Paralytic Shellfish Toxin Production Research Articles

Novel toxin biosynthetic gene cluster in harmful algal bloom-causing Heteroscytonema crispum: Insights into the origins of paralytic shellfish toxins.

Caused by both eukaryotic dinoflagellates and prokaryotic cyanobacteria, harmful algal blooms (HABs) are events of severe ecological, economic, and public health consequence, and their incidence has become more common of late. Despite coordinated research efforts to identify and characterize the genomes of HAB-causing organisms, the genomic basis and evolutionary origins of paralytic shellfish toxins (PSTs) produced by HABs remain at best incomplete. The PST saxitoxin has an especially complex genomic architecture and enigmatic phylogenetic distribution, spanning dinoflagellates and multiple cyanobacterial genera. Using filtration and extraction techniques to target the desired cyanobacteria from non-axenic culture, coupled with a combination of short and long read sequencing, we generated a reference-quality hybrid genome assembly for Heteroscytonema crispum UTEX LB 1556, a freshwater, PST-producing cyanobacterium thought to have the largest known genome in its phylum. We report a complete, novel biosynthetic gene cluster for the PST saxitoxin. Leveraging this biosynthetic gene cluster, we find support for the hypothesis that PST production has appeared in divergent Cyanobacteria lineages through widespread and repeated horizontal gene transfer. This work demonstrates the utility of long-read sequencing and metagenomic assembly toward advancing our understanding of PST biosynthetic gene cluster diversity and suggests a mechanism for the origin of PST biosynthetic genes.

Effects of Zooplankton Extracts on the Production of Paralytic Shellfish Toxins by Gymnodinium catenatum and Alexandrium pacificum

Effects of Zooplankton Extracts on the Production of Paralytic Shellfish Toxins by Gymnodinium catenatum and Alexandrium pacificum

Unraveling the risks of nAl2O3 on harmful algal blooms: Insights from paralytic shellfish toxins production of Alexandrium tamarense

As one of the commonly used and cost-effective nanomaterials, nanosized aluminum oxide (nAl2O3) posses unique properties and chemical stability. However, its extensive use and resultant dissemination into aquatic ecosystems prompt concerns over the proliferation and repercussions of harmful algal blooms, particularly those caused by dinoflagellates producing toxins. This study investigated the sub-chronic effects of nAl2O3 on growth, physiological activities, and paralytic shellfish toxins (PSTs) production in Alexandrium tamarense. Results showed dose-dependent inhibition in growth (EC50 = 20.6 mg L−1), esterase activity, and photosynthetic efficiency (Fv/Fm) during the sub-chronic exposure (13-day). The internalization of nAl2O3 in microalgal cells and the significant decrease in the total cellular PSTs content were observed under high nAl2O3 concentrations (>40 mg L−1). The study also demonstrated a clear decrease in the content of some derivatives of PSTs (GTX5, C1/2, and GTX2/3) with the increase in nAl2O3 concentrations, accompanied by the induction of an unknown derivative. Excessive ROS production, dissolved Al, and physical inhibition were suggested as potential mechanisms for nAl2O3 toxicity and changes in PSTs toxin profile. Overall, this research enhances our understanding of the potentiated risks and threats on the possible concurrent events of toxic dinoflagellate, such as Alexandrium species and nanoparticles in aquatic environments.

Elevated pCO2 may increase the edible safety risk of clams exposed to toxic Alexandrium spp.

