Anatoxin-a(S) is the most potent natural neurotoxin produced by fresh water cyanobacteria. It is also the least understood and monitored. Although this potent cholinesterase inhibitor was first reported in the 1970s and connected with animal poisonings, the lack of chemical standards and identified biosynthetic genes together with limited diagnostics and acute reactivity of this naturally-occurring organophosphate have limited our understanding of its environmental breadth and human health implications. Anatoxin-a(S) irreversibly inhibits acetylcholinesterase much like other organophosphate agents like paraoxon. It is however often confused with the similarly named anatoxin-a that has a completely different chemical structure, mechanism of action, and biosynthesis. Herein we propose renaming of anatoxin-a(S) to clarify its distinct structure and mechanism and to draw renewed attention to this potent natural poison. We propose the new name guanitoxin (GNT) to emphasize its distinctive guanidino organophosphate chemical structure.

Sea surface temperatures in the world’s oceans are projected to warm by 0.4–1.4 °C by mid twenty-first century causing many tropical and sub-tropical harmful dinoflagellate genera like Gambierdiscus, Fukuyoa and Ostreopsis (benthic harmful algal bloom species, BHABs) to exhibit higher growth rates over much of their current geographic range, resulting in higher population densities. The primary exception to this trend will be in the tropics where temperatures exceed species-specific upper thermal tolerances (30–31 °C) beyond which growth slows significantly. As surface waters warm, migration to deeper habitats is expected to provide refuge. Range extensions of several degrees of latitude also are anticipated, but only where species-specific habitat requirements can be met (e.g., temperature, suitable substrate, low turbulence, light, salinity, pH). The current understanding of habitat requirements that determine species distributions are reviewed to provide fuller understanding of how individual species will respond to climate change from the present to 2055 while addressing the paucity of information on environmental factors controlling small-scale distribution in localized habitats. Based on the available information, we hypothesized how complex environmental interactions can influence abundance and potential range extensions of BHAB species in different biogeographic regions and identify sentinel sites appropriate for long-term monitoring programs to detect range extensions and reduce human health risks.

Some benthic dinoflagellates produce toxins that can affect other organisms including humans, and their proliferation seems to be related to the environmental variability. For this reason, the present study aims to compare the structural variation of potentially toxic dinoflagellates associated with the seagrass Thalassia testudinum from two nearby systems, with different environmental characteristics in Colombian Caribbean, corresponding to a brackish water coastal lagoon and an adjacent bay. Between January 2014 and December 2015, leaves of T. testudinum were collected monthly to obtain the dinoflagellates. Salinity, temperature, dissolved oxygen, pH, nutrients and total suspended solids (TSS) were measured, and precipitation data and the Oceanic Niño Index (ONI) were obtained. Dinoflagellates were detached from the leaves, morphologically identified by analyzing their thecal plates arrangements, and quantified using a Sedgewick-Rafter chamber. The information was analyzed using standard statistics and regression models. Fourteen species of potentially toxic epiphytic dinoflagellate belonging to four genera were recorded, being Prorocentrum the most representative in number of species. The maximum density, dominated by P. lima, were found in Bahía Chengue during the rainy season of 2014 (18452 and 20109 cells g−1 w.w.), with salinity of 35.50, high temperatures (>29.60 °C), dissolved oxygen >6 mg L−1, pH close to 8 and TSS >85 mg L−1. Densities at the Lagoon were lower than 80 cells g−1 w.w. with the highest values of Prorocentrum sp.1 under different environmental conditions. With the statistical relationships between the most abundant species and the main environmental variables, fundamental niche models were proposed in which cells could proliferate. The degree of risk to human health due to the presence of these potentially toxic epiphytic dinoflagellates will not be resolved until their toxicity discarded.

Sexual reproduction (SR), hallmarked by meiosis, is widespread in eukaryotes. In phytoplankton, SR has been observed in many lineages, but molecular information on SR or meiosis of harmful algal bloom (HAB) species is scarce. The raphidophyte Heterosigma akashiwo is a globally distributed and devastating HAB species, but molecular evidence of its SR or meiosis is lacking. Here, to address the gap of knowledge, the presence of meiotic genes in H. akashiwo were examined. Interestingly, seven meiosis-specific or related genes (SPO11, MND, RAD21, RAD51, MSH2, MSH6 and MEI2) were identified from H. akashiwo transcriptomes. Furthermore, expression patterns of these genes except MSH6 (excluded due to primer failure) were investigated using quantitative reverse-transcription PCR. Results showed that the examined genes exhibited similar diel rhythms, typically, highest in early dark period and then gradually decreasing until mid-day. In addition, the expression of these six genes was not higher in the stationary growth stage than in the exponential stage, as would be expected if meiosis was to form cysts, and their elevated expression in response to colchicine treatment (arresting cells in the G2/M transition) indicated a potential role of these genes in cell division and population growth in H. akashiwo. Consistent with this, we also found a morning to afternoon shift in the expression of these genes during the bloom of H. akashiwo. This study documents a part of the typical SR gene repertoire and its potential role in regulating cell division in H. akashiwo, offering candidates for population growth markers for bloom monitoring although its linkage to meiosis and SR remain to be studied further in the future.

Massive cyanobacteria blooms occur almost every summer in the Baltic Sea but the capability to quantitatively predict their extent and intensity is poorly developed. Here we analyse statistical relationships between multi-decadal satellite-derived time series of the frequency of cyanobacteria surface accumulations (FCA) in the central Baltic Sea Proper and a suite of environmental variables. Over the decadal scale (∼5-20 years) FCA was highly correlated (R2 ∼ 0.69) with a set of biogeochemical variables related to the amount of phosphorus and hypoxia in bottom layers. Water temperature in the surface layer was also positively correlated with FCA at the decadal scale. In contrast, the inter-annual variations in FCA had no correlation with the biogeochemical variables. Instead, significant correlations were found with the solar shortwave direct flux in July and the sea-surface temperature, also in July. It thus appears that it is not possible to predict inter-annual fluctuations in cyanobacteria blooms from water chemistry. Moreover, environmental variables could only explain about 45% of the inter-annual variability in FCA, probably because year-to-year variations in FCA are significantly influenced by biological interactions.

Predicting algal population dynamics using models informed by experimental data has been used as a strategy to inform the management and control of harmful cyanobacterial blooms. We selected toxic bloom-forming species Microcystis spp. and Raphidiopsis raciborskii (basionym Cylindrospermopsis raciborskii) for further examination as they dominate in 78 % and 17 %, respectively, of freshwater cyanobacterial blooms (cyanoHABs) reported globally over the past 30 years. Field measurements of cyanoHABs are typically based on biomass accumulation, but laboratory experiments typically measure growth rates, which are an important variable in cyanoHAB models. Our objective was to determine the usefulness of laboratory studies of these cyanoHAB growth rates for simulating the species dominance at a global scale. We synthesized growth responses of M. aeruginosa and R. raciborskii from 20 and 16 culture studies, respectively, to predict growth rates as a function of two environmental variables, light and temperature. Predicted growth rates of R. raciborskii exceeded those of M. aeruginosa at temperatures ≳ 25 °C and light intensities ≳ 150 μmol photons m−2 s-1. Field observations of biomass accumulation, however, show that M. aeruginosa dominates over R. raciborskii, irrespective of climatic zones. The mismatch between biomass accumulation measured in the field, and what is predicted from growth rate measured in the laboratory, hinders effective use of culture studies to predict formation of cyanoHABs in the natural environment. The usefulness of growth rates measured may therefore be limited, and field experiments should instead be designed to examine key physiological attributes such as colony formation, buoyancy regulation and photoadaptation. Improving prediction of cyanoHABs in a changing climate requires a more effective integration of field and laboratory approaches, and an explicit consideration of strain-level variability.

Climate change is transforming aquatic ecosystems. Coastal waters have experienced progressive warming, acidification, and deoxygenation that will intensify this century. At the same time, there is a scientific consensus that the public health, recreation, tourism, fishery, aquaculture, and ecosystem impacts from harmful algal blooms (HABs) have all increased over the past several decades. The extent to which climate change is intensifying these HABs is not fully clear, but there has been a wealth of research on this topic this century alone. Indeed, the United Nations' Intergovernmental Panel on Climate Change's (IPCC) Special Report on the Ocean and Cryosphere in a Changing Climate (SROCC) approved in September 2019 was the first IPCC report to directly link HABs to climate change. In the Summary for Policy Makers, the report made the following declarations with “high confidence”: • Harmful algal blooms display range expansion and increased frequency in coastal areas since the 1980s in response to both climatic and non-climatic drivers such as increased riverine nutrients run-off. • The observed trends in harmful algal blooms are attributed partly to the effects of ocean warming, marine heatwaves, oxygen loss, eutrophication and pollution. • Harmful algal blooms have had negative impacts on food security, tourism, local economy, and human health. In addition, the report specifically outlines a series of linkages between heat waves and HABs. These statements about HABs and climate change and the high levels of confidence ascribed to them provides clear evidence that the field of HABs and climate change has matured and has, perhaps, reached a first plateau of certainty. While there are well-documented global trends in HABs being promoted by human activity, including climate change, individual events are driven by local, regional, and global drivers, making it critical to carefully evaluate the conditions and responses at appropriate scales. It is within this context that the first Special Issue on Climate Change and Harmful Algal Blooms is published in Harmful Algae.

