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oxic algae and their toxins represent an emerging public health issue, particularly due to global warming. The toxicological mechanisms of neurotoxic phycotoxins and their human health effects have been widely described (paralytic, neurological, amnesic, and ciguateric toxins). Food poisoning by bivalve shellfish (mussels, oysters...) can cause serious or even fatal neurological disorders, due to the accumulation in their flesh of natural toxins that develop in the water that surrounds them. However, retrospective study of human shellfish poisoning highlighted the difficulties in identifying cases related to neurotoxic phycotoxins by gathering clinical and biological diagnostic confirmations on the one hand, as well as environmental contaminations on the other. To improve the collection of shellfish poisoning data, the French Agency for Food, Environmental and Occupational Health and Safety (ANSES), French Poison Control Centers (PCCs), the French Research Institute for Exploitation of the Sea (Ifremer) and the French Defense Procurement Agency (DGA)have developed a prospective surveillance of neurotoxins from algal origin that combines monitoring of poisoning cases (clinical monitoring), of harmful algae and their toxins in shellfish production areas (environmental monitoring), as well as of toxins levels in seafood leftovers and in biological samples from poisoned subjects. This original clinical and environmental monitoring will allow us to have a more complete view of phycotoxin human exposures that are underreported, and to implement measures to protect consumers.

The planktonic diatom genus Pseudo-nitzschia contains several genetically closely related species that can produce domoic acid, a potent neurotoxin known to cause amnesic shellfish poisoning (ASP). An early identification and an adequate monitoring of the potential toxic Pseudo-nitzschia spp. are necessary. However, effective monitoring programs are time consuming due, in some cases, to the cell morphology similarities among species, determined with light microscopy, that can result in insufficient data to give a definitive species and toxins attribution. In this paper, Whole-Cell Fluorescent In Situ Hybridization (WC-FISH) has been evaluated as a powerful tool to detect and enumerate harmful cryptic and/or pseudo-cryptic Pseudo-nitzschia spp. collected in the Gulf of Naples. Fluorescently labelled probes directed against the ribosomal RNA (rRNA) of the 28S large subunit (LSU) were used. In particular, five probes detecting four cryptic species of Pseudo-nitzschia delicatissima complex and one specific for Pseudo-nitzschia multistriata gave good results for the molecular identification of potentially toxic target species in natural samples. Finally, we can state that the WC-FISH method, to identify Pseudo-nitzschia species, is faster and more cost-effective if compared with other rDNA-based methods.

Domoic acid (DA) is a marine neurotoxin produced as a defence compound by diatom Pseudo-nitzschia. Although its toxicity is well known in marine mammals and fish, data on DA cyto/genotoxicity in human non-target cells is still limited. Hence, we aimed to study the effect of DA (0.001 to 10µg/mL) on cell viability and proliferation kinetics of human hepatocellular carcinoma (HepG2) cells as well as DNA damage induction after 4, 24 and 72h of exposure. The results revealed that DA up to 10µg/mL did not elicit significant changes in HepG2 cell viability, proliferation and cell cycle at applied conditions. DA did not generate DNA double-strand breaks, while it exhibited significant dose- and time-dependent increase of DNA damage in the form of either DNA single-strand breaks or alkali labile sites. Additionally, increased malondialdehyde level after DA treatment indicated oxidative damage to lipids. Altogether, the results showed that neurotoxin DA induced only minor adverse genotoxic effects in non-target HepG2 cells that most probably occurred resulting from the oxidative stress. However, additional research is needed to further elucidate the mechanisms of DA toxicity, particularly in terms of chronic exposure, as well as to understand its potential influence on human non-target cells.

Plankton monitoring by microscopy offers a long-term ecological perspective of plankton communities, but detection approaches are uniquely biased. Genetic identification of marine plankton has become standard but is still not used in routine monitoring. This study assesses the value that genetic methods bring to microscopic and flow cytometry monitoring methods in the Western (English) Channel. An 18S high throughput sequencing (HTS) diversity survey of plankton taxa was performed on samples collected from an automated Water and Microplankton Sampler (WaMS) deployed on the Continuous Plankton Recorder platform (CPR) from 2011–2012. This survey detected contrasting but complementary taxa assemblages to that of microscopic surveys, mostly composed of smaller or naked or thin-walled plankton taxa, with most phytoplankton being under 10 µm infrequently recorded by other surveys. Most genetically-detected taxa in the survey were mixotrophic or heterotrophic. In comparison with microscopic phytoplankton counts from the CPR survey and Western Channel Observatory station L4, only 8–12 taxonomic families were common to all three surveys, most of them dinoflagellates, with a bias towards larger diatoms and dinoflagellate taxa in microscopy surveys. Additional quantitative real-time PCR detection of two potentially harmful taxa, the pelagophyte, Aureococcus anophagefferens and four Pseudo-nitzschia from 2011–2013. This revealed that growth of A. anophagefferens was elevated in the summer of 2011, whilst the appearance of Pseudo-nitzschia delicatissima in February of that year contrasted with it’s timing at a coastal station. Smaller phytoplankton measured by flow cytometry had distinct seasonality and abundance in the mid-Atlantic compared to coastal regions reflecting distinct conditions.

An inventory of phytoplankton diversity in 12 Adriatic ports was performed with the port baseline survey. Particular emphasis was put on the detection of harmful aquatic organisms and pathogens (HAOP) because of their negative impact on ecosystem, human health, and the economy. Phytoplanktonic HAOP are identified as species, either native or non-indigenous (NIS), which can trigger harmful algal blooms (HAB). A list of 691 taxa was prepared, and among them 52 were classified as HAB and five as NIS. Records of toxigenic NIS (Pseudo-nitzschia multistriata, Ostreopsis species including O. cf. ovata) indicate that the intrusion of non-native invasive phytoplankton species has already occurred in some Adriatic ports. The seasonal occurrence and abundance of HAOP offers a solid baseline for a monitoring design in ports in order to prevent ballast water uptake and possible expansion of HAOP outside their native region.

7 pages, 4 figures, 1 table, The saxitoxin-producing dinoflagellate Alexandrium minutum is a well-known microalga that causes paralytic shellfish poisoning (PSP) in many coastal regions of the world. In this study, we measured the valve movements of cultivated mussels Mytilus galloprovincialis feeding on toxic A. minutum (n = 29 mussels, shell length = 67.1 ± 3.2 mm, x̅ ± SD) or a morphologically-similar, but toxin-free phytoplankton, Heterocapsa triquetra (n = 24 mussels, shell length = 68.3 ± 2.9 mm). Phytoplankton inoculations were conducted in three sequential “pulses” intended to increase microalgal cell concentrations in a stepwise manner up to ~5000 cells l−1 over a 9-h period. M. galloprovincialis was connected to a non-invasive valvometry apparatus that concurrently measured the magnitude of valve openness once every 0.1 s. It was found that M. galloprovincialis tended to keep its valves open over the course of the experiment, regardless of the phytoplankton species present in water. Standard valve opening metrics, such as the opening duration and opening amplitude, were not significantly affected by the species of phytoplankton. However, the frequency of brief and partial valve closures (microclosures) was significantly influenced by phytoplankton species (P, Financial support was provided by Fisheries and Oceans Canada and the Spanish government through the Ministerio de Economía y Competitividad (projects AGL2013-45945-R and CTM2016-76146-C3-2-R)

Plastic debris carry fouling a variety of class-size organisms, among them harmful microorganisms that potentially play a role in the dispersal of allochthonous species and toxic compounds with ecological impacts on the marine environment and human health. We analyzed samples of marine plastics floating at the sea surface using a molecular qPCR assay to quantify the attached microalgal taxa, in particular, harmful species. Diatoms were the most abundant group of plastic colonizers with maximum abundance of 8.2 x 10(4) cells cm(-2) of plastics, the maximum abundance of dinoflagellates amounted to 1.1 x 10(3) cells cm(-2) of plastics. The most abundant harmful microalgal taxon was the diatom Pseudo-nitzschia spp., including at least 12 toxic species, and the dinoflagellate Ostreopsis cf. ovata with 6606 and 259 cells cm(-2), respectively. The abundance of other harmful microalgal species including the toxic allochthonous dinoflagellate Alexandrium pacificum ranged from 1 to 73 cells cm(-2). In the present study, a direct relationship between the abundance of harmful algal species colonizing the plastic substrates and their toxin production was found. The levels of potential toxins on plastic samples ranged from 101 to 10(2) ng cm(-2), considering the various toxin families produced by the colonized harmful microalgal species. We also measured the rate of adhesion by several target microalgal species. It ranged from 1.8 to 0.3 day(-1) demonstrating the capacity of plastic substrate colonizing rapidly by microalgae. The present study reports the first estimates of molecular quantification of microorganisms including toxin producing species that can colonize plastics. Such findings provide important insights for improving the monitoring practice of plastics and illustrate how the epi-plastic community can exacerbate the harmful effects of plastics by dispersal, acting as an alien and toxic species carrier and potentially being ingested through the marine trophic web. (C) 2018 Elsevier Ltd. All rights reserved.