Toxic harmful algal blooms (HABs) have received increasing attention owing to their threat to the health of aquatic life and seafood consumers. This study evaluated the impacts of elevated atmospheric partial pressure of CO2 (pCO2) on the production of paralytic shellfish toxins (PSTs) in different Alexandrium spp. strains, together with its further effects on the bioaccumulation/elimination dynamics of PSTs in bivalves contaminated with PSTs from toxic dinoflagellates. Our results showed that elevated pCO2 stimulated the growth of the two Alexandrium spp. (A. catenella and A. pacificum) isolated from the northern and southern coastal areas of China, respectively, and affected PST production including content and toxicity of the two strains differently. Further PSTs bioaccumulation/elimination in PSTs-contaminated Manila clam, Ruditapes philippinarum under high pCO2 also occurred. It is worth noting the biotransformation of neosaxitoxin (NEO) with high toxicity through trophic transfer with effect of elevated pCO2. When in microalgae cultured under the control (410 ppm) and elevated pCO2 conditions (495 and 850 ppm), the proportion of NEO in the PST content produced by A. catenella was reduced from 11.1 to 6.4 and 2.6 %, while the proportion of NEO in A. pacificum was increased from 3.1 to 3.6 and 4.7 %, respectively. NEO accounted for >50 % of total PST contents in clams, which were biotransformed via transfer from dinoflagellates and higher pCO2 enhanced this biotransformation leading to increased NEO accumulation. The negatively affected elimination of PSTs, especially NEO, in clams fed with A. catenella or A. pacificum, indicates that the detoxification of PSTs-contaminated clams may be more difficult under elevated pCO2. This study provides reference for developing models to assess the safety of bivalves under the co-stress of environmental change and toxic HABs, suggesting that ocean acidification may lead to the higher safety risk of Manila clams exposed to toxic HAB dinoflagellates.

Variability in Paralytic Shellfish Toxin Profiles and Dinoflagellate Diversity in Mussels and Seawater Collected during Spring in Korean Coastal Seawater.

Paralytic shellfish toxins (PSTs) are potent neurotoxins produced by certain microalgae, particularly dinoflagellates, and they can accumulate in shellfish in coastal seawater and thus pose significant health risks to humans. To explore the relationship between toxicity and PST profiles in seawater and mussels, the spatiotemporal variations in PST concentrations and profiles were investigated along the southern coast of Korea under peak PST levels during spring. Seawater and mussel samples were collected biweekly from multiple stations, and the toxin concentrations in the samples were measured. Moreover, the dinoflagellate community composition was analyzed using next-generation sequencing to identify potential PST-producing species. The PST concentrations and toxin profiles showed substantial spatiotemporal variability, with GTX1 and GTX4 representing the dominant toxins in both samples, and C1/2 tending to be higher in seawater. Alexandrium species were identified as the primary sources of PSTs. Environmental factors such as water temperature and salinity influenced PST production. This study demonstrates that variability in the amount and composition of PSTs is due to intricate ecological interactions. To mitigate shellfish poisoning, continuous monitoring must be conducted to gain a deeper understanding of these interactions.

Toxic dinoflagellate Centrodinium punctatum (Cleve) F.J.R. Taylor: An examination on the responses in growth and toxin contents to drastic changes of temperature and salinity

To understand environmental effects affecting paralytic shellfish toxin production of Centrodinium punctatum, this study examined the growth responses, and toxin contents and profiles of a C. punctatum culture exposed to drastic changes of temperature (5-30 °C) and salinity (15–40). C. punctatum grew over a temperature range of 15–25 °C, with an optimum of 20 °C., and over a salinity range of 25–40, with optimum salinities of 30–35. This suggests that C. punctatum prefers relatively warm waters and an oceanic habitat for its growth and can adapt to significant changes of salinity levels. When C. punctatum was cultivated at different temperature and salinity levels, the PST profile included four major analogs (STX, neoSTX, GTX1 and GTX4, constituted >80 % of the profile), while low amounts of doSTX and traces of dc-STX and dc-GTX2 were also observed. Interestingly, though overall toxin contents did not change significantly with temperature, increases in the proportion of STX, and decreases in proportions in GTX1 and GTX4 were observed with higher temperatures. Salinity did not affect either toxin contents or profile from 25 to 35. However, the total toxin content dropped to approximately half at salinity 40, suggesting this salinity may induce metabolic changes in C. punctatum.