Elevated seawater temperatures are linked to the development of harmful algal blooms (HABs), which pose a growing threat to marine birds and other wildlife. During late 2015 and early 2016, a massive die-off of Common Murres (Uria aalge; hereafter, murres) was observed in the Gulf of Alaska coincident with a strong marine heat wave. Previous studies have documented illness and death among seabirds resulting from exposure to the HAB neurotoxins saxitoxin (STX) and domoic acid (DA). Given the unusual mortality event, corresponding warm water anomalies, and recent detection of STX and DA throughout coastal Alaskan waters, HABs were identified as a possible factor of concern. To evaluate whether algal toxins may have contributed to murre deaths, we tested for STX and DA in a suite of tissues obtained from beach-cast murre carcasses associated with the die-off as well as from apparently healthy murres and Black-legged Kittiwakes (Rissa tridactyla; hereafter, kittiwakes) sampled in the preceding and following summers. We also tested forage fish and marine invertebrates collected in the Gulf of Alaska in 2015–2017 to evaluate potential sources of HAB toxin exposure for seabirds. Saxitoxin was present in multiple tissue types of both die-off (36.4 %) and healthy (41.7 %) murres and healthy kittiwakes (54.2 %). Among birds, we detected the highest concentrations of STX in liver tissues (range 1.4–10.8 μg 100 g−1) of die-off murres. Saxitoxin was relatively common in forage fish (20.3 %) and invertebrates (53.8 %). No established toxicity limits currently exist for seabirds, but concentrations of STX in birds and forage fish in our study were lower than values reported from most other bird die-offs in which STX intoxication was causally linked. We detected low concentrations of DA in a single bird sample and in 33.3 % of invertebrates and 4.0 % of forage fish samples. Although these results do not support the hypothesis that acute exposure to STX or DA was a primary factor in the 2015–2016 mortality event, additional information about the sensitivity of murres to these toxins is needed before we can discount their potential role in the die-off. The widespread occurrence of STX in seabirds, forage fish, and invertebrates in the Gulf of Alaska indicates that algal toxins should be considered in future assessments of seabird health, especially given the potential for greater occurrence of HABs in the future.

Many phytoplankton species, including many harmful algal bloom (HAB) species, survive long periods between blooms through formation of benthic resting stages. Because they are crucial to the persistence of these species and the initiation of new blooms, the physiology of benthic stages must be considered to accurately predict responses to climate warming and associated environmental changes. The benthic stages of dinoflagellates, called resting cysts, germinate in response to the combination of favorable temperature, oxygen-availability, and release from dormancy. The latter is a mechanism that prevents germination even when oxygen and temperature conditions are favorable. Here, evidence of temperature-mediated control of dormancy duration from the dinoflagellates Alexandrium catenella and Pyrodinium bahamense—two HAB species that cause paralytic shellfish poisoning (PSP)—is reviewed and presented alongside new evidence of complementary, temperature-based control of cyst quiescence (the state in which cysts germinate on exposure to favorable conditions). Interaction of the two temperature-based mechanisms with climate is explored through a simple model parameterized using results from recent experiments with A. catenella. Simulations demonstrate the importance of seasonal temperature cycles for the synchronization of cysts’ release from dormancy and are consistent with biogeography-based inferences that A. catenella is more tolerant of warming in habitats that experience a larger range of seasonal temperature variation (i.e., have higher temperature seasonality). Temperature seasonality is much greater in shallow, long-residence time habitats than in deep, open-water ones. As warming shifts species’ ranges, cyst beds may persist longer in more seasonally variable, shallow inshore habitats than in deep offshore ones, promoting HABs that are more localized and commence earlier each year. Recent field investigations of A. catenella also point to the importance of new cyst formation as a factor triggering bloom termination through mass sexual induction. In areas where temperature seasonality restricts the flux of new swimming cells (germlings) to narrow temporal windows, warming is unlikely to promote longer and more intense HAB impacts—even when water column conditions would otherwise promote prolonged bloom development. Many species likely have a strong drive to sexually differentiate and produce new cysts once concentrations reach levels that are conducive to new cyst formation. This phenomenon can impose a limit to bloom intensification and suggests an important role for cyst bed quiescence in determining the duration of HAB risk periods.

This review assesses harmful algal bloom (HAB) modeling in the context of climate change, examining modeling methodologies that are currently being used, approaches for representing climate processes, and time scales of HAB model projections. Statistical models are most commonly used for near-term HAB forecasting and resource management, but statistical models are not well suited for longer-term projections as forcing conditions diverge from past observations. Process-based models are more complex, difficult to parameterize, and require extensive calibration, but can mechanistically project HAB response under changing forcing conditions. Nevertheless, process-based models remain prone to failure if key processes emerge with climate change that were not identified in model development based on historical observations. We review recent studies on modeling HABs and their response to climate change, and the various statistical and process-based approaches used to link global climate model projections and potential HAB response. We also make several recommendations for how the field can move forward: 1) use process-based models to explicitly represent key physical and biological factors in HAB development, including evaluating HAB response to climate change in the context of the broader ecosystem; 2) quantify and convey model uncertainty using ensemble approaches and scenario planning; 3) use robust approaches to downscale global climate model results to the coastal regions that are most impacted by HABs; and 4) evaluate HAB models with long-term observations, which are critical for assessing long-term trends associated with climate change and far too limited in extent.

A ubiquitous dinoflagellate, Alexandrium, produces paralytic shellfish toxin (PST), and its outbreaks have negative impacts on aquaculture, fisheries, human health, and the marine ecosystem. To minimize such damages, a routine monitoring program of toxic species must be implemented with a suitable analytical technique for their identification and quantification. However, the taxonomic identification and cell quantification of Alexandrium species based on their external morphology under a light microscope, or by using conventional molecular approaches have limited sensitivity and reproducibility. To address these challenges, we have developed an advanced protocol using droplet-digital PCR (ddPCR) for the discrimination and enumeration of three co-occurring Alexandrium species (A. affine, A. catenella, and A. pacificum) in environmental samples. Copies of species-specific internal transcribed spacer (ITS) per cell, which were calculated from environmental samples spiked with various numbers of culture cells, were used to estimate the abundance of species in the field samples. There were no significant differences in ITS copies estimated by the digital PCR assay between environmental samples from different localities, spiked artificially with a consistent number of cells from Alexandrium cultures. This sensitive assay was applied to determine the abundance and vertical distribution of those populations in the southern coastal waters of Korea. In spring, A. catenella was the dominant species, followed by the non-toxic A. affine in summers. A novel digital PCR assay can also be used to monitor other harmful marine protists that require high sample throughput and low detection limit with high accuracy and precision.

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

Cyanobacteria are notorious for producing water blooms and for toxin formation. Toxic cyanobacterial blooms present an ever-increasing serious threat to both the quality of drinking water and recreational uses and severely disrupt aquatic ecosystems, worldwide. In many cases, such blooms are dominated by toxic Microcystis sp. that produce a family of structurally similar hepatotoxins, known as microcystins (MCs). Here we present a retrospective analysis of Microcystis seasonal blooms from Lake Kinneret (Sea of Galilee, Israel) indicating that the population is composed of at least 25 different genotypes and two different chemo-types, whose relative abundance changes over decades. Based on a long-term record of biotic and abiotic parameters and laboratory experiments we propose that minor increase in water temperature, but not in salinity, may affect Microcystis community structure by changing the relative abundance of species/strains from toxic to less or non-toxic species.

In autumn of 2013 an immense dinoflagellate bloom developed in Kachemak Bay, AK, USA. Much of the Bay was discolored a dark amber color and raised public concerns as small scale fish kills were reported in a few locations. Light microscopy revealed a monospecific bloom of gymnodinoid dinoflagellates that were previously unknown from the Bay. Gene sequencing of SSU rDNA from cells collected from the bloom confirmed the causative species to be Karenia mikimotoi. This represents the first report of a K. mikimotoi bloom in Alaska. After the bloom organism was confirmed, a K. mikimotoi species-specific qPCR assay was developed and used to assess K. mikimotoi abundances in DNA extracted from phytoplankton samples from Kachemak Bay and Lower Cook Inlet (LCI) obtained over a six-year period. The K. mikimotoi abundances were compared with corresponding time series of environmental variables (water temperature, salinity, water column stability, nutrients, precipitation and wind speed) to assess the factors contributing to the development of the bloom. The results showed early bloom development occurred in August when snow melt reduced salinities and increased water column stability during a period of calm winds. Peak bloom concentrations occurred in late September (107 cell eq. L-1) even as water temperatures were decreasing. The bloom gradually declined over the winter but persisted until April of 2014. Karenia mikimotoi cells were not detected two years prior or three years following the bloom, suggesting cells were introduced to Kachemak Bay at a time when conditions allowed K. mikimotoi to thrive.

Monitoring drinking water quality is an important public health issue and pathogenic organisms present a particularly serious health hazard in freshwater bodies. However, many pathogenic bacteria, including cyanobacteria, and pathogenic protozoa can be swept into coastal lagoons and into near-shore marine environments where they continue to grow and pose a health threat to marine mammals and invertebrates. In this study, we tested the suitability of a phylochip (microarray for species detection) developed for freshwater pathogenic organisms to be applied to samples taken across a marine/freshwater interface at monthly intervals for two years. Toxic cyanobacteria and pathogenic protozoa were more numerous in a coastal lagoon than at the freshwater or marine site, indicating that this microarray can be used to detect the presence of these pathogens across a marine/freshwater interface and thus the potential for toxicity to occur within the entire watershed.