The Peruvian sea is one of the most productive ecosystems in the world. Phytoplankton production provides food for fish, mammals, mollusks and birds. This trophic network is affected by the presence of toxic phytoplankton species. In July 2017, samples of phytoplankton were obtained from Paracas Bay, an important zone for scallop (Argopecten purpuratus) aquaculture in Peru. Morphological analysis revealed the presence of the genus Pseudo-nitzschia, which was isolated and cultivated in laboratory conditions. Subsequently, the monoclonal cultures were observed by scanning electron microscopy (SEM), and identified as P. multistriata, based on both the morphological characteristics, and internal transcribed spacers region (ITS2) sequence phylogenetic analysis. Toxin analysis using liquid chromatography (LC) with high-resolution mass spectrometry (HRMS) revealed the presence of domoic acid (DA) with an estimated amount of 0.004 to 0.010 pg cell−1. This is the first report of DA from the coastal waters of Peru and its detection in P. multistriata indicates that it is a potential risk. Based on our results, routine monitoring of this genus should be considered in order to ensure public health.

Author(s): Rabinowitz, Sarah; Solano, Joshua J. | Abstract: Introduction: Histamine-mediated angioedema is a potentially life-threatening reaction following exposures that incite mast cell activation. In Florida, red tides are a frequent phenomenon caused by overgrowth of the harmful algae species Karenia brevis, which contain environmentally detrimental brevetoxins. Even in low concentrations, brevetoxins can cause disease in humans through inducing histamine release. We report the first documented case of angioedema associated with red tide exposure.Case Report: A 52-year-old-male presented with severe angioedema encompassing both lips within a few hours after exposure to red tide algae. Other symptoms included voice changes and difficulty swallowing. Laboratory findings revealed complement factors that were within reference range, which ruled out a bradykinin-mediated pathology and supported the diagnosis of histaminergic angioedema. Symptoms resolved after 24 hours in the intensive care unit under management with epinephrine, diphenhydramine, methylprednisolone, and famotidine.Conclusion: In coastal regions, red tide algae should be recognized as a rare cause of acute angioedema. Emergency management of histamine-mediated angioedema should focus on preventing respiratory compromise with frequent airway monitoring and treatment with steroids, antihistamines, and epinephrine.

Harmful algal bloom (HAB) species Alexandrium catenella and Dinophysis acuminata are associated with paralytic shellfish poisoning (PSP) and diarrhetic shellfish poisoning (DSP) in humans, respectively. While PSP and DSP have been studied extensively, less is known about the effects of these HAB species or their associated toxins on shellfish. This study investigated A. catenella and D. acuminata toxicity in a larval oyster (Crassostrea virginica) bioassay. Larval activity and mortality were examined through 96-h laboratory exposures to live HAB cells (10–1000 cells/mL), cell lysates (1000 cells/mL equivalents), and purified toxins (10,000 cells/mL equivalents). Exposure to 1000 cells/mL live or lysed D. acuminata caused larval mortality (21.9 ± 7.0%, 10.2 ± 4.0%, respectively) while exposure to any tested cell concentration of live A. catenella, but not lysate, caused swimming arrest and/or mortality in >50% of larvae. Exposure to high concentrations of saxitoxin (STX) or okadaic acid (OA), toxins traditionally associated with PSP and DSP, respectively, had no effect on larval activity or mortality. In contrast, pectenotoxin-2 (PTX2) caused rapid larval mortality (49.6 ± 5.8% by 48 h) and completely immobilized larval oysters. The results indicate that the toxic effects of A. catenella and D. acuminata on shellfish are not linked to the primary toxins associated with PSP and DSP in humans, and that PTX2 is acutely toxic to larval oysters.

Interactions between algae and bacteria represent an important inter-organism association in aquatic environments, which often have cascading bottom-up influences on ecosystem-scale processes. Despite the increasing recognition of linkages between bacterioplankton and dynamics of dinoflagellate blooms in the field, knowledge about the forms and functions of dinoflagellate-bacteria associations remains elusive, mainly due to the ephemeral and variable conditions in the field. In this study, we characterized the bacterial community associated with laboratory cultures of 144 harmful algal strains, including 130 dinoflagellates (covering all major taxonomic orders of dinoflagellates) and 14 non-dinoflagellates, via high-throughput sequencing for 16S rRNA gene amplicons. A total of 4577 features belonging to bacteria kingdom comprising of 24 phyla, 55 classes, 134 orders, 273 families, 716 genera, and 1104 species were recovered from the algal culture collection, and 3 phyla (Proteobacteria, Bacteroidetes, and Firmicutes) were universally present in all the culture samples. Bacterial communities in dinoflagellates cultures exhibited remarkable conservation across different algal strains, which were dominated by a relatively small number of taxa, most notably the γ-proteobacteria Methylophaga, Marinobacter and Alteromonas. Although the bacterial community composition between dinoflagellates and non-dinoflagellate groups did not show significant difference in general, dinoflagellates harbored a large number of unique features (up to 3811) with relatively low individual abundance and enriched in the potential methylotrophs Methylophaga. While the bacterial assemblages associated with thecate and athecate dinoflagellates displayed no general difference in species composition and functional groups, athecate dinoflagellates appeared to accommodate more aerobic cellulolytic members of Actinobacteria, implying a more possible reliance on cellulose utilization as energy source. The extensive co-occurrence discovered here implied that the relationships between these algal species and the bacterial consortia could be viewed as either bilaterally beneficial (i.e., mutualism) or unilaterally beneficial at least to one party but virtually harmless to the other party (i.e., commensalism), whereas both scenarios support a long-term and stable co-existence rather than an exclusion of one or the other. Our results demonstrated that dinoflagellates-associated bacterial communities were similar in composition, with enrichment of potential uncultured methylotrophs to one-carbon compounds. This work enriches the knowledge about the fundamental functions of bacteria consortia associated with the phycospheres of dinoflagellates and other HABs-forming microalgae.

Cyanobacteria often cause harmful algal blooms and release toxic substances that can harm humans and animals. Accurately modeling these phytoplankton is a step towards predicting, preventing, and controlling such blooms. Certain cyanobacteria species are known to migrate vertically in the water column on a daily cycle. Capturing this behavior is one aspect of modeling their dynamics. Previous studies on modeling cyanobacterial vertical migration are reviewed and summarized. Several models of cyanobacteria vertical movement are tested using data from field studies. These models are applied using both continuum and particle-tracking frameworks. Models range in complexity from simple functions of time to more complicated calculations of cyanobacteria buoyancy. Simple models were often able to predict cyanobacteria migration at low values of vertical diffusion in both types of modeling frameworks. More complicated models of buoyancy change performed better in the particle-tracking framework than in the continuum framework. Analysis of the models developed and tested provides information on the applicability of these models in more complex hydrodynamic and water quality models.

Several coastal countries including France have experienced serious and increasing problems related to Pseudo‐nitzschia toxic blooms. These toxic blooms occur in estuarine and coastal waters potentially subject to fluctuations in salinity. In this study, we document for the first time the viability, growth, photosynthetic efficiency and toxin production of two strains of Pseudo‐nitzschia australis grown under conditions with sudden salinity changes. Following salinity variation, the two strains survived over a restricted salinity range of 30 to 35, with favorable physiological responses, as the growth, effective quantum yield and toxin content were high compared to the other conditions. In addition, high cellular quotas of domoic acid (DA) were observed at a salinity of 40 for the strain IFR‐PAU‐16.1 in comparison with the other strain IFR‐PAU‐16.2 where the cell DA content was directly released into the medium. On the other hand, the osmotic stress imposed at lower salinities, 20 and 10, resulted in cell lysis and a sudden DA leakage in the medium. Intra‐specific variability was observed in growth and toxin production, with the strain IFR‐PAU‐16.1 apparently able to withstand higher salinities than the strain IFR‐PAU‐16.2. On the whole, DA does not appear to act as an osmolyte in response to sudden salinity changes. Since most of the shellfish harvesting areas of bivalve molluscs in France are located in areas where the salinity generally varies between 30 and 35, Pseudo‐nitzschia australis blooms might potentially impact public health and commercial shellfish resources in these places.