ROS meditated paralytic shellfish toxins production changes of Alexandrium tamarense caused by microplastic particles

A variety of studies have investigated the toxic effects of microplastics (MPs) on microalgae, but few of them considered their influence on dinoflagellate toxins production, which could cause significant ecological safety concerns in coastal areas. This research investigated the impacts of 5 μg L−1 and 5 mg L−1 polystyrene (PS) MPs on the changes of paralytic shellfish toxins (PSTs) production and their relationship with cellular oxidative stress of Alexandrium tamarense, a common harmful algal blooms causative dinoflagellate. The results showed elevation of reactive oxygen species (ROS) levels, activation of antioxidant system and overproduction of PSTs were positively correlated under PS MPs exposure (especially under 5 mg L−1 PS MPs), and the PSTs changes were eliminated by the ROS inhibitor. Further transcriptomic analysis revealed that ROS could enhance biosynthesis of glutamate, providing raw materials for PSTs precursor arginine, accompanied with enhanced acetyl-CoA and ATP production, finally leading to the overproduction of PSTs. Moreover, the oxidative intracellular environments might block the reduction process from STX to C1&C2, leading to the increase of STX and decrease of C1&C2 proportions. This work brings the first evidence that ROS could mediate PSTs production and compositions of Alexandrium under MPs exposure, with important scientific and ecological significance.

Genomic copy number variability at the genus, species and population levels impacts in situ ecological analyses of dinoflagellates and harmful algal blooms

The application of meta-barcoding, qPCR, and metagenomics to aquatic eukaryotic microbial communities requires knowledge of genomic copy number variability (CNV). CNV may be particularly relevant to functional genes, impacting dosage and expression, yet little is known of the scale and role of CNV in microbial eukaryotes. Here, we quantify CNV of rRNA and a gene involved in Paralytic Shellfish Toxin (PST) synthesis (sxtA4), in 51 strains of 4 Alexandrium (Dinophyceae) species. Genomes varied up to threefold within species and ~7-fold amongst species, with the largest (A. pacificum, 130 ± 1.3 pg cell−1 /~127 Gbp) in the largest size category of any eukaryote. Genomic copy numbers (GCN) of rRNA varied by 6 orders of magnitude amongst Alexandrium (102– 108 copies cell−1) and were significantly related to genome size. Within the population CNV of rRNA was 2 orders of magnitude (105 – 107 cell−1) in 15 isolates from one population, demonstrating that quantitative data based on rRNA genes needs considerable caution in interpretation, even if validated against locally isolated strains. Despite up to 30 years in laboratory culture, rRNA CNV and genome size variability were not correlated with time in culture. Cell volume was only weakly associated with rRNA GCN (20–22% variance explained across dinoflagellates, 4% in Gonyaulacales). GCN of sxtA4 varied from 0–102 copies cell−1, was significantly related to PSTs (ng cell−1), displaying a gene dosage effect modulating PST production. Our data indicate that in dinoflagellates, a major marine eukaryotic group, low-copy functional genes are more reliable and informative targets for quantification of ecological processes than unstable rRNA genes.

Thecal plate morphology, molecular phylogeny, and toxin analyses reveal two novel species of Alexandrium (Dinophyceae) and their potential for toxin production

This study describes two novel species of marine dinophytes in the genus Alexandrium. Morphological characteristics and phylogenetic analyses support the placement of the new taxa, herein designated as Alexandrium limii sp. nov. and A. ogatae sp. nov. Alexandrium limii, a species closely related to A. taylorii, is distinguished by having a shorter 2ʹ/4ʹ suture length, narrower plates 1ʹ and 6ʹʹ, with larger length: width ratios, and by the position of the ventral pore (Vp). Alexandrium ogatae is distinguishable with its metasert plate 1ʹ having almost parallel lateral margins, and by lacking a Vp. Production of paralytic shellfish toxins (PSTs), cycloimines, and goniodomins (GDs) in clonal cultures of A. ogatae, A. limii, and A. taylorii were examined analytically and the results showed that all strains contained GDs, with GDA as major variants (6–14 pg cell−1) for all strains except the Japanese strain of A. limii, which exclusively had a desmethyl variant of GDA (1.4–7.3 pg cell−1). None of the strains contained detectable levels of PSTs and cycloimines.