San Francisco Bay (SFB), California, USA is the largest estuary in the western United States and is home to more than 7 million people in nine counties and 101 cities. It is highly nutrient enriched and is directly connected to the Gulf of the Farallones and coastal Pacific ocean through the Golden Gate strait. The Gulf of the Farallones is one of several “hotspots” for the neurotoxin domoic acid, produced by members of the genus Pseudo-nitzschia. Despite the close proximity, SFB has few reports of harmful algal blooms and low concentrations of domoic acid, suggesting that SFB is somehow resistant to toxic blooms. Here we evaluate the potential growth and toxicity of the dominant toxigenic species in California coastal waters, P. australis and P. multiseries, to directly test the hypothesis that SFB waters confer resistance to blooms. We specifically evaluate the effect of varying temperature, salinity, and to a lesser extent, nutrients on growth and toxin production. Results show equivalent growth in SFB water (maximum growth rates of 0.71 and 1.35 d−1 for P. multiseries and P. australis) compared to open-coast water, and comparable or greater toxicity (0 to >100 pg DA cell−1). The historical resistance to blooms in SFB is hypothesized to be caused by a combination of insufficient acclimation time for advected Pseudo-nitzschia populations to become established and suppression of toxin production in warm waters.

In the present study, the abundance of Prorocentrum and the molecular phylogeny, distribution, and DST production of P. lima complex and P. caipirignum in Japan were investigated. First, the cell densities of Prorocentrum were assessed from the temperate to subtropical zones in Japan between 2014 and 2018. The cell density in the subtropical zone [19.0 ± 40.2 cells/g wet weight (ww) algae] was significantly higher than that in the temperate zone (1.4 ± 3.4 cells/g ww algae). A total of 244 clonal strains were established from the temperate and subtropical zones. Phylogenetic analyses based on the large-subunit ribosomal DNA D1/D2 revealed that the strains were separated into four species/species complex/phylotypes (P. limacomplex, P. caipirignum, and new phylotypes Prorocentrum spp. types 1 and 2). The strains of P. lima complex could be separated into two clades (1 and 3). Furthermore, the strains of clades 1 and 3 could be separated into nine subclades (1a, 1c, 1d, 1e, 1f, 1 g, 1 h, 1i, and 1 j) and three subclades (3a, 3b, and 3c), respectively. The strains of P. caipirignum were separated into two subclades (b and e). Each phylotype/subclade showed a unique distribution pattern in Japan: P. lima complex subclades 1a, 1c, and 3a and P. caipirignum subclades b and e were widespread from the temperate to subtropical zones. On the other hand, P. lima complex subclades 1e and 1i were restricted to the temperate zone, and P. lima complex subclades 1d, 1f, 1 g, 1 h, 1 j, 3b, and 3c and Prorocentrum spp. types 1 and 2 were restricted to the subtropical zone. Furthermore, the DST production of the 243 clonal strains was assessed by LC/MS/MS analysis. The results revealed that all strains produced okadaic acid (OA) and that the OA contents of P. lima complex subclades 1d and 1f, P. caipirignum subclades b and e, and Prorocentrum sp. type 2 tended to be higher than those of the other subclades. While P. lima complex subclades 1a, 1e, 1f, and 1i produced DTX1, the other phylotype/subclades produced either no or low quantities of DTX1. A strain of P. lima complex subclade 1e showed the highest OA and DTX1 contents (55.27 and 70.73 pg/cell, respectively) in the world. These results suggest that there are potential risks for DST accumulation in benthic animals in Japan.

Despite nearly annual blooms of the neurotoxic dinoflagellate Karenia brevis (Davis) G. Hansen and Moestrup in the Gulf of Mexico, defining the suite of biological traits that explain its proliferation has remained challenging. Studies have described K. brevis as a low-light-adapted species, incapable of sustaining growth under high light, which is at odds with observed surface aggregations sometimes within centimeters of the sea surface and also with short-term experiments showing photosynthetic machinery accommodating high irradiances. Here, growth and photophysiology of three K. brevis isolates were evaluated under a range of environmentally relevant irradiances (10–1500 μmol photons m−2 s−1) in the laboratory. No differences in growth–irradiance curves were observed among isolates; all sustained maximum growth rates at the highest irradiances examined, even in exposures as long as three weeks. The growth efficiency α of K. brevis under light-limiting conditions appeared mediocre among dinoflagellates, and poorer than that of other phytoplankton (e.g., diatoms, cyanobacteria), implying that K. brevis is not a low-light specialist. This finding substantially alters earlier parameterizations of K. brevis growth–irradiance curves. Therefore, a model was developed to contextualize how these new growth–irradiance curves might affect bottom growth rates. This model was subsequently applied to a case study comparing seasonal light forcing offshore of Pinellas County, FL, USA, with a single empirical value for light attenuation, and seasonal bottom water temperatures. Predictions suggested that light may limit bottom growth as close as 1 km from shore in winter, but would only begin limiting growth 20 km from shore in summer. Population maintenance (no net growth) was possible as far offshore as 90 km in summer and 68 km in winter. These ranges intercept areas thought to be important for bloom initiation.

Global increases in atmospheric CO2 and temperatures will impact aquatic systems, with freshwater habitats being affected. Some studies suggest that these conditions will promote cyanobacterial dominance. There is a need for a clearer picture of how algal species and strains within species will respond to higher temperatures and CO2, especially in combination. This study examined two chlorophytes (Monoraphidium and Staurastrum), and two strains of the cyanobacterium Raphidiopsis raciborskii (straight S07 and coiled C03), to determine how the combination of higher temperature and CO2 levels will affect their growth and maximum cell concentrations. Continuous cultures were used to compare the steady state cell concentrations at 28 °C and 30 °C, and CO2 partial pressures (pCO2), 400 and 750 ppm for all cultures, and in addition 1000 ppm at 28 °C for R. raciborskii strains. This study showed that, for all species, water temperature had a greater effect than higher pCO2 on cell concentrations. There were clear differences in response between the chlorophyte species, with Monoraphidium preferring 28 °C and Staurastrum preferring 30 °C. There were also differences in response of the R. raciborskii strains to increasing temperature and pCO2, with S07 having a greater increase in cell concentration. Genome analysis of R. raciborskii showed that the straight strain has five additional carbon acquisition genes (β-CA, chpY, cmpB, cmpD and NdhD4), indicative of increased carbon metabolism. These differences in the strains’ response to elevated pCO2 will lead to changes in the species population structure and distribution in the water column. This study shows that it is important to examine the effects of both pCO2 and temperature, and to consider strain variation, to understand how species composition of natural systems may change under future climate conditions.

Within the past few decades, harmful algal blooms (HABs) have occurred frequently in Indonesian waters, resulting in environmental degradation, economic loss and human health problems. So far, HAB related studies mainly addressed ecological traits and species distribution, yet toxin measurements were virtually absent for Indonesian waters. The aim of the present study was to explore variability of the potentially toxic marine diatom genus Pseudo-nitzschia, as well as its neurotoxin domoic acid as a function of environmental conditions in Ambon Bay, eastern Indonesia. Weekly phytoplankton samples, oceanographic (CTD, nutrients) and meteorological (precipitation, wind) parameters were analyzed at 5 stations in the bay during the dry and wet seasons of 2018. Liquid chromatography – tandem mass spectrometry (LC–MS/MS) was used to detect particulate DA (pDA). Vegetative cells of Pseudo-nitzschia spp. and pDA were found in 98.6% and 51.4% of the samples, respectively. pDA levels were low, yet detected throughout the campaign, implying that Ambon Bay might potentially be subject to amnesic shellfish poisoning. The highest levels of both Pseudo-nitzschia spp. cell abundance and pDA were found in the wet season, showing a strong positive correlation between both parameters, compared to the dry season, (r = 0.87 and r = 0.66 (p < 0.01), respectively). Statistical analyses revealed that temperature and mixed layer depth positively correlated with Pseudo-nitzschia spp. and pDA during the dry season, while ammonium showed positive correlations in both seasons. This study represents the first successful investigation of the presence and variability of Pseudo-nitzschia spp. and its neurotoxin DA in Indonesian waters.

Enrichment of the oceans with CO2 may be beneficial for some marine phytoplankton, including harmful algae. Numerous laboratory experiments provided valuable insights into the effects of elevated pCO2 on the growth and physiology of harmful algal species, including the production of phycotoxins. Experiments close to natural conditions are the next step to improve predictions, as they consider the complex interplay between biotic and abiotic factors that can confound the direct effects of ocean acidification. We therefore investigated the effect of ocean acidification on the occurrence and abundance of phycotoxins in bulk plankton samples during a long-term mesocosm experiment in the Gullmar Fjord, Sweden, an area frequently experiencing harmful algal blooms. During the experimental period, a total of seven phycotoxin-producing harmful algal genera were identified in the fjord, and in accordance, six toxin classes were detected. However, within the mesocosms, only domoic acid and the corresponding producer Pseudo-nitzschia spp. was observed. Despite high variation within treatments, significantly higher particulate domoic acid contents were measured in the mesocosms with elevated pCO2. Higher particulate domoic acid contents were additionally associated with macronutrient limitation. The risks associated with potentially higher phycotoxin levels in the future ocean warrants attention and should be considered in prospective monitoring strategies for coastal marine waters.