In this report, we establish proof-of-principle demonstrating for the first time genetic engineering of a photoautotrophic microorganism for bioremediation of naturally occurring cyanotoxins. In model cyanobacterium Synechocystis sp. PCC 6803 we have heterologously expressed Sphingopyxis sp. USTB-05 microcystinase (MlrA) bearing a 23 amino acid N-terminus secretion peptide from native Synechocystis sp. PCC 6803 PilA (sll1694). The resultant whole cell biocatalyst displayed about 3 times higher activity against microcystin-LR compared to a native MlrA host (Sphingomonas sp. ACM 3962), normalized for optical density. In addition, MlrA activity was found to be almost entirely located in the cyanobacterial cytosolic fraction, despite the presence of the secretion tag, with crude cellular extracts showing MlrA activity comparable to extracts from MlrA expressing E. coli. Furthermore, despite approximately 9.4-fold higher initial MlrA activity of a whole cell E. coli biocatalyst, utilization of a photoautotrophic chassis resulted in prolonged stability of MlrA activity when cultured under semi-natural conditions (using lake water), with the heterologous MlrA biocatalytic activity of the E. coli culture disappearing after 4 days, while the cyanobacterial host displayed activity (3% of initial activity) after 9 days. In addition, the cyanobacterial cell density was maintained over the duration of this experiment while the cell density of the E. coli culture rapidly declined. Lastly, failure to establish a stable cyanobacterial isolate expressing native MlrA (without the N-terminus tag) via the strong cpcB560 promoter draws attention to the use of peptide tags to positively modulate expression of potentially toxic proteins.

Benthic dinoflagellates of the genus Gambierdiscus produce ciguatoxins, compounds that when metabolized in fish and consumed by humans cause ciguatera poisoning (CP). This syndrome, which is widespread in tropical and subtropical regions, has recently been reported also in subtropical-temperate latitudes such as the Canary Islands where CP events have been regularly detected since 2004. This study examined the effect of temperature on the growth of Gambierdiscus isolated from Canary waters: G. australes, G. caribaeus, G. carolinianus, G. excentricus, and G. silvae. From the temperature vs. growth curves, the maximum growth (µm), optimum temperature range for growth (Topt), and the temperature yielding maximum growth (Tm) were estimated for each species. The results revealed temperature-dependent differences in the growth parameters. G. caribaeus had the highest Tm and Topt, followed by G. australes, G. carolinianus, G. silvae, and G excentricus. G. australes tolerated the widest range of temperatures (from 15 ◦C to 29 ◦C), which may explain its broader geographic distribution, both worldwide and across the Canary archipelago. Neither G. excentricus nor G. silvae survived at 29 ◦C whereas G. caribaeus reached mean growth rates (± standard deviation) up to 0.19 ± 0.01 div.day− 1 at that temperature, followed by G. australes (0.16 ± 0.01 div.day− 1) and G. carolinianus (0.14 ± 0.04 div.day− 1). G. caribaeus showed no measurable growth at 19◦C, whereas G. excentricus and G. silvae along with G. australes appeared as the species better adapted to lower temperatures. In an intraspecific variability study of 12 strains of G. australes, the mean (± standard deviation) of µm and Tm were 0.17 ± 0.01 div.day− 1 and 27.7 ± 0.5 ◦C, respectively. An analysis of the shapes and position of the cell nuclei at the different temperatures showed that nuclei characteristic of vegetative cells appeared mainly at 26 ◦C but extreme temperatures resulted in nuclei with a more variable morphology. The presence of putative zygotes at extreme temperatures (17 ◦C, 19 ◦C and 29 ◦C) suggests that sexual reproduction is promoted as an adaptive strategy which could play an important role in the expansion of geographic distribution of Gambierdiscus species., SI

Phytoplankton abundance in relation to some physicochemical characters of the costal water of Arabian Gulf (Saudi Arabia) was studied for one year. The sampling program included 15 locations in Dammam, Saihat, Al-Qatif, Al-Awamia and Safwa. Water samples were analyzed monthly for these parameters; temperature, pH, salinity, dissolved oxygen, nitrite, nitrate, ammonia, carbon dioxide, total chloride, reactive orthophosphate and total phosphorus and alkalinity, also phytoplankton communities were identified and Chlorophyll a was estimated. The results showed that, the high phytoplankton density attaining the maximum (190.3 × 104/m3) during May and June, and the minimum (10.4 × 104/m3) during November and December. Forty Five species belonging to 5 phytoplankton groups were recorded. Bacillariophyceae was the first dominant group forming 48% of the total phytoplankton communities (23 species). The dominant species of Bacillariophyceae were Pleurosigma strigosum, Pleurosigma elongatum, Lyrella clavata, Rhizosolenia shrubsolei, Cylindrotheca closterium, Nitzschia panduriform, Nitzschia longissimia, Amphora sp and Stephanopyxis. Dinophyceae was the second dominant group and formed 31% of the total phytoplankton communities (10 species); the dominant species were Ceratium fusus, Heterosigma sp, Ceratium furca, Prorocentrum triestium, Protoperidinium sp, Gyrodinium spirale, Noctiluca scintillans and Scrippsiella trochoidea. Cyanophyceae formed 13% (5 species) where Nostoc sp, Oscillatoria and Merismopedia sp were the dominant species. Chlorophyceae had 8% (6 species); Scendesmus sp., Chlorella sp., Chlamydomonas sp., Dunaliella salina and Nannochloropsis sp were the dominant species. The Euglinophyceae was rare only one species (Euglina sp). The relationship was positive between the phytoplankton, chlorophyll a and carbon dioxide while negative amongst dissolved oxygen and total nitrogen. This research indicated that the relation between water quality and phytoplankton organisms at the coast of Arabian Gulf (Saudi Arabia) was extremely overlapping and interdependent; any changes in the one component may significantly affect those of the other components.

Prymnesium parvum is a bloom forming haptophyte that has been responsible for numerous fish kill events across the world. The toxicity of P. parvum has been attributed to the production of large polyketide compounds, collectively called prymnesins, which based on their structure can be divided into A-, B- and C-type. The polyketide chemical nature of prymnesins indicates the potential involvement of polyketide synthases (PKSs) in their biosynthesis. However, little is known about the presence of PKSs in P. parvum as well as the potential molecular trade-offs of toxin biosynthesis. In the current study, we generated and analyzed the transcriptomes of nine P. parvum strains that produce different toxin types and have various cellular toxin contents. Numerous type I PKSs, ranging from 37 to 109, were found among the strains. Larger modular type I PKSs were mainly retrieved from strains with high cellular toxin levels and eight consensus transcripts were present in all nine strains. Gene expression variance analysis revealed potential molecular trade-offs associated with cellular toxin quantity, showing that basic metabolic processes seem to correlate negatively with cellular toxin content. These findings point towards the presence of metabolic costs for maintaining high cellular toxin quantity. The detailed analysis of PKSs in P. parvum is the first step towards better understanding the molecular basis of the biosynthesis of prymnesins and contributes to the development of molecular tools for efficient monitoring of future blooms.

Estuaries are coastal wetlands that deserve special attention because they are vulnerable, biodiversity- and service-rich environments. However, estuaries of tropical areas have been scarcely studied regarding water quality and biodiversity in spite of strong developments of their agricultural and livestock sectors. Two estuaries on the Pacific Nicaragua in which snapper cages had been set up were studied regarding water physicochemical properties, microalgae and cyanobacteria along with their dynamics over a hydrological year. Both environments showed excellent water quality (oligotrophy and absence of harmful organisms) arising from their fluvial features. During the rainy season, there was a decline in chemical compounds and microorganisms. In the dry period nutrients and salinity promoted different assemblages of microscopic primary producers that were never too dense. Nutrient inputs from shrimp farms, agricultural crops and snapper cages did not appear to increase eutrophication and cyanobacterial blooms did not occur. Furthermore, the strong fluvial character of these estuaries seems to prevent the entry of harmful dinoflagellates from marine areas.

International audience; This study investigated the genetic diversity of phytoplankton communities in six shallow lagoons located on the French coast of the northwestern Mediterranean Sea that represented a trophic gradient ranging from oligotrophic to hypereutrophic. The phytoplankton communities were sampled once a month from spring (May) to the beginning of autumn (September/early October) in 2012 and fractionated by size. Metabarcodes were generated from cDNAs by targeting the D1-D2 region of the 28S rRNA gene and pyrosequenced using Roche 454 technology. Examination of the annotated barcodes revealed harmful algal species not previously documented in these lagoons. Three ichthyotoxic species belonging to Pfiesteriaceae were detected: Luciella masanensis was relatively widespread and abundant in many samples, whereas Pfiesteria piscicida and Stoeckeria changwonensis were found as single barcode sequences. Furthermore, a phylogenetic analysis of barcodes annotated as belonging to Pfiesteriaceae suggested the existence of two previously undescribed clades. The other toxic or potentially harmful dinoflagellates detected through rare barcodes were Dinophysis acuminata, Vulcanodinium rugosum, Alexandrium andersonii and A. ostenfeldii. The two most abundant dinoflagellate taxa were Gymnodinium litoralis and Akashiwo sanguinea with respect to sequence numbers. Four diatom species from the genus Pseudo-nitzschia that potentially produce domoic acid were identified (P. galaxiae, P. delicatissima, P. brasiliana and P. calliantha). These observations are discussed in terms of the literature and monitoring records related to the identified taxa in this Mediterranean area.