Uncovering the regulation effect of modified clay on toxin production in Alexandrium pacificum: From physiological insights

As a common dinoflagellate, Alexandrium pacificum can produce paralytic shellfish toxins (PSTs). It can be removed from water by Polyaluminium chloride modified clay (PAC-MC), but it is unclear whether PAC-MC can prevent PSTs content and toxicity from increasing and whether PAC-MC can stimulate PSTs biosynthesis by A. pacificum. Effect of PAC-MC on PSTs and the physiological mechanism were analysed here. The results showed total PSTs content and toxicity decreased respectively by 34.10 % and 48.59 % in 0.2 g/L PAC-MC group at 12 days compared with control group. And the restriction of total PSTs by PAC-MC was mainly achieved via inhibition of algal cell proliferation, by affecting A. pacificum physiological processes and changing phycosphere microbial community. Meanwhile, single-cell PSTs toxicity did not increase significantly throughout the experiment. Moreover, A. pacificum treated with PAC-MC tended to synthesize sulfated PSTs such as C1&2. Mechanistic analysis showed that PAC-MC induced upregulation of sulfotransferase sxtN (related to PSTs sulfation), and functional prediction of bacterial community also showed significant enrichment of “sulfur relay system” after PAC-MC treatment, which might also promote PSTs sulfation. The results will provide theoretical guidance for the application of PAC-MC to field control of toxic Alexandrium blooms.

Proteomic and toxicological analysis of the response of dinoflagellate Alexandrium catenella to changes in NaNO3 concentration

Dinoflagellates of the genus Alexandrium cause Harmful Algal Blooms (HABs) in coastal waters worldwide, damaging marine environments, aquaculture, and human health. They synthesize potent neurotoxic alkaloids known as PSTs (i.e., Paralytic Shellfish Toxins), the etiological agents of PSP (i.e., Paralytic Shellfish Poisoning). In recent decades, the eutrophication of coastal waters with inorganic nitrogen (e.g., nitrate, nitrite, and ammonia) has increased the frequency and scale of HABs. PSTs concentrations within Alexandrium cells can increase by up to 76% after a nitrogen enrichment event; however, the mechanisms that underlie their biosynthesis in dinoflagellates remains unclear. This study combines mass spectrometry, bioinformatics, and toxicology and investigates the expression profiles of PSTs in Alexandrium catenella grown in 0.4, 0.9 and 1.3 mM NaNO3. Pathway analysis of protein expression revealed that tRNA amino acylation, glycolysis, TCA cycle and pigment biosynthesis were upregulated in 0.4 mM and downregulated in 1.3 mM NaNO3 compared to those grown in 0.9 mM NaNO3. Conversely, ATP synthesis, photosynthesis and arginine biosynthesis were downregulated in 0.4 mM and upregulated in 1.3 mM NaNO3. Additionally, the expression of proteins involved in PST biosynthesis (sxtA, sxtG, sxtV, sxtW and sxtZ) and overall PST production like STX, NEO, C1, C2, GTX1-6 and dcGTX2 was higher at lower nitrate concentrations. Therefore, increased nitrogen concentrations increase protein synthesis, photosynthesis, and energy metabolism and decrease enzyme expression in PST biosynthesis and production. This research provides new clues about how the changes in the nitrate concentration can modulate different metabolic pathways and the expression of PST biosynthesis in toxigenic dinoflagellates.

Genetic association of toxin production in the dinoflagellate Alexandrium minutum.

Dinoflagellates of the genus Alexandrium are responsible for harmful algal blooms and produce paralytic shellfish toxins (PSTs). Their very large and complex genomes make it challenging to identify the genes responsible for toxin synthesis. A family-based genomic association study was developed to determine the inheritance of toxin production in Alexandrium minutum and identify genomic regions linked to this production. We show that the ability to produce toxins is inheritable in a Mendelian way, while the heritability of the toxin profile is more complex. We developed the first dinoflagellate genetic linkage map. Using this map, several major results were obtained: 1. A genomic region related to the ability to produce toxins was identified. 2. This region does not contain any polymorphic sxt genes, known to be involved in toxin production in cyanobacteria. 3. The sxt genes, known to be present in a single cluster in cyanobacteria, are scattered on different linkage groups in A. minutum. 4. The expression of two sxt genes not assigned to any linkage group, sxtI and sxtG, may be regulated by the genomic region related to the ability to produce toxins. Our results provide new insights into the organization of toxicity-related genes in A. minutum, suggesting a dissociated genetic mechanism for the production of the different analogues and the ability to produce toxins. However, most of the newly identified genes remain unannotated. This study therefore proposes new candidate genes to be further explored to understand how dinoflagellates synthesize their toxins.