Harmful events associated with epibenthic dinoflagellates, have been reported more frequently over the last decades. Occurrence of potentially toxic benthic dinoflagellates, on the leaves of two magnoliophytes (Cymodocea nodosa and Zostera noltei) and thalli of the macroalgae (Ulva rigida), was monitored over one year (From May 2015 to April 2016) in the Bizerte Bay and Lagoon (North of Tunisia, Southern Mediterranean Sea). The investigated lagoon is known to be highly anthropized. This is the first report on the seasonal distribution of epibenthic dinoflagellates hosted by natural substrates, from two contrasted, adjacent coastal Mediterranean ecosystems. The environmental factors promoting the development of the harmful epibenthic dinoflagellates Ostreopsis spp., Prorocentrum lima and Coolia monotis were investigated. The highest cell densities were reached by Ostreopsis spp. (1.9 × 103 cells g−1 FW, in October 2015), P. lima (1.6 × 103 cells g−1 FW, in June 2015) and C. monotis (1.1 × 103 cells g−1 FW, in May 2015). C. nodosa and Z. noltei were the most favorable host macrophytes for C. monotis (in station L2) and Ostreopsis spp. (in station L3), respectively. Positive correlations were recorded between Ostreopsis spp. and temperature. Densities of the epibenthic dinoflagellates varied according to the collection site, and a great disparity was observed between the Bay and the Lagoon. Maximum concentrations were recorded on C. nodosa leaves from the Bizerte Bay, while low epiphytic cell abundances were associated with macrophytes sampled from the Bizerte Lagoon. The observed differences in dinoflagellate abundances between the two ecosystems (Bay-Lagoon) seemed not related to the nutrients, but rather to the poor environmental conditions in the lagoon.

Mass spectrometric methods for the quantitative and qualitative analyses of algal biotoxins are often complicated by co-eluting compounds that present analytically as interferences. This issue is particularly critical for organic polyamines, where co-eluting materials can suppress the formation of cations during electrospray ionization. Here we present an extraction procedure designed specifically to overcome matrix-derived ion suppression of algal toxins in samples of Lyngbya wollei, a filamentous benthic algae known to produce several saxitoxin analogues. Lyngbya wollei samples were collected from a large, persistent harmful algal bloom in Lake Wateree, SC. Six known Lyngbya wollei-specific toxins (LWT1–6) were successfully resolved and quantified against saxitoxin using hydrophilic interaction liquid chromatography coupled with triple quadrupole and quadrupole time-of-flight mass spectrometry. The parent ions [M2+ – H+]+ were observed for LWTs 1–6 and the [M]2+ ion was observed for LWT5. High resolution mass spectra and unique fragmentation ions were obtained for LWTs 1–6. A dilution factor of 50 resulted in a linear calibration of saxitoxin in the algae matrix. Ion suppression was resolved by sample dilution, which led to linear, positive correlations between peak area and mass of the extracted sample (R2 > 0.96). Optimized sample extraction method and instrument parameters are presented.

Karenia mikimotoi is a worldwide bloom-forming dinoflagellate in the genus Karenia. Blooms of this alga have been observed since the 1930s and have caused mass mortalities of fish, shellfish, and other invertebrates in the coastal waters of many countries, including Japan, Norway, Ireland, and New Zealand. This species has frequently bloomed in China, causing great financial losses (more than 2 billion yuan, Fujian Province, 2012). K. mikimotoi can adapt to various light, temperature, salinity, and nutrient conditions, which together with its complex life history, strong motility, and density-dependent allelopathy, allows it to form blooms that are lethal to almost all marine organisms. However, its toxicity differs between subspecies and some target-species-specific toxicity has also been recorded. Significant gill disorder is observed in affected fish, to which the massive fish kills are attributed, rather than to the hypoxia that occurs in the fading stage of a bloom. However, although this species is haemolytic and cytotoxic, and generates reactive oxygen species, none of the isolated toxins or lipophilic extracts have toxic effects as extreme as those of the intact algal cells. The toxic effects of K. mikimotoi are strongly related to contact with intact cells. Several reasonable hypotheses of how and why this species blooms and causes mass mortalities have been proposed, but further research is required.

The Benguela Current in the Atlantic is one of the four major upwelling systems on the Eastern boundary of the world ocean. Thus the coastal regions off Namibia are prone to high primary productivity that can lead to Harmful Algae Blooms as this nutrient rich water reaches the euphotic zone. Yessotoxins (YTXs) produced by G. spinifera were detected in Namibian phytoplankton field samples in 2011. Isolation of G. spinifera cultures from this location in 2012 enabled molecular genetics work and further liquid chromatography-mass spectrometry assessment of toxin profiles. The molecular work grouped the Benguela G. spinifera with other toxic G. spinifera strains originating from Italy and New Zealand. The main YTX analogs present in the Benguela G. spinifera are homo-YTX, YTX and a hydroxylated analogue. This work adds important knowledge on the occurrence of Harmful Algae Blooms in this region and is of relevance for safety.

Climate projections suggest–with substantial certainty–that global warming >1.5 °C will occur by mid-century (2050). Population is also projected to increase, amplifying the demands for food, fuel, water and sanitation, which, in turn, escalate nutrient pollution. Global projections of nutrient pollution, however, are less certain than those of climate as there are regionally decreasing trends projected in Europe, and stabilization of nutrient use in North America and Australia. In this review of the effects of eutrophication and climate on harmful algae, some of the complex, subtle, and non-intuitive effects and interactions on the physiology of both harmful and non-harmful taxa are emphasized. In a future ocean, non-harmful diatoms may be disproportionately stressed and mixotrophs advantaged due to changing nutrient stoichiometry and forms of nutrients, temperature, stratification and oceanic pH. Modeling is advancing, but there is much yet to be understood, in terms of physiology, biogeochemistry and trophodynamics and how both harmful and nonharmful taxa may change in an uncertain future driven by anthropogenic activities.

Climate change is predicted to increase the severity and prevalence of harmful algal blooms (HABs). In the past twenty years, omics techniques such as genomics, transcriptomics, proteomics and metabolomics have transformed that data landscape of many fields including the study of HABs. Advances in technology have facilitated the creation of many publicly available omics datasets that are complementary and shed new light on the mechanisms of HAB formation and toxin production. Genomics have been used to reveal differences in toxicity and nutritional requirements, while transcriptomics and proteomics have been used to explore HAB species responses to environmental stressors, and metabolomics can reveal mechanisms of allelopathy and toxicity. In this review, we explore how omics data may be leveraged to improve predictions of how climate change will impact HAB dynamics. We also highlight important gaps in our knowledge of HAB prediction, which include swimming behaviors, microbial interactions and evolution that can be addressed by future studies with omics tools. Lastly, we discuss approaches to incorporate current omics datasets into predictive numerical models that may enhance HAB prediction in a changing world. With the ever-increasing omics databases, leveraging these data for understanding climate-driven HAB dynamics will be increasingly powerful.

Marine and freshwater ecosystems are warming, acidifying, and deoxygenating as a consequence of climate change. In parallel, the impacts of harmful algal blooms (HABs) on these ecosystems are intensifying. Many eutrophic habitats that host recurring HABs already experience thermal extremes, low dissolved oxygen, and low pH, making these locations potential sentinel sites for conditions that will become more common in larger-scale systems as climate change accelerates. While studies of the effects of HABs or individual climate change stressors on aquatic organisms have been relatively common, studies assessing their combined impacts have been rare. Those doing so have reported strong species- and strain-specific interactions between HAB species and climate change co-stressors yielding outcomes for aquatic organisms that could not have been predicted based on investigations of these factors individually. This review provides an ecological and physiological framework for considering HABs as a climate change co-stressor and considers the consequences of their combined occurrence for coastal ecosystems. This review also highlights critical gaps in our understanding of HABs as a climate change co-stressor that must be addressed in order to develop management plans that adequately protect fisheries, aquaculture, aquatic ecosystems, and human health. Ultimately, incorporating HAB species into experiments and monitoring programs where the effects of multiple climate change stressors are considered will provide a more ecologically relevant perspective of the structure and function of marine ecosystems in future, climate-altered systems.

Harmful cyanobacterial blooms (=cyanoHABs) are an increasing feature of many waterbodies throughout the world. Many bloom-forming species produce toxins, making them of particular concern for drinking water supplies, recreation and fisheries in waterbodies along the freshwater to marine continuum. Global changes resulting from human impacts, such as climate change, over-enrichment and hydrological alterations of waterways, are major drivers of cyanoHAB proliferation and persistence. This review advocates that to better predict and manage cyanoHABs in a changing world, researchers need to leverage studies undertaken to date, but adopt a more complex and definitive suite of experiments, observations, and models which can effectively capture the temporal scales of processes driven by eutrophication and a changing climate. Better integration of laboratory culture and field experiments, as well as whole system and multiple-system studies are needed to improve confidence in models predicting impacts of climate change and anthropogenic over-enrichment and hydrological modifications. Recent studies examining adaptation of species and strains to long-term perturbations, e.g. temperature and carbon dioxide (CO2) levels, as well as incorporating multi-species and multi-stressor approaches emphasize the limitations of approaches focused on single stressors and individual species. There are also emerging species of concern, such as toxic benthic cyanobacteria, for which the effects of global change are less well understood, and require more detailed study. This review provides approaches and examples of studies tackling the challenging issue of understanding how global changes will affect cyanoHABs, and identifies critical information needs for effective prediction and management.