Kongsfjorden (Spitsbergen, Svalbard) is an inlet treated as a model site for studies on the impact of climate change in the Arctic due to its hydrological features. In this research, seven-days monitoring was carried out to evaluate the effects of hydrological variability on phytoplankton biomass and diversity in the late summer period. Temperature, salinity, nutrients, total suspended matter, phytoplankton abundance and biomass were determined for each sample. The thermo-haline properties of the column water seemed to affect phytoplankton communities. Their abundances and biomass were correlated with the amount of the total suspended matter. Moreover, species composition and biomass dramatically changed throughout the study period. Cold-water and Atlantic species were replaced by temperatewarm water dinoflagellates, including harmful species. An increase in phytoplankton biomass as well as the presence of dinoflagellate aggregations, mainly composed of Prorocentrum cf. gracile, were detected. This kind of algal accumulation is a new phenomenon in the Arctic and was probably related to the mobilization of sediment-rich glacial meltwaters. These findings, even if preliminary, suggest the need to study how additional biomass pulses and the increase of harmful species may alter the food web structure and the biogeochemical cycles, leading to major ecosystem changes.

Aquaculture industries are under threat from noxious red tides, but harm can be mitigated by precautions such as early harvesting and restricting fish feeding to just before the outbreak of a red tide. Therefore, accurate techniques for forecasting red-tide outbreaks are strongly needed. Omics analyses have the potential to expand our understanding of the eco-physiology of these organisms at the molecular level, and to facilitate identification of molecular markers for forecasting their population dynamics and occurrence of damages to fisheries. Red tides of marine raphidophytes, especially Chattonella species, often extensively harm aquaculture industries in regions with a temperate climate around the world. A red tide of Chattonella tends to develop just after an input of nutrients along the coast. Chattonella displays diurnal vertical migration regulated by a weak blue light, so it photosynthesizes in the surface layer during the daytime and takes up nutrients in the bottom layer during the nighttime. Superoxide produced by Chattonella cells is a strong candidate for the cause of its toxicity to bacteria and fishes. Here we conducted mRNA-seq of Chattonella antiqua to identify genes with functions closely related to the dynamics of the noxious red tide, such as photosynthesis, photoreception, nutrient uptake, and superoxide production. The genes related to photosynthetic pigment biosynthesis and nutrient uptake had high similarity with those of model organisms of plants and algae and other red-tide microalgae. We identified orthologous genes of photoreceptors such as aureochrome (newly five genes), the cryptochrome/photolyase (CRY/PHR) family (6-4PHR, plant CRY or cyclobutane pyrimidine dimer [CPD] Class III, CPD Class II, and CRY-DASH), and phytochrome (four genes), which regulate various physiological processes such as flagellar motion and cell cycle in model organisms. Six orthologous genes of NADPH oxidase, which produces superoxide on the cell membrane, were found and divided into two types: one with 5–6 transmembrane domains and another with 11 transmembrane domains. The present study should open the way for analyzing the eco-physiological features of marine raphidophytes at the molecular level.

Prorocentrum minimum is a species of marine dinoflagellate that occurs worldwide and can be responsible for harmful algal blooms (HABs). Some studies have reported it to produce tetrodotoxin, however, results have been inconsistent. qPCR and molecular barcoding (amplicon sequencing) using high-throughput sequencing have been increasingly applied to quantify HAB species for ecological analyses and monitoring. Here, we isolated a strain of P. minimum from eastern Australian waters, where it commonly occurs, and developed and validated a qPCR assay for this species based on a region of ITS rRNA in relation to abundance estimates from the cultured strain as determined using light microscopy. We used this tool to quantify and examine ecological drivers of P. minimum in Botany Bay, an estuary in southeast Australia, for over ~14 months in 2016–2017. We compared abundance estimates using qPCR with those obtained using molecular barcoding based on an 18S rRNA amplicon. There was a significant correlation between the abundance estimates from amplicon sequencing and qPCR, but the estimates from light microscopy were not significantly correlated, likely due to the counting method applied. Using amplicon sequencing, ~600 unique actual sequence variants (ASVs) were found, much larger than the known phytoplankton diversity from this region. P. minimum abundance in Botany Bay was found to be significantly associated with lower salinities and higher dissolved CO2 levels.

Tropical shrimp aquaculture systems in New Caledonia regularly face major crises resulting from outbreaks of Vibrio infections. Ponds are highly dynamic and challenging environments and display a wide range of trophic conditions. In farms affected by vibriosis, phytoplankton biomass and composition are highly variable. These conditions may promote the development of harmful algae increasing shrimp susceptibility to bacterial infections. Phytoplankton compartment before and during mortality outbreaks was monitored at a shrimp farm that has been regularly and highly impacted by these diseases. Combining information from flow cytometry, microscopy, pigment and phylogenetic analysis, the presence of Picocyanobacteria, Prasinophyceae and Diatomophyceae were detected as dominant phytoplankton groups and Cryptophyceae, Prymnesiophyceae and Dinophyceae as minor components. At the onset of the first shrimp mortalities, Bacillariophyceae increased while Cyanobacteria, Prymnesiophyceae and Dinophyceae decreased in the water column, followed by proliferation of Prasinophyceae. Several taxa were identified as potential harmful algae (Cyanobacteria, dinoflagellates and Phaeocystis).

6 pages, 3 figures, 1 table, The genus Prymnesium includes several species that produce toxins with cytotoxic, ichthyotoxic, neurotoxic and haemolytic activity. Bloom episodes of Prymnesium species have been reported from several parts of the world (North America, Europe, Africa, Asia and Australia), especially from temperate and subtropical regions and most of them from brackish waters. Blooms cause great economic losses to aquaculture and fisheries around the world. The ichthyotoxic and allelopathic effects of Prymnesium have been linked to the presence of Haemolysin 1, Prymnesins 1 and 2 and, more recently, fatty acids and fatty acid amides. The toxicology of this genus with regard to different growth conditions such as light, nutrients and other parameters has been well documented. It is unknown, however, whether different species and strains from the Prymnesium genus all produce the same types and level of toxins. In this study, we have determined the haemolytic activity of eight different strains from the genus Prymnesium in both exponential and stationary phases of growth. We have also evaluated the efficiency of the extraction solvent., This work was supported by the projects EHUA 12/22 and EHUA 15/30 from the University of the Basque Country (UPV/EHU).

Harmful algae blooms (HABs) cause acute effects on marine ecosystems due to their production of endogenous toxins or their enormous biomass, leading to significant impacts on local economies and public health. Although HAB monitoring has been intensively performed at spatiotemporal scales in coastal areas of the world over the last decades, procedures have not yet been standardized. HAB monitoring procedures are complicated and consist of many methodologies, including physical, chemical, and biological water sample measurements. Each monitoring program currently uses different combinations of methodologies depending on site specific purposes, and many prior programs refer to the procedures in quotations. HAB monitoring programs in Chile have adopted the traditional microscopic and toxin analyses but not molecular biology and bacterial assemblage approaches. Here we select and optimize the HAB monitoring methodologies suitable for Chilean geography, emphasizing on metabarcoding analyses accompanied by the classical tools with considerations including cost, materials and instrument availability, and easiness and efficiency of performance. We present results from a pilot study using the standardized stepwise protocols, demonstrating feasibility and plausibility for sampling and analysis for the HAB monitoring. Such specific instructions in the standardized protocol are critical obtaining quality data under various research environments involving multiple stations, different analysts, various time-points, and long HAB monitoring duration.

Pinnatoxin G (PnTx-G) is a marine cyclic imine toxin produced by the dinoflagellate Vulcanodinium rugosum, frequently detected in edible shellfish from Ingril Lagoon (France). As other pinnatoxins, to date, no human poisonings ascribed to consumption of PnTx-G contaminated seafood have been reported, despite its potent antagonism at nicotinic acetylcholine receptors and its high and fast-acting toxicity after intraperitoneal or oral administration in mice. The hazard characterization of PnTx-G by oral exposure is limited to a single acute toxicity study recording lethality and clinical signs in non-fasted mice treated by gavage or through voluntary food ingestion, which showed differences in PnTx-G toxic potency. Thus, an acute toxicity study was carried out using 3 h-fasted CD-1 female mice, administered by gavage with PnTx-G (8&ndash, 450 &micro, g kg&minus, 1). At the dose of 220 &micro, 1 and above, the toxin induced a rapid onset of clinical signs (piloerection, prostration, hypothermia, abdominal breathing, paralysis of the hind limbs, and cyanosis), leading to the death of mice within 30 min. Except for moderate mucosal degeneration in the small intestine recorded at doses of 300 &micro, 1, the toxin did not induce significant morphological changes in the other main organs and tissues, or alterations in blood chemistry parameters. This acute oral toxicity study allowed to calculate an oral LD50 for PnTx-G equal to 208 g kg&minus, 1 (95% confidence limits: 155&ndash, 281 &micro, 1) and to estimate a provisional NOEL of 120 &micro, 1.