Formation mechanism and environmental drivers of Alexandrium catenella bloom events in the coastal waters of Qinhuangdao, China

In the last 5 years, paralytic shellfish toxins (PSTs) have been recurrently detected in mollusks farmed in the mussel culture area of Qinhuangdao city, along with the occurrence of toxic outbreaks linked to dinoflagellate species of the Alexandrium genus. To understand the formation mechanism and variation of these events, continuous and comprehensive PSTs monitoring was carried out between 2017 and 2020. Through the analysis of both phytoplankton and cysts via light microscopy and quantitative polymerase chain reaction, it was shown that Alexandrium catenella was responsible for the production of PSTs, which consisted mainly of gonyautoxins 1,4 (GTX1/4, 87%) and GTX2/3 (13%). During bloom events in 2019, mussels accumulated the highest PSTs value (929 μg STX di-HCl eq·kg−1) in conjunction with the peak of cell abundances, and toxin profiles were consistent with high distributions of GTX1/4, GTX2/3, and Neosaxitoxin. Toxin metabolites vary in different substances and mainly transferred to a stable proportion of α-epimer: β-epimers 3:1. The environmental drivers of Alexandrium blooms included the continuous rise of water temperature (>4 °C) and calm weather with low wind speed and no significant precipitation. By comparing toxin profiles and method sensitivity, it was found that dissolved toxins in seawater are more useful for early warning. These results have important implications for the effective monitoring and management of paralytic shellfish poisoning outbreaks.

Toxin production of dinoflagellate Gymnodinium catenatum isolated from the East China Sea

Dinoflagellate Gymnodinium catenatum is an important producer of paralytic shellfish toxins (PSTs), including a novel group of hydroxybenzoate derivatives named GC toxins. In the East China Sea, G. catenatum has been considered as the causative agent for several paralytic shellfish poisoning (PSP) episodes, yet the knowledge on their toxin production was still quite limited. In this study, toxins produced by a strain of G. catenatum (MEL11) isolated from the East China Sea were determined, using high performance liquid chromatography coupled with tandem mass spectrometry (HPLC-MS/MS). Changes of toxin profile in the stain MEL11 in response to nutrient and temperature variations were also examined. A total of 11 PST components dominated by hydroxybenzoate analogs and N-sulfocarbamoyl toxins were detected, which was different from other G. catenatum strains previously established in the East China Sea in the presence of GC5 and the lack of dcGTX2&3. Cellular toxin composition and content of the strain had no apparent change within a range of temperature from 20°C to 26°C. In contrast, nutrient limitation and nitrogen source had notable impacts on toxin production. The molar percentage of GC toxins decreased remarkably at the stationary growth phase under nutrient-deprived conditions of both nitrogen (N) and phosphorus (P). The replacement of nitrate with ammonium as the source of N significantly promoted PST production by G. catenatum. The study revealed the potential diversity of toxin profiles of G. catenatum in the East China Sea, and highlighted the effects of nutrients on production of GC toxins by G. catenatum.