Rising concentrations of atmospheric CO2 results in higher equilibrium concentrations of dissolved CO2 in natural waters, with corresponding increases in hydrogen ion and bicarbonate concentrations and decreases in hydroxyl ion and carbonate concentrations. Superimposed on these climate change effects is the dynamic nature of carbon cycling in coastal zones, which can lead to seasonal and diel changes in pH and CO2 concentrations that can exceed changes expected for open ocean ecosystems by the end of the century. Among harmful algae, i.e. some species and/or strains of Cyanobacteria, Dinophyceae, Prymnesiophyceae, Bacillariophyceae, and Ulvophyceae, the occurrence of a CO2 concentrating mechanisms (CCMs) is the most frequent mechanism of inorganic carbon acquisition in natural waters in equilibrium with the present atmosphere (400 μmol CO2 mol−1 total gas), with varying phenotypic modification of the CCM. No data on CCMs are available for Raphidophyceae or the brown tide Pelagophyceae. Several HAB species and/or strains respond to increased CO2 concentrations with increases in growth rate and/or cellular toxin content, however, others are unaffected. Beyond the effects of altered C concentrations and speciation on HABs, changes in pH in natural waters are likely to have profound effects on algal physiology. This review outlines the implications of changes in inorganic cycling for HABs in coastal zones, and reviews the knowns and unknowns with regard to how HABs can be expected to ocean acidification. We further point to the large regions of uncertainty with regard to this evolving field.

Red tide and green tide are two common algal blooms that frequently occur in many areas in the global oceans. The algae causing red tide and green tide often interact with each other in costal ecosystems. However, little is known on how future CO2-induced ocean acidification combined with temperature variation would affect the interaction of red and green tides. In this study, we cultured the red tide alga Skeletonema costatum and the green tide alga Ulva linza under ambient (400 ppm) and future CO2 (1000 ppm) levels and three temperatures (12, 18, 24 °C) in both monoculture and coculture systems. Coculture did not affect the growth rate of U. linza but significantly decreased it for S. costatum. Elevated CO2 relieved the inhibitory effect of U. linza on the growth of S. costatum, particularly for higher temperatures. At elevated CO2, higher temperature increased the growth rate of S. costatum but reduced it for U. linza. Coculture with U. linza reduced the net photosynthetic rate of S. costatum, which was relieved by elevated CO2. This pattern was also found in Chl a content, indicating that U. linza may inhibit growth of S. costatum via harming pigment synthesis and thus photosynthesis. In monoculture, higher temperature did not affect respiration rate of S. costatum but increased it in U. linza. Coculture did not affect respiration of U. linza but stimulated it for S. costatum, which was a signal of responding to biotic and/abiotic stress. The increased growth of S. costatum at higher temperature and decreased inhibition of U. linza on S. costatum at elevated CO2 suggest that red tides may have more advantages over green tides in future warmer and CO2-enriched oceans.

A tourism dependent state such as Florida relies on its environment and climate to attract visitors and generate revenue. HABs can certainly have an impact on the coastal waters of the Gulf, but does this necessarily drive away tourist related activity? To determine not only if the impact of HABs is significant, but also at what magnitude, a time series econometric model was used to study effects of persistent and severe blooms on counties in Southwestern Florida, particularly Sarasota County, hit hardest by blooms in 2006 and 2018 that lasted multiple months. Lodging and restaurant sectors of the economy were found to have monthly losses of 15% and 1.75% respectively, during months when red tide was present. Neighboring counties unaffected by severe blooms did not experience significant losses to these sectors. These results support the intuition that effects of HABs reach far beyond the waters of the Gulf, and as red tide grows in frequency and severity, more economic loss could lie ahead.

Cyanophage MaMV-DC is a member of Myoviridae that was reported to specifically infect and lyse Microcystis aeruginosa FACHB-524 among 21 selected cyanobacterial strains. We reidentified the infection specificity of MaMV-DC among seven other Microcystis strains of different species. In our experiments, MaMV-DC infected three Microcystis strains but did not form plaque in Microcystis lawns. This indicated that MaMV-DC is at least a genus- rather than strain-specific virus. Cyanophage MaMV-DC genes were transcribed in M. aeruginosa FACHB-524, M. flos-aquae TF09, M. aeruginosa TA09 and M. wesenbergii DW09, and the growth of these Microcystis strains was inhibited by the addition of MaMV-DC. The predicted defense of eight Microcystis strains by CRISPR–Cas systems has shown mixed consistency with the infection experiment results, suggesting other defense or anti-defense systems play roles during infection process. Restriction–modification (RM) system analysis revealed an abundance of four types of RM proteins that may play roles in defense against cyanophages.

Oxygen-based productivity and respiration rates were determined in West Florida coastal waters to evaluate the proportion of community respiration demands met by autotrophic production within a harmful algal bloom dominated by Karenia brevis. The field program was adaptive in that sampling during the 2006 bloom occurred where surveys by the Florida Wildlife Research Institute indicated locations with high cell abundances. Net community production (NCP) rates from light-dark bottle incubations during the bloom ranged from 10 to 42 µmole O2 L-1 day-1 with highest rates in bloom waters where abundances exceeded 105 cells L-1. Community dark respiration (R) rates in dark bottles ranged from <10 to 70 µmole O2 L-1 day-1 over 24 h. Gross primary production derived from the sum of NCP and R varied from ca. 20 to 120 µmole O2 L-1 day-1. The proportion of GPP attributed to NCP varied with the magnitude of R during day and night periods. Most surface communities exhibited net autotrophic production (NCP > R) over 24 h, although heterotrophy (NCP < R) characterized the densest sample where K. brevis cell densities exceed 106 cells L-1.

Cyanobacterial harmful algal blooms (CyanoHAB) are thought to be increasing globally over the past few decades, but relatively little quantitative information is available about the spatial extent of blooms. Satellite remote sensing provides a potential technology for identifying cyanoHABs in multiple water bodies and across geo-political boundaries. An assessment method was developed using MEdium Resolution Imaging Spectrometer (MERIS) imagery to quantify cyanoHAB surface area extent, transferable to different spatial areas, in Florida, Ohio, and California for the test period of 2008 to 2012. Temporal assessment was used to evaluate changes in satellite resolvable inland waterbodies for each state of interest. To further assess cyanoHAB risk within the states, the World Health Organization's (WHO) recreational guidance level thresholds were used to categorize surface area of cyanoHABs into three risk categories: low, moderate, and high-risk bloom area. Results showed that in Florida, the area of cyanoHABs increased largely due to observed increases in high-risk bloom area. California exhibited a slight decrease in cyanoHAB extent, primarily attributed to decreases in Northern California. In Ohio (excluding Lake Erie), little change in cyanoHAB surface area was observed. This study uses satellite remote sensing to quantify changes in inland cyanoHAB surface area across numerous water bodies within an entire state. The temporal assessment method developed here will be relevant into the future as it is transferable to the Ocean Land Colour Instrument (OLCI) on Sentinel-3A/3B missions.

Inshore and offshore waters of the Gulf of Maine (USA) have spring/summer harmful algal blooms (HABs) of the toxic dinoflagellate Alexandrium fundyense, which is responsible for paralytic shellfish poisoning (PSP) in humans. The calanoid copepod Calanus finmarchicus co-occurs with A. fundyense during the seasonal blooms. At that time, C. finmarchicus population abundances are high, dominated by immature copepods preparing for diapause, and by actively-reproducing adults. High survival has been reported for copepods exposed to toxic A. fundyense, but little is known about possible sublethal effects. In this study, C. finmarchicus adult females were fed either a control diet of non-toxic Rhodomonas spp. or one of two diets containing either low dose (LD) or high dose (HD) levels (50 and 200 cells mL-1, respectively) of toxic A. fundyense for a total of 7 days in two independent experiments. As expected, ingestion of the dinoflagellate had no effect on copepod survival and grazing activity. However, significant reductions of egg production and egg viability were observed in C. finmarchicus females fed on either experimental diet. After the 7-day experiment, total nauplius production by females on the LD and HD diets was reduced by 35% to 75% compared to the control females. These results suggest that blooms of A. fundyense in the Gulf of Maine may be an environmental challenge for C. finmarchicus populations, with a potential negative effect on copepod recruitment.