One of the main goals of monitoring cyanobacteria blooms in aquatic environments is to reveal the relationship between cyanobacterial abundance and environmental variables. Studies typically correlate data that were simultaneously sampled. However, samplings occur sparsely over time and may not reveal the short-term responses of cyanobacterial abundance to environmental changes. In this study, we tested the hypothesis that stronger cyanobacteria x environment relationships in monitoring are found when the temporal variability of sampling points is incorporated in the statistical analyses. To this end, we investigated relationships between cyanobacteria and seven environmental variables that were sampled twice yearly for three years across 11 reservoirs, and data from an intensive monitoring in one of these reservoirs. Poor correlations were obtained when correlating data simultaneously sampled. In fact, the 'highly recurrent' role of phosphorus in cyanobacteria blooms is not properly observed in all sampling periods. On the other hand, the strongest correlation values for the total phosphorus x cyanobacteria relationship were observed when we used the variation of sampling points. We have also shown that environment variables better explain cyanobacteria when a time lag is considered. We conclude that, in cyanobacteria monitoring, the best approach to reveal determinants of cyanobacteria blooms is to consider environmental variability. Um dos principais objetivos de monitorar as florações de cianobactérias em ambientes aquáticos é revelar a relação entre a abundância de cianobactérias e as variáveis ambientais. Esses estudos tipicamente correlacionam dados coletados simultaneamente. No entanto, as amostragens ocorrem esparsamente ao longo do tempo e não são capazes de revelar respostas de curto prazo na abundância de cianobactérias frente às mudanças ambientais. Neste estudo, testamos a hipótese de que relações mais fortes entre cianobactérias e variáveis ambientais no monitoramento de cianobactérias são encontradas quando a variabilidade temporal dos pontos de amostragem é incorporada nas análises estatísticas. Para isso, nós investigamos as relações entre cianobactérias e sete variáveis ambientais que foram amostradas duas vezes por ano, durante três anos, em 11 reservatórios, e dados de um monitoramento intensivo em um desses reservatórios. Relações fracas foram encontradas correlacionando dados coletados simultaneamente. De fato, o papel "altamente recorrente" do fósforo nas florações de cianobactérias não foi devidamente observado em todos os períodos de amostragem. Por outro lado, coeficientes de correlação mais fortes entre fósforo total e cianobactérias foram observados quando usamos a variabilidade dos pontos de amostragem. Nós também mostramos que as variáveis ambientais explicam melhor a densidade de cianobactérias quando um atraso é considerado. Nós concluímos que, no monitoramento de cianobactérias, a melhor abordagem para revelar os determinantes das florações de cianobactérias é considerar a variabilidade ambiental.

The ecology of Ostreopsis cf. ovata blooms in the Conero Riviera (N Adriatic Sea) was investigated based on sampling carried out from 2007 to 2012, in order to assess the role of environmental factors associated with blooms. Generally, the temporal trend of blooms showed the first cell appearance at the end of July/early August, the maximum abundances in late-summer at end of September/early October reaching up to 10 6 cells g −1 fw on macrophyte samples, and the decline of the blooms at end October/early November. Calm conditions appeared to be a prerequisite for blooms. When suitable hydrodynamic conditions exist, O . cf. ovata blooms appear to be triggered by a combination of optimal temperature and available nutrients. A water temperature threshold of 25 °C plays a key role in the germination of O . cf. ovata cysts and therefore in bloom onset, and an N:P ratio around Redfield value is a necessary condition to allow cell proliferation. The synergy of higher temperatures and optimal N:P ratios resulted in a higher net growth rate of O. cf. ovata cells. After the onset, blooms can be maintained at temperature values even below 20 °C and at N:P ratios that are in excess of the Redfield ratio. Once the bloom has started it may be maintained not necessarily through high growth rates, but likely through other physiological mechanisms. Bloom decline occurred when temperatures dropped below 18 °C. The net effect of the synergy between local hydrodynamic conditions, temperature, and N and P availability may help to explain why blooms in the northern Adriatic Sea occur differently from those in other Mediterranean regions.

Algae and cyanobacteria are present in all aquatic environments. We do not have a good sense of the extent of human and animal exposures to cyanobacteria or their toxins, nor do we understand the public health impacts from acute exposures associated with recreational activities or chronic exposures associated with drinking water. We describe the Harmful Algal Bloom-related Illness Surveillance System (HABISS) and summarize the collected reports describing bloom events and associated adverse human and animal health events. For the period of 2007-2011, Departments of Health and/or Environment from 11 states funded by the National Center for Environmental Health (NCEH), Centers for Disease Control and Prevention contributed reports for 4534 events. For 2007, states contributed 173 reports from historical data. The states participating in the HABISS program built response capacity through targeted public outreach and prevention activities, including supporting routine cyanobacteria monitoring for public recreation waters. During 2007-2010, states used monitoring data to support196 public health advisories or beach closures. The information recorded in HABISS and the application of these data to develop a wide range of public health prevention and response activities indicate that cyanobacteria and algae blooms are an environmental public health issue that needs continuing attention.

International audience; Effects of simultaneous exposure of Pacific oyster, Crassostrea gigas, to both a harmful dinoflagellate that produces Paralytic Shellfish Toxins (PST), Alexandrium minutum, and cadmium (Cd) and copper (Cu), were assessed. Oysters were exposed to a mix of Cd-Cu with two different diets (i.e. A. minutum or Tisochrysis lutea) and compared to control oysters fed A. minutum or T. lutea, respectively, without metal addition. Metals and PST accumulations, digestive gland lipid composition, and cellular and biochemical hemolymph variables were measured after 4 days of exposure. Oysters exposed to Cd-Cu accumulated about thirty-six times less PSTs than oysters exposed to A. minutum alone. Exposure to Cd-Cu induced significant changes in neutral lipids (increase in diacylglycerol - DAG - and decrease in sterols) and phospholipids (decreases in phosphatidylcholine, phosphatidylethanolamine, cardiolipin and ceramide aminoethylphosphonate) of digestive gland suggesting that lipid metabolism disruptions and/or lipid peroxidation have occurred. Simultaneously, concentrations, percentages of dead cells and phenoloxidase activity of hemocytes increased in oysters exposed to metals while reactive oxygen species production of hemocytes decreased. Feeding on the harmful dinoflagellate A. minutum resulted in significant decreases in monoacylglycerol (MAG) and DAG and ether glycerides (EG), as well as significant increases in hemocyte concentration and phagocytic activity as compared to oysters fed T. lutea. Finally, the present study revealed that short-term, simultaneous exposure to Cd-Cu and A. minutum may induce antagonistic (i.e. hemocyte concentration and phagocytosis) or synergic (i.e. DAG content in digestive gland) effects upon cellular and tissular functions in oysters.

The distribution and abundance of epiphytic and planktonic Coolia monotis Meunier along the Alexandria coast were studied through five annual cycles; from summer 2005 to summer 2010 at four sites: Abu Qir Beach, Stanly, Eastern Harbour and Mex Beach. The morphology of C. monotis was characterised by noticeable small size specimens in the E.H., the D.V. diameter is not exceeding 20 μm and width between 10 and 15 μm. On the other hand, the epiphytic specimens showed a normal cell size. The species was most abundant as planktonic form in the E.H., reaching 15.2 × 103 cell l−1 during summer 2010. The epiphytic forms showed lower density, with a maximum of 454 cell g−1 fwm during autumn 2005, 2006 and summer from 2007 to 2010. The fluctuations of cell abundance of C. monotis showed a significant negative correlation with salinity and a weak positive correlation with temperature. This study represents the first report of Coolia monotis Meunier in the Egyptian Mediterranean waters.

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.