Chlorophyll-a and Sea Surface Temperature Changes in Relation to Paralytic Shellfish Toxin Production off the East Coast of Tasmania, Australia

Toxic phytoplankton have been detrimental to the fishing and aquaculture industry on the east coast of Tasmania, causing millions of dollars in loss due to contaminated seafood. In 2012–2017, shellfish stocks were poisoned by Alexandrium catenella, a dinoflagellate species that produces paralytic shellfish toxins (PST). Remote sensing data may provide an environmental context for the drivers of PST events in Tasmania. We conducted spatial and temporal trend analyses of the Multi-Scale Ultra-High-Resolution Sea Surface Temperature (MUR SST) and Ocean Color Climate Change Initiative chlorophyll-a (OC-CCI chl-a) to determine if SST and chl-a correlated with the major toxin increases from 2012 to 2017. Along with the trends, we compare the remotely sensed oceanographic parameters of SST and chl-a to toxin events off the east coast of Tasmania to provide environmental context for the high-toxin period. Spatial and temporal changes for chl-a differ based on the north, central, and southeast coast of Tasmania. For sites in the north, chl-a was 5.3% higher from the pre-PST period relative to the PST period, 5.1% along the central part of the coast, and by 6.0% in the south based on deviations from the coastal study area time series. Overall, SST has slightly decreased from 2007 to 2020 (tau = −0.011, p = 0.827) and chl-a has significantly decreased for the east coast (tau = −0.164, p = 1.58 × 10−3). A negative relationship of SST and PST values occurred in the north (r = −0.530, p = 5.32 × 10−5) and central sites (r = −0.225, p = 0.157). The correlation between satellite chl-a (from OC-CCI, Visible Infrared Imaging Radiometer Suite (VIIRS), and Moderate-Resolution Imaging Spectrometer (MODIS) Aqua) and in situ data is weak, which makes it difficult to assess relationships present between chl-a and toxin concentrations. Moving forward, the development of a regional chl-a algorithm and increased in situ chl-a collection and plankton sampling at a species level will help to improve chl-a measurements and toxic phytoplankton production monitoring around Tasmania.

The effect of iron on Chilean Alexandrium catenella growth and paralytic shellfish toxin production as related to algal blooms

The dinoflagellate Alexandrium catenella is a well-known paralytic shellfish toxin producer that forms harmful algal blooms (HABs) worldwide. Blooms of this species have repeatedly brought severe ecological and economic impacts to Chile, especially in the southern region, where the shellfish and salmon industries are world-famous. The mechanisms of such HABs have been intensively studied but are still unclear. Nutrient overloading is one of the often-discussed drivers for HABs. The present study used the A. catenella strain isolated from southern Chile to investigate how iron conditions could affect their growth and toxin production as related to HAB. Our results showed that an optimum concentration of iron was pivotal for proper A. catenella growth. Thus, while excess iron exerted a toxic effect, low iron media led to iron insufficiency and growth inhibition. In addition, the study shows that the degree of paralytic shellfish toxin production by A. catenella varied depending on the iron concentration in the culture media. The A. catenella strain from southern Chile produced GTX1-4 exclusively in the fmol cell−1 scale. Based on these findings, we suggest that including iron and paralytic shellfish toxin measurements in the fields can improve the current HAB monitoring and contribute to an understanding of A. catenella bloom dynamics in Chile.

Novel Non-paralytic Shellfish Toxin and Non-spirolide Toxicity to Finfish, Brine Shrimp, and Rotifer Observed in a Culture of the Dinoflagellate Alexandrium insuetum Isolated From the Coastal Water of China