Embayments and salt ponds along the coast of Massachusetts can host localized blooms of the toxic dinoflagellate Alexandrium fundyense. One such system, exhibiting a long history of toxicity and annual closures of shellfish beds, is the Nauset Marsh System (NMS) on Cape Cod. In order measure net growth rates of natural A. fundyense populations in the NMS during spring 2012, incubation experiments were conducted on seawater samples from two salt ponds within the NMS (Salt Pond and Mill Pond). Seawater samples containing natural populations of grazers and A. fundyense were incubated at ambient temperatures. Concentrations of A. fundyense after incubations were compared to initial abundances to determine net increases from population growth, or decreases presumed to be primarily due to grazing losses. Abundances of both microzooplankton (ciliates, rotifers, copepod nauplii and heterotrophic dinoflagellates) and mesozooplankton (copepodites and adult copepods, marine cladocerans, and meroplankton) grazers were also determined. This study documented net growth rates that were highly variable throughout the bloom, calculated from weekly bloom cell counts from the start of sampling to bloom peak in both ponds (Mill Pond range = 0.12 - 0.46 d-1; Salt Pond range = -0.02 - 0.44 d-1). Microzooplankton grazers that were observed with ingested A. fundyense cells included polychaete larvae, rotifers, tintinnids, and heterotrophic dinoflagellates of the genera Polykrikos and Gymnodinium. Significant A. fundyense net growth was observed in two incubation experiments, and only a single experiment exhibited significant population losses. For the majority of experiments, due to high variability in data, net changes in A. fundyense abundance were not significant after the 24-hr incubations. However, experimental net growth rates through bloom peak were not statistically distinguishable from estimated long-term average net growth rates of natural populations in each pond (Mill Pond = 0.27 d-1 and Salt Pond = 0.20 d-1), which led to peak bloom concentrations on the order of 106 cells l-1 in both ponds. Experimental net growth rates from the incubations underestimated the observed natural net growth rates at several time intervals prior to bloom peak, which may indicate that natural populations experienced additional sources of vegetative cells or periods of reduced losses that the 24-hr incubation experiments did not capture, or that the experimental procedure introduced containment artifacts.

We report on a dense bloom (~1.80 × 105 cells mL-1) of the marine dinoflagellate species Amphidinium carterae (Genotype 2) in a shallow, small intermittently open coastal lagoon in south eastern Australia. This bloom co-occurred with the deaths of >300 individuals of three different species of fish. The opening of the lagoon to the ocean, as well as localized high nutrient levels, preceded the observations of very high cell numbers. A. carterae is usually benthic and sediment-dwelling, but temporarily became abundant throughout the water column in this shallow (<2 m) sandy habitat. Histopathological results showed that the Anguilla reinhardtii individuals examined had damage to epithelial and gill epithelial cells. An analysis of the bloom water indicated the presence of a compound with a retention time and UV spectra similar to Luteophanol A, a compound known from a strain of Amphidinium. Assays with a fish gill cell line were conducted using a purified compound from cells concentrated from the bloom, and was found to cause a loss of 87% in cell viability in 6 h. The fish deaths were likely due to the low dissolved oxygen levels in the water and/or the presence of Luteophanol A-like compounds released during the bloom.

Cyanobacteria are known to produce a wide variety of bioactive, toxic secondary metabolites generally described as hepatotoxins, neurotoxins, cytotoxins, or dermatoxins. In Brazil, the regular monitoring of cyanobacterial toxins has intensified after the death of 65 patients in a hemodialysis clinic in Caruaru in the state of Pernambuco due to microcystin exposure. The primary objective of this study was to use multivariate statistics that incorporated environmental parameters (both biotic and abiotic) to forecast blooms of cyanobacteria and their toxic secondary metabolites in 20 drinking water reservoirs managed by the Water Treatment Company of Ceará (CAGECE) in the semi-arid region of Ceará, Brazil. Across four years (January 2013 to January 2017), 114 different phytoplankton taxa were identified, including 24 cyanobacterial taxa. In general, Ceará reservoirs were dominated by cyanobacteria due to eutrophication but also because of the dry and warm climate found throughout the region. Interestingly, specific cyanobacterial taxa were influenced by different biotic and abiotic factors. For example, nitrogen-to-phosphorus (N:P) and evaporation were positively related to saxitoxin-producing taxa, especially Raphidiopsis raciborskii, while temperature, electrical conductivity, total phosphorus, and transparency (measured as Secchi depth) were positively associated with microcystin-producing taxa, such as Microcystis aeruginosa. Climate forecasts predict higher evaporation and temperatures in the semi-arid Ceará region, which will likely magnify droughts and water scarcity as well as promote toxic cyanobacterial blooms in reservoirs in the future. Therefore, understanding the factors associated with algal blooms dominated by specific taxa is paramount for water resource management.

Species composition and seasonal variations of free-floating Ulva species were investigated in the source area of the world's largest macroalgal blooms during 2009-2015. Based on a combination of a morphological analysis and sequences of nuclear-encoded ITS and 5S rDNA spacer regions, the dominant species in the free-floating Ulva community at the early stage of green tides were Ulva compressa, Ulva flexuosa, and Ulva linza. The first appearance of Ulva prolifera on the sea surface was in mid-May and it dominated the floating Ulva community in June from 2009 to 2011. From 2012-2015, U. prolifera was not only the first species to appear on the sea surface but also the dominant species during the whole early stage of green tides. To explain the successional mechanism, the effects of environmental factors on the growth of four Ulva species were examined in the laboratory under different combinations of light intensity and temperature. It was found that the highest growth rate of U. prolifera was 44.9%/d, which was much higher than the other three Ulva species. The strong tolerance of U. prolifera to extreme conditions also helps it survive and bloom in the Yellow Sea.

Cyanobacterial survival following their release in water from major headwaters reservoirs was compared in five New South Wales rivers. Under low flow conditions, cyanobacterial presence disappeared rapidly with distance downstream in the Cudgegong and Hunter Rivers, whereas the other three rivers were contaminated for at least 300 km. Cyanobacterial survival is likely to be impacted by the geomorphology of each river, especially the extent of gravel riffle reaches (cells striking rocks can destroy them) and by the different turbulent flow conditions it produces within each. Flow conditions at gauging stations were used to estimate the turbulent strain rate experienced by suspended cyanobacteria. These indicate average turbulent strain rates in the Cudgegong and Hunter Rivers can be above 33 and 83 s-1 while for the Murray, Edward and Macquarie Rivers average strain rate was estimated to be less than 30 s-1. These turbulent strain rate estimates are substantially above published thresholds of approximately 2 s-1 for impacts indicated from laboratory tests. Estimates of strain rate were correlated with changes in cyanobacterial biovolume at stations along the rivers. These measurements indicate a weak but significant negative linear relationship between average strain rate and change in cyanobacterial biomass. River management often involves releasing cold deep water with low cyanobacterial presence from these reservoirs, leading to ecological impacts from cold water pollution downstream. The pollution may be avoided if cyanobacteria die off rapidly downstream of the reservoir, allowing surface water to be released instead. However high concentrations of soluble cyanotoxins may remain even after the cyanobacterial cells have been destroyed. The geomorphology of the river (length of riffle reaches) is an important consideration for river management during cyanobacterial blooms in headwater reservoirs.

Azaspiracids (AZA) are the most recently discovered group of lipophilic marine biotoxins of microalgal origin, and associated with human incidents of shellfish poisoning. They are produced by a few species of Amphidomataceae, but diversity and occurrence of the small-sized dinophytes remain poorly explored for many regions of the world. In order to analyze the presence and importance of Amphidomataceae in a highly productive area of Argentinean coastal waters (El Rincón area, SW Atlantic), a scientific cruise was performed in 2015 to sample the early spring bloom. In a multi-method approach, light microscopy was combined with real-time PCR molecular detection of Amphidomataceae, with chemical analysis of AZA, and with the establishment and characterization of amphidomatacean strains. Both light microscopy and PCR revealed that Amphidomataceae were widely present in spring plankton communities along the El Rincón area. They were particularly abundant offshore at the shelf front, reaching peak densities of 2.8 × 105 cells L-1, but no AZA were detected in field samples. In total, 31 new strains were determined as Az. dalianense and Az. spinosum, respectively. All Az. dalianense were non-toxigenic and shared the same rRNA sequences. The large majority of the new Az. spinosum strains revealed for the first time the presence of a non-toxigenic ribotype of this species, which is otherwise the most important AZA producer in European waters. One of the new Az. spinosum strains, with a particular slender shape and some other morphological peculiarities, clustered with toxigenic strains of Az. spinosum from Norway and, exceptionally for the species, produced only AZA-2 but not AZA-1. Results indicate a wide diversity within Az. spinosum, both in terms of sequence data and toxin profiles, which also will affect the qualitative and quantitative performance of the specific qPCR assay for this species. Overall, the new data provide a more differentiated perspective of diversity, toxin productivity and occurrence of Amphidomataceae in a poorly explored region of the global ocean.

Lake Taihu, the third largest freshwater lake in China, suffers from harmful cyanobacteria blooms caused by Microcystis spp., which do not fix nitrogen (N). Reduced N (i.e., NH4+, urea and other labile organic N compounds) is an important factor affecting the growth of Microcystis. As the world use of urea as fertilizer has escalated during the past decades, an understanding of how urea cycling relates to blooms of Microcystis is critical to predicting, controlling and alleviating the problem. In this study, the cycling rates of urea-N in Lake Taihu ranged from non-detectable to 1.37 μmol N L-1 h-1 for regeneration, and from 0.042 μmol N L-1 h-1 to 2.27 μmol N L-1 h-1 for potential urea-N removal. The fate of urea-N differed between light and dark incubations. Increased 15NH4+ accumulated and higher quantities of the removed urea-15N remained in the 15NH4+ form were detected in the dark than in the light. A follow-up incubation experiment with 15N-urea confirmed that Microcystis can grow on urea but its effects on urea dynamics were minor, indicating that Microcystis was not the major factor causing the observed fates of urea under different light conditions in Lake Taihu. Bacterial community composition and predicted functional gene data suggested that heterotrophic bacteria metabolized urea, even though Microcystis spp. was the dominant bloom organism.