We present experiments that examined the grazing and survivorship of zooplankton native (Acartia tonsa) and non-native (Artemia salina) to NY (USA) estuaries when exposed to blooms and cultures of the three harmful algae native to NY, Alexandrium fundyense, Aureococcus anophagefferens (strains CCMP 1850 and CCMP 1984) and Cochlodinium polykrikoides. During experiments with cultures of A. anophagefferens, clearance rates (CR) of A. salina were significantly greater than those of A. tonsa for both algal strains examined. A. salina fed on cultures of C. polykrikoides at higher rates than all phytoplankton species examined, including the control diet (Rhodomonas salina), and faster than rates of A. tonsa fed C. polykrikoides. During experiments with A. fundyense, A. salina actively grazed all cell concentrations (250–1500 cells ml−1) while A. tonsa did not feed at any concentration. Percent mortality of A. salina and A. tonsa fed A. fundyense for 48 h were 43 ± 7.7% and 72 ± 7.8%, respectively, percentages significantly higher than those of individuals fed all other algal diets. During 25 field experiments using natural blooms of the three HAB species performed across six NY estuaries, A. salina significantly (p < 0.05) reduced cell densities of A. anophagefferens, C. polykrikoides, and A. fundyense relative to the control treatments in all but one experiment. The sum of these findings demonstrates that a failure to graze these HABs by the indigenous copepod, A. tonsa, may permit blooms to occur. In addition, the ability of A. salina to graze these HABs at densities that were inhibitory to A. tonsa suggests that A. salina could, in some circumstances, be considered as a part of mitigation strategy for these events. Fil: Marcoval, Maria Alejandra. Stony Brook University; Estados Unidos. Consejo Nacional de Investigaciones Científicas y Técnicas; Argentina Fil: Pan, Jerónimo. Stony Brook University; Estados Unidos Fil: Tang, Yingzhong. Stony Brook University; Estados Unidos Fil: Gobler, Christopher J.. Stony Brook University; Estados Unidos

18 pages, 6 figures, 6 tables, The present study identified and quantified dinoflagellate cysts in surface sediments from three Mediterranean lagoons. Sediment samples were recovered from 11 stations in May 2009 at Cabras Lagoon, eight stations in May 2010 at Corru S'Ittiri Lagoon, and five stations in May 2011 at Santa Giusta Lagoon. Fifty-three dinoflagellate cyst morphotypes were identified. Sixteen species are first reports for the lagoons, and two for the Mediterranean Sea. Moreover, a new Scrippsiella species was discovered in Cabras. Seven harmful algal species were identified, primarily belonging to the potentially toxic genus Alexandrium. Total cyst abundance, number of morphotypes, and assemblages varied among lagoons, and each lagoon showed a distinct morphotype composition. A degree of heterogeneity was also detected within lagoon. Cabras and Santa Giusta cyst assemblages were characterised by morphotypes belonging to the autotrophic genus Scrippsiella , whereas Corru S'Ittiri assemblages showed dominance of heterotrophic morphotypes, including Protoperidinium cf tricingulatum. Differentiation among lagoons was also evident according to environmental conditions. Salinity proved to be a fundamental variable in determining total cyst abundance, morphotype number, and composition. This study was among the first to examine dinoflagellate cyst composition in coastal lagoons, especially from the Mediterranean region, and contributed data that increased our knowledge of cyst-producing dinoflagellates in these environments., The Autonomous Region of Sardinia provided the principle financial support for the Cabras and Santa Giusta monitoring activities, and the Zoumgest project provided the financial support required for the Corru S'Ittiri Lagoon. The Autonomous Region of Sardinia funded Research Project – Master and Back financed C. T. Satta work. A Postdoctoral Grant from the Spanish Ministry of Economy and Competitiveness partially funded S. Anglès work

International audience; Blood cells or hemocytes are the major component of the cellular immune responses in bivalves. Recent evidences have demonstrated that the immune functions of these cells can be impaired by harmful algae and their toxins. In this study, we compared the in vitro effect of the toxic dinoflagellate Alexandrium minutum and its purified paralytic toxin, saxitoxin (STX) on the oyster Crassostrea gigas hemocytes. Oyster hemocytes were incubated with A. minutum or STX, and their size, complexity, viability, phagocytic activity and reactive oxygen species (ROS) production were analyzed through flow cytometry. Also, expression of eleven defense-related genes was evaluated through real-time quantitative PCR. Cell incubation with both, the noxious microalga and STX did not alter hemocyte viability neither complexity. However, their phagocytic capacity and ROS production were significantly affected by both treatments. On the other hand, only A. minutum exposure resulted in increase of hemocyte size. Transcript levels of the gene encoding the cytokine Interleukin-17 was significantly higher (4.5-fold) after hemocytes were exposed to A. minutum, which contrasted with a decreased (7-fold) number of transcripts, when hemocytes were treated with STX. The purified toxin also led to lower transcript levels of a cytochrome P450 gene, the CYP356A1 (3.5-fold). On the other hand, hemocytes incubated with STX presented higher mRNA levels of antimicrobial peptide defensin-2 (7-fold) and heat shock protein Hsp70 (2-fold) genes. Altogether, these results suggest that the noxious alga A. minutum and STX can indeed negatively affect immunocompetence of C. gigas hemocytes, although in distinct manners. These differences could be due to a lower STX bioavailability in living microalgal cultures or alternatively to the presence of allelopathic compounds produced by living microalgae. This is the first report comparing the transcription of defense-related genes in oysters exposed to a harmful alga and its purified toxin.

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.

14 pages, 10 figures, 7 tables, supplementary data http://dx.doi.org/10.1016/j.ecolmodel.2016.07.009, The dinoflagellate Karlodinium and the diatom Pseudo-nitzschia are bloom-forming genera frequently present in Alfacs Bay. Both microalgae are associated with toxic events. Therefore, understanding their population dynamics and predict their occurrence in short-term is crucial for an optimal management of toxic events for the local shellfish production and ecosystem managers. Artificial neural networks have been successfully used to model the complex nonlinear dynamics of phytoplankton. In this study, this approach was applied to predict absence-presence and abundance of Karlodinium and Pseudo-nitzschia microalgae in Alfacs Bay (NW Mediterranean) using biological and/or environmental variables. Neural Interpretation Diagram (NID) and Connection Weight Approach (CWA) methodologies were applied to obtain ecological information from the models. The dataset used was long-term (1990–2015) time series of environmental and phytoplankton variables from different monitoring stations established in Alfacs Bay (Ebre Delta), meteorological data and Ebre River flow rates. Several models were presented. The best ones were achieved for one-week ahead procedures performed with environmental and biological variables using all the available data. A sensitivity analysis showed the larger the data set used, the better the models obtained. However, Karlodinium absence-presence models developed with five years of data present high accuracy. The size of the neural networks denotes complex relationships between environmental and phytoplankton variables. The environmental variables had stronger influence on the abundance models while biological variables had more importance in the absence-presence models. These results highlight a complex ecosystem in Alfacs Bay involving anthropogenic, climatic and hydrologic factors forcing phytoplankton dynamics. In addition, a change in the ecosystem dynamics regarding Karlodinium is detected. The configuration and the accuracy achieved with the models allow their use in different real-world applications as automated systems and/or monitoring programs, This research, including CG contract, has been funded by the Economy and Competitiveness Ministry of Spain under the program INNPACTO 2011 (“PURGADEMAR” project; IPT-2011-1707-310000) and INIA (project PROMAQUA RTA2013-00096-00-00)

In many coastal areas the abundant proliferation of microalgae producing biotoxins determines the occurrence of Harmful Algal Blooms (HABs). Their presence in temperate waters is well documented and often associated with marine toxin-derived disease. The occurrence and toxicity of three harmful microalgae (Amphidinium carterae, Coolia cfr. monotis and Ostreopsis cfr. ovata) from the northern Ionian Sea (Mediterranean Sea) is hereby reported. The three dinoflagellates were sampled both on macroalgae and water and their morphology and occurrence were compared to those of other Mediterranean sites. The toxicity of the three cultured strains was tested by Artemia salina and hemolysis tests and their effects on the first stages of the sea urchin development was also evaluated. The contemporary presence of the three species inhibited the in vitro sea urchin embryonic development. But this action could be ascribed to the sole Ostreopsis as the addition of the single species to the sea urchins embryos evidenced no effects in presence of Amphidinium or Coolia cells, and an irregular segmentation in presence of Ostreopsis. In particular, this latter species exerted a cytotoxic effect in a dose-dependent manner, with a production of deformed embryos even at very low cell concentration (42 cells mL−1). Nevertheless, when algal cell lysate was added, some effects on the sea urchin development was detected for each dinoflagellate, and also in this case Ostreopsis has proved to be the most toxic species. However, the lysate of Amphidinium and Ostreopsis strongly affects the A. salina nauplii vitality, while the hemolytic activity was very low for Amphidinium and Coolia lysate and very strong for Ostreopsis. Our results highlight the importance to monitoring the presence of these dinoflagellates whose effects may also be reflected on the early life stages of marine organisms, especially those species that are important from both an ecological and economic point of view, as the sea urchins are.

10 páginas. -- Magda vila ... et al., As shown in this report, the integration of a research project with a monitoring program improves the detection and management of Ostreopsis blooms in Catalonia. The research project benefits from information previously obtained from several localities by the monitoring program, which in turn profits from the specific findings and conclusions contributed by the research project.

