iDCs; mature dendritic cells, mDCs) THP-1 cells. At 10-11 M, PLTX induced interleukin (IL)-6 and IL-8 release from HaCaT keratinocytes after 24 h of continuous exposure to the toxin, as well as after 23 h in toxin-free medium preceded by 1 h exposure to PLTX. Under the same experimental conditions, release of the inflammatory mediators prostaglandin-E2 and histamine was also found after both short and long exposures to the toxin. The conditioned media collected from HaCaT cells treated with PLTX increased the migration of the differentiated and undifferentiated THP-1 cells (potency rank order: monocytes ≥ iDCs > mDCs > macrophages) but did not induce cell differentiation. These results indicate that keratinocytes can be actively involved in the recruitment of inflammatory cells in response to cutaneous contact with PLTX. The lack of a significant effect on monocyte differentiation towards mature immune cells suggests that PLTX is endowed with irritant rather than sensitizing properties.

The new benthic toxic dinoflagellate Ostreopsis fattorussoi sp. nov. is described from eastern Mediterranean Sea Lebanon and Cyprus coasts, supported by morphological and molecular data. The plate formula, Po, 3’, 7’’, 6c, 7s, 5’’’, 2’’’’, is typical for the Ostreopsis genus. It differs from all other Ostreopsis species in that i) the curved suture between plates 1’ and 3’ makes them approximately hexagonal, ii) the 1’ plate lies in the left-half of the epitheca and is obliquely orientated leading to a characteristic shape of plate 6’’. The round thecal pores are bigger than the other two Mediterranean species (O. cf. ovata and O. cf. siamensis). O. fattorussoi is among the smallest species of the genus (DV: 60.07±5.63 μm, AP: 25.66±2.97 μm, W: 39.81±5.05 μm) along with O. ovata. Phylogenetic analyses based on the LSU and ITS rDNA shows that O. fattorussoi belongs to the Atlantic/Mediterranean Ostreopsis spp. clade separated from the other Ostreopsis species. O. fattorussoi produces OVTX-a and structural isomers OVTX-d and –e, O. cf. ovata is the only other species of this genus known to produce these toxins. The Lebanese O. fattorussoi did not produce the new palytoxin-like compounds (ovatoxin-i, ovatoxin-j1, ovatoxin-j2, and ovatoxin-k) that were previously found in O. fattorussoi from Cyprus. The toxin content was in the range of 0.28-0.94 pg. cell−1. In Lebanon coast, O. fattorussoi was recorded throughout the year 2015 (temperature range 18-31.5 °C), with peaks in June and August. This article is protected by copyright. All rights reserved.

Palytoxin is one of the most potent non protein marine toxins known and it is suspected to cause severe, and sometimes even lethal intoxications in humans. Its origin has long been controversial and different potential production sources have been proposed. Nowadays, dinoflagellates belonging to the genus Ostreopsis are the most accredited producers of palytoxins. Since the isolation of palytoxin in 1971, a number of palytoxin congeners have been isolated so far from different biological sources. In this chapter the main aspects of chemistry of palytoxins are reported.

The genus Ostreopsis includes species largely distributed from tropical to temperate marine areas worldwide. Among the nine species of the genus, O. siamensis, O. mascarenensis, O. lenticularis and O. cf. ovata can produce toxins of the palytoxin group. In the last decade Ostreopsis cf. ovata and O. cf. siamensis originated intense blooms in all the rocky Mediterranean Sea coastal areas, typically during summer-late summer. The correct identification of Ostreopsis species in field samples is often problematic as Ostreopsis species are morphologically plastic and hardly discriminable under light microscopy and, therefore, molecular analyses are required. Ostreopsis blooms are often associated with noxious effects on health of both humans and benthic marine organisms mainly carried by aerosol and direct contact with seawater. Environmental factors have been shown to affect toxin content of Ostreopsis which generally produces more toxins per cell when growing under suboptimal conditions. O. cf. ovata is able to produce both temporary and resting cysts. In particular, the resting cysts are able to germinate in laboratory conditions for as long as 5 months after their formation at 25°C, but not at 21°C; the presence of a temperature threshold affecting cyst germination in the laboratory suggests that temperature represents a key factor for Ostreopsis cf. ovata bloom onset in natural environments as well. Several studies conducted to assess the role of abiotic factors (mainly hydrodynamics, water temperature and nutrients) on the bloom dynamics, revealed that the synergic effects of hydrodynamics, temperature and N:P ratios would lead the Ostreopsis blooms in temperate areas. Ostreopsis abundances showed a significant decrease with depth, likely related to light availability, although there are conflicting data about the relationship between light intensity and Ostreopsis growth in experimental conditions. The relationship between Ostreopsis blooms and salinity is not completely clear, complicated by the influence of high nutrient levels often associated to low salinity waters. Finally, Ostreopsis colonize a variety of substrata, although living substrata seems to allow lower concentration of epibionts than any other substrate, probably due to the production of some allelopathic compounds.