Large-scale blooms formed by pico-sized phytoplankton, which strongly inhibited feeding activity and growth of cultured scallops, have been recorded along the coast of Qinhuangdao in the Bohai Sea since 2009. Based on pigment profiles and clone library analysis of phytoplankton samples during the blooms, the major bloom-forming species was primarily identified as pelagophyte Aureococcus anophagefferens Hargraves et Sieburth, the causative species of intensive brown tides in the United States and South Africa. Due to the indistinct morphological features of the bloom-forming microalgae and limited knowledge on the composition of phytoplankton communities, there were still disputes concerning the causative species of the blooms. In this study, the method of high-throughput sequencing targeted 18S rDNA V4 region was used to study the composition of pico- and nano-sized phytoplankton communities in 2013 and 2014. A total of 18 groups of eukaryotic microalgae at the class level and more than 2000 operational taxonomic units (OTUs) were identified in phytoplankton samples collected from the brown-tide zone in the Qinhuangdao coastal waters. For both years, A. anophagefferens was the most dominant species during the bloom period and its maximum relative abundance exceeded 60 percent. Along with other evidence, the results further confirm that A. anophagefferens is the major causative species of the pico-sized phytoplankton blooms in the Bohai Sea. The outbreak of brown tide exhibited a strong inter-annual variation between 2013 and 2014, and an increasing dominance of dinoflagellates could be observed in the Qinhuangdao coastal waters.

The dinoflagellate Prorocentrum donghaiense is a dominant harmful algal bloom (HAB) species on the East China Sea (ECS) coast. The co-occurrence of Karlodinium veneficum with P. donghaiense is often observed and can later develop into dense blooms. However, the role of K. veneficum in P. donghaiense population dynamics is unknown. In the current study, three K. veneficum (GM1, GM2, and GM3) strains were isolated from the ECS with one (GM1) from a mixed, dense bloom of P. donghaiense and other HAB species. All three isolates had identical ITS sequences that were concordant with the species designation. Unique karlotoxin congeners were isolated from one strain (GM2). The sterol compositions of P. donghaiense and K. veneficum were consistent with sensitivity to karlotoxin in the former and insensitivity in the latter. Additional experimentation showed that: (1)in monocultures, higher growth rate of P. donghaiense than K. veneficum is observed in nutrient-enriched and nutrient-depleted media. In co-cultures, the growth of P. donghaiense is inhibited; (2) feeding on P. donghaiense by K. veneficum is clearly demonstrated by fluorescent dye tracking; and (3) the isolated karlotoxin is lethal to P. donghaiense in a concentration-dependent manner. From these studies we propose that K. veneficum may play a negative role in P. donghaiense bloom maintenance and that P. donghaiense may in turn be a bloom initiator as a prey item for K. veneficum.

Thirteen isolates of Prorocentrum species were established from the coral reefs of Perhentian Islands Marine Park, Malaysia and underwent morphological observations and molecular characterization. Six species were found: P. caipirignum, P. concavum, P. cf. emarginatum, P. lima, P. mexicanum and a new morphotype, herein designated as P. malayense sp. nov. Prorocentrum malayense, a species closely related to P. leve, P. cf. foraminosum, P. sp. aff. foraminossum, and P. concavum (Clade A sensu Chomérat et al. 2018), is distinguished from its congeners as having larger thecal pore size and a more deeply excavated V-shaped periflagellar area. Platelet arrangement in the periflagellar area of P. malayense is unique, with the presence of platelet 1a and 1b, platelet 2 being the most anterior platelet, and a broad calabash-shaped platelet 3. The species exhibits consistent genetic sequence divergences for the nuclear-encoded large subunit ribosomal RNA gene (LSU rDNA) and the second internal transcribed spacer (ITS2). The phylogenetic inferences further confirmed that it represents an independent lineage, closely related to species in Clade A sensu Chomérat et al. Pairwise comparison of ITS2 transcripts with its closest relatives revealed the presence of compensatory base changes (CBCs). Toxicity analysis showed detectable levels of okadaic acid in P. lima (1.0-1.6 pg cell-1) and P. caipirignum (3.1 pg cell-1); this is the first report of toxigenic P. caipirignum in the Southeast Asian region. Other Prorocentrum species tested, including the new species, however, were below the detection limit.

The "Applied Simulations and Integrated Modelling for the Understanding of Harmful Algal Blooms" (Asimuth) project sought to develop a harmful algal bloom (HAB) alert system for Atlantic Europe. This was approached by combining, at a national or regional level, regulatory monitoring phytoplankton and biotoxin data with satellite remote sensing and other information on current marine conditions, coupled with regional scale models that included a representation of HAB transport. Synthesis of these products was achieved by expert interpretation within HAB risk alert bulletins that were prepared on a regular basis (typically weekly) for use by the aquaculture industry. In this preface to the Asimuth Special Issue we outline the main HAB species of concern in the region and the strengths and limitations of different methodologies to provide early warning of their blooms.

β-N-methylamino-l-alanine (BMAA), a non-protein amino acid with neurodegenerative features, is known to be produced by cyanobacteria, diatoms and a dinoflagellate. BMAA research has intensified over the last decade, and knowledge has been gained about its bioaccumulation in aquatic and terrestrial ecosystems, toxic effects in model organisms and neurotoxicity in vivo and in vitro. Nevertheless, knowledge of the actual physiological role of BMAA in the producing species or of the ecological factors that regulate BMAA production is still lacking. A few studies propose that BMAA functions to signal nitrogen depletion in cyanobacteria. To investigate whether BMAA might have a similar role in diatoms, two diatom species - Phaeodactylum tricornutum and Thalassiosira weissflogii - were exposed to exogenous BMAA at environmental relevant concentrations, i.e. 0.005, 0.05 and 0.5μM. BMAA was taken up in a concentration dependent manner in both species in the BMAA free fraction and in the protein fraction of T. weissflogii. As a result of the treatments, the diatom cells at some of the time points and at some of the BMAA concentrations exhibited lower concentrations of chlorophyll a and protein, in comparison to controls. At the highest (0.5μM) concentration of BMAA, extracellular ammonia was found in the media of both species at all time points. These results suggest that BMAA interferes with nitrogen metabolism in diatoms, possibly by inhibiting ammonium assimilation via the GS/GOGAT pathway.

A 21-year time series of phytoplankton community structure was analysed in relation to Phaeocystis spp. to elucidate its contribution to the annual carbon budget at station L4 in the western English Channel (WEC). Between 1993-2014 Phaeocystis spp. contributed ∼4.6% of the annual phytoplankton carbon and during the March - May spring bloom, the mean Phaeocystis spp. biomass constituted 17% with a maximal contribution of 47% in 2001. Upper maximal weekly values above the time series mean ranged from 63 to 82% of the total phytoplankton carbon (∼42-137mg carbon (C)m-3) with significant inter-annual variability in Phaeocystis spp. Maximal biomass usually occurred by the end of April, although in some cases as early as mid-April (2007) and as late as late May (2013). The effects of elevated pCO2 on the Phaeocystis spp. spring bloom were investigated during a fifteen-day semi-continuous microcosm experiment. The phytoplankton community biomass was estimated at ∼160mgCm-3 and was dominated by nanophytoplankton (40%, excluding Phaeocystis spp.), Phaeocystis spp. (30%) and cryptophytes (12%). The smaller fraction of the community biomass comprised picophytoplankton (9%), coccolithophores (3%), Synechococcus (3%), dinoflagellates (1.5%), ciliates (1%) and diatoms (0.5%). Over the experimental period, total biomass increased significantly by 90% to ∼305mgCm-3 in the high CO2 treatment while the ambient pCO2 control showed no net gains. Phaeocystis spp. exhibited the greatest response to the high CO2 treatment, increasing by 330%, from ∼50mgCm-3 to over 200mgCm-3 and contributing ∼70% of the total biomass. Taken together, the results of our microcosm experiment and analysis of the time series suggest that a future high CO2 scenario may favour dominance of Phaeocystis spp. during the spring bloom. This has significant implications for the formation of hypoxic zones and the alteration of food web structure including inhibitory feeding effects and lowered fecundity in many copepod species.