There may be biogeographic aspects of interactions between toxic phytoplankton and their zooplankton grazers. Grazers that naturally co-occur with a given species of toxic phytoplankton may have evolved mechanisms to counteract adverse effects of phytoplankton toxins. Conversely, grazers that have no co-evolutionary experience with a toxic phytoplankter may be more susceptible to its effects. This may also be true for toxic metabolites other than neurotoxins that adversely affect copepod reproduction and development such as the well-described diatom oxylipins. In order to test this hypothesis, we investigated interactions in terms of feeding, reproductive success and survival of a copepod feeding on a potentially toxic dinoflagellate species with which the copepod does not naturally co-occur: the red tide dinoflagellate Karenia brevis from the Gulf of Mexico and the copepod Calanus helgolandicus from the Mediterranean. We additionally compared interactions of these two allopatric plankters with various co-occurring sympatric ones, including the copepod Temora stylifera and a non-neurotoxic strain of the dinoflagellate Alexandrium tamarense, both from the Mediterranean, and as a control, the non-toxic dinoflagellate Prorocentrum minimum. Both Mediterranean copepods fed, reproduced and survived on diets of K. brevis from the Gulf of Mexico, but neither copepod did as well on this diet as on diets of A. tamarense from the Mediterranean or the control diet of P. minimum. Of the two copepods, T. stylifera fared worse on a diet of K. brevis than did C. helgolandicus. Egg hatching success for both copepods feeding on K. brevis was sharply diminished over the seven days of the experiments. This was surprising, since we subsequently discovered that our culture of K. brevis contained no brevetoxins. However, our culture appeared to contain another as yet unidentified compound. Similarly, we have previously demonstrated that our culture of A. tamarense, which contained no saxitoxins or aldehydes, did contain another unidentified compound that reduced egg hatching success by inhibiting sperm from fertilizing eggs. Thus, although diets of the dinoflagellate from the Gulf of Mexico (K. brevis) caused adverse effects on both of the Mediterranean copepods, T. stylifera was also adversely affected by the Mediterranean dinoflagellate (A. tamarense). The copepod T. stylifera which co-occurs in the Gulf of Mexico with the dinoflagellate K. brevis was far more adversely affected by feeding on a sole diet of this dinoflagellate than was the Mediterranean copepod (C. helgolandicus) which never naturally co-occurs with this dinoflagellate. Thus, any putative biogeographic effects of copepods feeding on allopatric versus sympatric dinoflagellates did not clearly emerge from our data. However, since both species of dinoflagellates appear to contain additional undescribed chemicals other than the neurotoxins for which they are best known, and these compounds are potentially related to the adverse effects we recorded, it appears that understanding of the biogeography must await further defining the biochemistry of some harmful algae:grazer interactions.

Pseudochattonella farcimen (Eikrem, Edvardsen, et Throndsen) is a unicellular alga belonging to the Dictyochophyceae (Heterokonta). It forms recurring blooms in Scandinavian coastal waters, and has been associated to fish mortality. Here we report the sequencing and analysis of 10,368 expressed sequence tags (ESTs) corresponding to 8,149 unique gene models from this species. Compared to EST libraries from other heterokonts, P. farcimen contains a high number of genes with functions related to cell communication and signaling. We found several genes encoding proteins related to fatty acid metabolism, including eight fatty acid desaturases and two phospholipase A2 genes. Three desaturases are highly similar to Δ4-desaturases from haptophytes. P. farcimen also possesses three putative polyketide synthases (PKSs), belonging to two different families. Some of these genes may have been acquired via horizontal gene transfer by a common ancestor of brown algae and dictyochophytes, together with genes involved in mannitol metabolism, which are also present in P. farcimen. Our findings may explain the unusual fatty acid profile previously observed in P. farcimen, and are discussed from an evolutionary perspective and in relation to the ichthyotoxicity of this alga.

Ostreopsis cf. ovata, a benthic dinoflagellate often blooming along the Mediterranean coasts, has been associated with toxic events ranging from dyspnea to mild dermatitis. In late September 2009, an Ostreopsis cf. ovata bloom occurred in the Gulf of Trieste (Northern Adriatic Sea; Italy), causing pruritus and mild dermatitis in beachgoers. An integrated study was initiated to characterize Ostreopsis cells by light and confocal microscopy, PCR techniques, immunocytochemistry, and high resolution liquid chromatography-mass spectrometry (HR LC-MS). The presence of Ostreopsis cf. ovata of the Atlantic/Mediterranean clade was unambiguously established by morphological and genetic analyses in field samples. Several palytoxin-like compounds (ovatoxin-a,-b,-c,-d,-e) were identified by HR LC-MS, ovatoxin-a being the most abundant (45-64 pg/cell). Surprisingly, no palytoxin was detected. For the first time, monoclonal and polyclonal antipalytoxin antibodies revealed the intracellular cytoplasmic localization of ovatoxins, suggesting their cross-reactivity with these antibodies. Since harmful dinoflagellates do not always produce toxins, the immunocytochemical localization of ovatoxins, although qualitative, can provide an early warning for toxic Ostreopsis cells before their massive diffusion and/or concentration in seafood.

The Greek coastal waters are subjected to harmful algal bloom (HAB) phenomena due to the occurrence of species characterized as toxic (TX), potentially toxic (PT), and non-toxic, high biomass (HB) producers causing harm at multiple levels. The total number of (TX), (PT) and (HB) algae reported in this work are 61, but only 16 species have been associated with the occurrence of important HABs causing damage in the marine biota and the water quality. These phenomena are sporadic in time, space and recurrence of the causative species, and are related to the anthropogenically-induced eutrophication conditions prevailing in the investigated areas.

Blooms of the cyanobacterium Trichodesmium occur in massive colored patches over large areas of tropical and subtropical oceans. Recently, the interest in such events has increased given their role in major nitrogen and carbon dioxide oceanic fluxes. Trichodesmium occurs all along the Brazilian coast and patches frequently migrate towards the coast. In this paper we screen the toxicity and toxin content of Trichodesmium blooms off the coast of Bahia state. Four samples, collected from February to April 2007, were analyzed. Organisms were identified and assessed for toxicity by means of several methods. Analogues of microcystins, cylindrospermopsins and saxitoxins were analyzed using HPLC. Microcystins were also assayed through ELISA. Results showed dominance of T. erythraeum, which makes up as much as 99% of cell counts. Other organisms found in smaller quantities include the dinoflagellates Prorocentrum minimum and P. rhathymum. Extracts from all samples delayed or interrupted sea urchin larval development, but presented no acute toxicity during a mouse bioassay. Saxitoxin congeners and microcystins were present at low concentrations in all samples, occurrences that had not previously been reported in the literature. Despite our finding of saxitoxin analogues and microcystins in Trichodesmium blooms, these toxins do not represent a potential harm to human health by primary contact. We conclude, based on our results and those reported in the recent literature, which differ from results published in 1963, that although toxins are present, there is no evidence that T. erythraeum blooms represent a threat to humans.

Blooms of the marine dictyochophyte Pseudochattonella farcimen have been associated with fish kills, but attempts to verify ichthyotoxicity of this microalga under experimental conditions have not been successful. In the early spring of 2009 and 2011, P. farcimen bloomed in the inner Danish waters. The blooms occurred at a seawater temperature of ~2°C and correlated with extensive kills of farmed salmonid fish (2009) and wild populations (2011). Several strains of P. farcimen were isolated from the 2009 bloom. However, exposure of rainbow trout Oncorhynchus mykiss to laboratory-grown P. farcimen cultures did not reveal any toxic effects. During the 2011 bloom, fish were exposed to bloom water under both laboratory and field conditions. While no clinical effect was observed on fish incubated in bloom water in the laboratory trial, a remarkable difference was seen in the field trial between rainbow trout kept in tanks supplied with a continuous flow of filtered versus non-filtered bloom water. Histological examination of the gill tissue revealed karyorrhexis and epithelial loosening in the affected fish. Microscopy analysis of algal cell morphology suggested that mucocysts detected on the cell surface only in freshly sampled bloom water might be associated with ichtyotoxicity. Blooms of the marine dictyochophyte Pseudochattonella farcimen have been associated with fish kills, but attempts to verify ichthyotoxicity of this microalga under experimental conditions have not been successful. In the early spring of 2009 and 2011, P. farcimen bloomed in the inner Danish waters. The blooms occurred at a seawater temperature of ~2°C and correlated with extensive kills of farmed salmonid fish (2009) and wild populations (2011). Several strains of P. farcimen were isolated from the 2009 bloom. However, exposure of rainbow trout Oncorhynchus mykiss to laboratory-grown P. farcimen cultures did not reveal any toxic effects. During the 2011 bloom, fish were exposed to bloom water under both laboratory and field conditions. While no clinical effect was observed on fish incubated in bloom water in the laboratory trial, a remarkable difference was seen in the field trial between rainbow trout kept in tanks supplied with a continuous flow of filtered versus non-filtered bloom water. Histological examination of the gill tissue revealed karyorrhexis and epithelial loosening in the affected fish. Microscopy analysis of algal cell morphology suggested that mucocysts detected on the cell surface only in freshly sampled bloom water might be associated with ichtyotoxicity.

In the ocean, microscopic plankton algae constitute a crucial food supply for filter- feeding bivalves, shellfish (oyster, mussels, scallop and clams), as well as larvae of commercially important crustaceans and finfish. In most areas, the proliferation of plankton algae (so called "algal blooms"; consisting of millions of cells per liter) is beneficial for aquaculture, recreational and commercial fisheries. However, sometimes algal blooms may have a negative impact. Mass occurrences of harmful microalgae, harmful algal blooms (HABs), are a globally growing concern. Most HAB species cause harm due to their production of toxins. HAB species may kill marine wildlife directly, or the algal toxins may accumulate in the food web, causing illness and mortality of fish, seabirds, marine mammals and humans consuming the toxic seafood products. There is scientific consensus that the HAB problem is increasing globally, with increasing numbers of toxic blooms and associated economical loss. The most significant reasons for the increased occurrence of HABs are increased eutrophication of marine coastal areas and spreading of harmful species to new areas. During the last three decades, the Arabian Gulf area has experienced massive marine mortalities, resulting in serious economic losses (Kuwait, Saudi Arabia, Iran, U A E, Oman, Bahrain and Qatar). A documented example is an eight-month-bloom during 2008-2009 of the dinoflagellate Cochlodinium polykrikoides, killing thousands of tons of fish, hampering traditional fisheries, impacting tourism, forcing closure of desalination plants, and damaging coral reefs. The frequency and severity of HAB events are increasing in the Arabian Gulf, and the distribution of harmful species within the region appears to be expanding. In this situation, exchange of information and cooperative research has become obvious to scientists working in the Gulf region. To date, only a small number of studies have been conducted on diversity, distribution and toxicity of HABs in the Arabian Gulf, including Qatari waters. A project on harmful algae is needed here, focusing on biodiversity and ecology based on state of the art techniques, and involving capacity building of local Qatari staff.

Puffer fish poisoning (PFP) is usually caused by ingestion of tetrodotoxins (TTXs) found naturally in certain species of puffer fish (Halstead 1967; Mosher and Fuhrmann 1984). In Japan, 20–100 people die annually from PFP, in spite of stringent controls by authorities (Ogura 1971). TTXs can cause fatal human poisoning, which is similar to paralytic shellfish poisoning (PSP) caused by saxitoxins (STXs). PSP is caused by the consumption of toxic shellfish (Shumway 1990) and rarely by fish that have have become toxic after feeding on STX-producing microalgae (Maclean 1979). As well as TTXs, STXs have also been found in at least 12 marine and freshwater puffer fish species in Asia (Ahmed et al. 2001; Kodama et al. 1983; Kungsuwan et al. 1997; Nakamura et al. 1984; Nakashima et al. 2004; Sato et al. 1997, 2000; Zaman et al. 1997), but their bioorigin has not been identified. TTXs are chemically distinct from STXs, but both neurotoxins produce similar symptoms in mammals because they act on site 1 of the voltage-dependent sodium channel, blocking the influx of sodium into excitable cells and restricting signal transmission along nerve and muscle membranes (Ahmed 1991). The symptoms of traditional PFP from TTXs and of PSP from STXs include tingling and numbness of the mouth, lips, tongue, face, and fingers; paralysis of the extremities; nausea; vomiting; ataxia; drowsiness; difficulty in speaking; progressively decreasing ventilatory efficiency; and finally in extreme cases, death by asphyxiation caused by respiratory paralysis (Ahmed 1991; Catterall 1985; Kao 1993). PFP cases in Europe (Kao 1993) and Mexico (Nunez-Vazquez et al. 2000) have occasionally been reported. In the United States, PFP has been associated with imports of puffer fish [Centers for Disease Control and Prevention (CDC) 1996]; rarely have fatalities occurred after the consumption of indigenous puffer fish. In Hawaii, white-spotted puffer fish, Arothron hispidus, were implicated in seven deaths (Ahmed 1991). Until 1974, seven PFP cases in Florida, outside of the Indian River Lagoon (IRL), were caused by the consumption of locally caught “blowfish” or puffer fish (Ahmed 1991; Benson 1956; Bigler 1999; Hemmert 1974; Mosher and Fuhrmann 1984). These cases included three fatalities, likely from TTX; for example, one woman died 45 min after consuming toxic liver from a checkered puffer fish (Sphoeroides testudineus) (Benson 1956). The toxins involved in the previous Florida PFP cases were not characterized, but because PFP is usually associated with TTX, investigators likely assumed that TTX was the cause (Benson 1956; Bigler 1999; Hemmert 1974). Tissues from Florida bandtail (Sphoeroides spengleri), checkered, and southern puffer fish (Sphoeroides nephelus) were found to be lethal in the mouse bioassay (MBA) (Burklew and Morton 1971; Lalone et al. 1963), but, again, the toxins were not determined. Until January 2002 the harvest and consumption of puffer fish from the IRL was not a risk to public health. Since then (until May 2004), however, 28 PFP cases occurring in Florida (n = 21), New Jersey (n = 3), Virginia (n = 2), and New York (n = 2) caused by puffer fish originating from the IRL were reported (Bodager 2002; CDC 2002a, 2002b). Analyses of toxins from unidentified puffer fish fillet remnants from one of the early 2002 PFP cases in New Jersey revealed STXs (Quilliam et al. 2004), not TTXs, a distinction that alone could not be made on the basis of consumer symptoms or traditional screening methods (i.e., MBA). During 2002–2004, all PFP cases were linked to puffer fish originating from the northern IRL and the Banana River on Florida’s east coast (Figure 1). Except for one case, where puffer fish were commercially harvested and reached a New Jersey fish market, puffer fish were caught recreationally [Bodager 2002; Florida Fish and Wildlife Conservation Commission (FWC) 2004]. In April 2002, state and federal officials issued health advisories, and the FWC banned puffer fish harvesting in the IRL, a ban that currently remains in effect. In New York on 14 October 2002, two PFP cases were caused from fish caught near Titusville, Florida, but frozen in March 2002 before the harvesting ban (Bodager D, personal communication). This case demonstrated the stability of toxins in puffer fish frozen for almost 9 months. Figure 1 Map showing locations (circles) in the Indian River Lagoon, Florida, where toxic puffer fish in the SPFP incidents originated (FWC 2004). Sample collections of puffer fish and Pyrodinium bahamense were conducted throughout this area and further south ... Because STXs had not previously been identified in Florida’s marine waters and their distribution, source, and origin were unknown in April 2002 (Abbott et al. 2003; Landsberg et al. 2002), we initiated an intensive survey of biota in the IRL. In this article we present a summary from 3 years of monitoring, as well as the first report of the putative toxin source.

In Japan, a paralytic shellfish poison (PSP) event caused by Gymnodinium catenatum Graham, a naked, colonial and PSP dinoflagellate, was first reported from Senzaki Bay, western part of the Sea of Japan in 1986. Since then, the geographical distribution of this species seems to have expanded through West Japan. To clarify the possibility of its migration being due to anthropogenic agency the historical occurrence of G. catenatum in Japanese coastal waters was examined; core sediments that preserve an historical record of this species were collected from several coastal areas where it had been observed. From the cores collected from Imari Bay, smaller spherical cysts ornamented with fine reticulated structure and with epicystal archeopyle are found which are probably different from Gymnodinium nolleri. Low cyst concentrations of G. catenatum in these cores may have resulted from such a characteristic physiological feature as chain formation of its vegetative form. The earliest occurrence of this species was recorded in sediment of Omura Bay dated to ca. 1700 A.D. This fact strongly suggests that the occurrence of G. catenatum in Japan is natural and that it has not been artificially introduced from different areas., Paleontological research. 1997, 10(2), p.117-125

This investigation included a 2-year monitoring program aimed at assessing the abundance and distribution of harmful marine phytoplankton along the Southern Adriatic coast of Italy. Monthly sampling was conducted from April 1995 to March 1997 at four sampling stations along four transects, to determine the temporal and spatial presence/abundance of the potentially toxic Pseudonitzschia species. The study focussed on the most abundant taxa Pseudo-nitzschia calliantha and Pseudo-nitzschia delicatissima, the identities of which were confirmed by TEM on cleaned net material. The distribution patterns of these potential Amnesic Shellfish Poisoning (ASP) toxin producers were statistically analysed by Generalised Linear Model ANalysis Of VAriance, Principal Component Analysis (GLM ANOVA, PCA) and Spearman’s correlation analysis in order to address relationships between environmental variable and population dynamics. Abundances displayed horizontal and vertical structure in the study area. Inter-annual variability was also observed for both species that appeared to respond differently to the environmental factors investigated. Distributions of P. calliantha showed a stronger seasonality and was more correlated with winter water conditions than P. delicatissima, which in turn exhibited a broader temporal distribution and appeared independent from major environmental constraints. This is the first report of the occurrence and dynamics of P. calliantha and P. delicatissima populations in Southern Adriatic coastal waters.