The freshwater cyanobacterium Microcystis is a nuisance species. It forms large blooms on the water surface and overwhelmingly dominates the ecosystem through the formation of colonies from single cells surrounded by mucilage; however, the mechanism of colony formation is poorly understood. Two mechanisms of Microcystis colony formation have been proposed: cell-division, where cells remain attached after binary fission; and cell-adhesion, where single cells stick together. This paper examined the published literature on Microcystis colony formation to clarify the mechanism of colony formation and its relationship to environmental drivers. This meta-analysis showed that in laboratory experiments, colony formation by cell-division was mainly induced by zooplankton filtrate, high Pb2+ concentrations, the presence of the cyanobacterium Cylindrospermopsis raciborskii, heterotrophic bacteria, and low temperature and low light intensities. Alternatively, colony formation by cell-adhesion was mainly induced by zooplankton grazing, high Ca2+ concentrations, and microcystins. Therefore, colony formation by cell-division appears to be a slower process and to occur under an environmental stress factor, while cell-adhesion occurs more quickly to an environmental threat. Applying the criteria to the different morphospecies of Microcystis, it was found that under natural conditions M. ichthyoblabe colonies formed predominantly through cell-division, whereas M. wesenbergii colonies formed predominantly through cell-adhesion. This study provides new insights into the mechanisms and environmental drivers of colony formation by Microcystis.

Phosphorus (P) is an essential nutrient for marine phytoplankton as for other living organisms, and the preferred form, dissolved inorganic phosphate (DIP), is often quickly depleted in the sunlit layer of the ocean. Phytoplankton have developed mechanisms to utilize organic forms of P (DOP). Hydrolysis of DOP to release DIP by alkaline phosphatase is believed to be the most common mechanism of DOP utilization. Little effort has been made, however, to understand other potential molecular mechanisms of utilizing different types of DOP. This study investigated the bioavailability of glucose-6-phosphate (G6P) and its underlying molecular mechanism in the dinoflagellate Karenia mikimotoi. Suppression Subtraction Hybridization (SSH) was used to identify genes up- and down-regulated during G6P utilization compared to DIP condition. The results showed that G6P supported the growth and yield of K. mikimotoi as efficiently as DIP. Neither DIP release nor AP activity was detected in the cultures grown in G6P medium, however, suggesting direct uptake of G6P. SSH analysis and RT-qPCR results showed evidence of metabolic modifications, particularly that mitochondrial ATP synthase f1gamma subunit and thioredoxin reductase were up-regulated while diphosphatase and pyrophosphatase were down-regulated in the G6P cultures. All the results indicate that K. mikimotoi has developed a mechanism other than alkaline phosphatase to utilize G6P.

In the present study the geographical distribution, abundance and composition of Gambierdiscus was described over a 600km longitudinal scale in the Canary Islands. Samples for cell counts, isolation and identification of Gambierdiscus were obtained from five islands (El Hierro, Tenerife, Gran Canaria, Fuerteventura and Lanzarote). Average densities of Gambierdiscus spp. between 0 and 2200cellsg-1 blot dry weight of macrophyte were recorded. Morphological (light microscopy and SEM techniques) and molecular analyses (LSU and SSU rDNA sequencing of cultures and single cells from the field) of Gambierdiscus was performed. Five Gambierdiscus species (G. australes, G. caribaeus, G. carolinianus, G. excentricus and G. silvae), together with a new putative species (Gambierdiscus ribotype 3) were identified. These results suggest that some cases of CFP in the region could be associated with the accumulation of ciguatoxins in the marine food web acquired from local populations of Gambierdiscus. This unexpected high diversity of Gambierdiscus species in an area which a priori is not under risk of ciguatera, hints at an ancient settlement of Gambierdiscus populations, likely favored by warmer climate conditions in the Miocene Epoch (when oldest current Canary Islands were created), in contrast with cooler present ones. Currently, warming trends associated with climate change could contribute to extend favorable environmental conditions in the area for Gambierdiscus growth especially during winter months.

The genus Pseudo-nitzschia has attracted attention because of production of the toxin, domoic acid (DA), causing Amnesic Shellfish Poisoning (ASP). Pseudo-nitzschia blooms occur frequently in Chinese coastal waters, and DA has been detected in several marine organisms, but so far no Pseudo-nitzschia strains from Chinese waters have been shown to produce DA. In this study, monoclonal Pseudo-nitzschia strains were established from Chinese coastal waters and examined using light microscopy, electron microscopy and molecular markers. Five strains, sharing distinct morphological and molecular features differentiating them from other Pseudo-nitzschia species, represent a new species, Pseudo-nitzschia simulans sp. nov. Morphologically, the taxon belongs to the P. pseudodelicatissima group, cells possessing a central nodule and each stria comprising one row of poroids. The new species is characterized by the poroid structure, which typically comprises two sectors, each sector located near opposite margins of the poroid. The production of DA was examined by liquid chromatography tandem mass spectrometry (LC-MS/MS) analyses of cells in stationary growth phase. Domoic acid was detected in one of the five strains, with concentrations around 1.05-1.54 fg cell-1. This is the first toxigenic diatom species reported from Chinese waters.

Recent molecular phylogenetic studies of Gambierdiscus species flagged several new species and genotypes, thus leading to revitalizing its systematics. The inter-relationships of clades revealed by the primary sequence information of nuclear ribosomal genes (rDNA), however, can sometimes be equivocal, and therefore, in this study, the taxonomic status of a ribotype, Gambierdiscus sp. type 6, was evaluated using specimens collected from the original locality, Marakei Island, Republic of Kiribati; and specimens found in Rawa Island, Peninsular Malaysia, were further used for comparison. Morphologically, the ribotype cells resembled G. scabrosus, G. belizeanus, G. balechii, G. cheloniae and G. lapillus in thecal ornamentation, where the thecal surfaces are reticulate-foveated, but differed from G. scabrosus by its hatchet-shaped Plate 2', and G. belizeanus by the asymmetrical Plate 3'. To identify the phylogenetic relationship of this ribotype, a large dataset of the large subunit (LSU) and small subunit (SSU) rDNAs were compiled, and performed comprehensive analyses, using Bayesian-inference, maximum-parsimony, and maximum-likelihood, for the latter two incorporating the sequence-structure information of the SSU rDNA. Both the LSU and SSU rDNA phylogenetic trees displayed an identical topology and supported the hypothesis that the relationship between Gambierdiscus sp. type 6 and G. balechii was monophyletic. As a result, the taxonomic status of Gambierdiscus sp. type 6 was revised, and assigned as Gambierdiscus balechii. Toxicity analysis using neuroblastoma N2A assay confirmed that the Central Pacific strains were toxic, ranging from 1.1 to 19.9 fg P-CTX-1 eq cell-1, but no toxicity was detected in a Western Pacific strain. This suggested that the species might be one of the species contributing to the high incidence rate of ciguatera fish poisoning in Marakei Island.

The increased frequency and intensity of drought with climate change may cause an increase in the magnitude and toxicity of freshwater cyanobacteria harmful algal blooms (CHABs), including Microcystis blooms, in San Francisco Estuary, California. As the fourth driest year on record in San Francisco Estuary, the 2014 drought provided an opportunity to directly test the impact of severe drought on cyanobacteria blooms in SFE. A field sampling program was conducted between July and December 2014 to sample a suite of physical, chemical, and biological variables at 10 stations in the freshwater and brackish reaches of the estuary. The 2014 Microcystis bloom had the highest biomass and toxin concentration, earliest initiation, and the longest duration, since the blooms began in 1999. Median chlorophyll a concentration increased by 9 and 12 times over previous dry and wet years, respectively. Total microcystin concentration also exceeded that in previous dry and wet years by a factor of 11 and 65, respectively. Cell abundance determined by quantitative PCR indicated the bloom contained multiple potentially toxic cyanobacteria species, toxic Microcystis and relatively high total cyanobacteria abundance. The bloom was associated with extreme nutrient concentrations, including a 20-year high in soluble reactive phosphorus concentration and low to below detection levels of ammonium. Stable isotope analysis suggested the bloom varied with both inorganic and organic nutrient concentration, and used ammonium as the primary nitrogen source. Water temperature was a primary controlling factor for the bloom and was positively correlated with the increase in both total and toxic Microcystis abundance. In addition, the early initiation and persistence of warm water temperature coincided with the increased intensity and duration of the Microcystis bloom from the usual 3 to 4 months to 8 months. Long residence time was also a primary factor controlling the magnitude and persistence of the bloom, and was created by a 66% to 85% reduction in both the water inflow and diversion of water for agriculture during the summer. We concluded that severe drought conditions can lead to a significant increase in the abundance of Microcystis and other cyanobacteria, as well as their associated toxins.

Many toxic secondary metabolites used for defense are also toxic to the producing organism. One important way to circumvent toxicity is to store the toxin as an inactive precursor. Several sulfated diesters of the diarrhetic shellfish poisoning (DSP) toxin okadaic acid have been reported from cultures of various dinoflagellate species belonging to the genus Prorocentrum. It has been proposed that these sulfated diesters are a means of toxin storage within the dinoflagellate cell, and that a putative enzyme mediated two-step hydrolysis of sulfated diesters such as DTX-4 and DTX-5 initially leads to the formation of diol esters and ultimately to the release of free okadaic acid. However, only one diol ester and no sulfated diesters of DTX-1, a closely related DSP toxin, have been isolated leading some to speculate that this toxin is not stored as a sulfated diester and is processed by some other means. DSP components in organic extracts of two large scale Prorocentrum lima laboratory cultures have been investigated. In addition to the usual suite of okadaic acid esters, as well as the free acids okadaic acid and DTX-1, a group of corresponding diol- and sulfated diesters of both okadaic acid and DTX-1 have now been isolated and structurally characterized, confirming that both okadaic acid and DTX-1 are initially formed in the dinoflagellate cell as the non-toxic sulfated diesters.