The genus Alexandrium is one of the major harmful algal blooms (HABs)-forming dinoflagellate group and at least half of ~40 described species have been reported to produce paralytic shellfish toxins (PSTs). The potentially harmful species Alexandrium insuetum has been reported from many countries of Asia and Europe, and to have paralytic shellfish poisoning toxicity, but no mortality of marine animals was observed during its bloom. Therefore, it is ecologically important to characterize the possible toxicity and toxins of this organism. In this study, based on the establishment of two clonal cultures through cyst germination collected from the Yellow Sea, we identified A. insuetum from China as the first record via light microscopy (LM) and scanning electron microscopy (SEM) observations and phylogenetic analyses. The cultures of A. insuetum were further observed to be toxic to finfish and zooplankton and deleterious to rotifer eggs via laboratory bioassays. The exposure bioassays using rotifer (Brachionus plicatilis), brine shrimp (Artemia salina), and larval finfish (Oryzias melastigma) demonstrated that A. insuetum caused significant lethal effects on finfish and zooplankton species. Rotifer bioassays using cell-free culture medium, heat-treated cultures, and water, methanol, and trichloromethane extracts of algal cells revealed that A. insuetum produced heat-labile, water-soluble toxin(s) that could be excreted from A. insuetum cells and steadily accumulated in the medium during the growth phases. Hatching success of rotifer eggs was also found to be seriously affected by the exposure to A. insuetum. Importantly, ultra-high performance liquid chromatography-tandem mass spectrometry [UPLC (or LC)-MS-MS] analyses suggest the above-described toxicity of A. insuetum was caused by neither PSTs nor spiroimines (13-desmethyl spirolide C and gymnodimine). Collectively, our findings demonstrated the novel toxicity to finfish and zooplankton in A. insuetum, which is ecologically important in not only possibly contributing to population dynamics and even the formation of HABs of the species, but also affecting the on-the-spot survival and the reproduction potency of marine animals. The present work is believed to set a cornerstone for the monitoring and risk assessment of the species along the coastal waters of China and for understanding the general ecology of A. insuetum.

Toxicity and histopathological effects of toxic dinoflagellate, Alexandrium catenella exudates on larvae of blue mussel, Mytilus galloprovincialis, and Pacific oyster, Crassostrea gigas

HighlightToxicity and pathological effects of A. catenella were investigated on shellfish larvaeUnfiltered exudates of A. catenella caused significant mortality of blue mussel larvaeApplication of 0.22 mm filtration on A. fundyense exudates potentially decrease the toxicity effectsPathological effects of A. catenella occurred as early as 3 h after exposureThe prevalence and intensity of necrosis increased with exposure duration to A. catenella exudatesAbstractBlooms of the toxic dinoflagellate Alexandrium catenella have affected shellfish industries globally due to their capacity to produce paralytic shellfish toxins(PST). This study aimed to investigate the toxicity effect of exudate A. catenella on larvae of blue mussel Mytilus galloprovincialis and Pacific oyster Crassostrea gigas and filtration methods to reduce the toxic effect. Blue mussel and Pacific oyster larvae were assessed their survival and histopathological changes after exposure to extracellular exudates of A. catenella ranging from 100 to 1,000 cells ml-1 . The results showed that exposure to exudate A. catenella caused significantly higher larval mortality (39 to 52%) than exposure to an equivalent biovolume of the nontoxic species, Tisochrysis lutea (33%) or unfed controls (17%). Filter-sterilization (0.22 µm) of exudates and activated carbon filtration decreased the mortality of Pacific oyster larvae to a level similar to controls (unfed), with the exception of the highest concentrations (600 and 1,000 cells ml-1 ) and mortality of bluemussel larvae mortality by 32% respectively. Blue mussel larvae exposed to exudate A. catenella showed pathological changes mainly in the stomach (digestive gland and style sac) as early as three hours after onset of exposure. The findings of this study suggest that early detection of blooms in the vicinity of mussel and Pacific oyster hatcheries and taking steps to mitigate their effects, is important to reduce the effects of A. catenella blooms on shellfish larval rearing.

A Mediterranean Alexandrium taylorii (Dinophyceae) Strain Produces Goniodomin A and Lytic Compounds but Not Paralytic Shellfish Toxins.

Species of the dinophyte genus Alexandrium are widely distributed and are notorious bloom formers and producers of various potent phycotoxins. The species Alexandrium taylorii is known to form recurrent and dense blooms in the Mediterranean, but its toxin production potential is poorly studied. Here we investigated toxin production potential of a Mediterranean A. taylorii clonal strain by combining state-of-the-art screening for various toxins known to be produced within Alexandrium with a sound morphological and molecular designation of the studied strain. As shown by a detailed thecal plate analysis, morphology of the A. taylorii strain AY7T from the Adriatic Sea conformed with the original species description. Moreover, newly obtained Large Subunit (LSU) and Internal Transcribed Spacers (ITS) rDNA sequences perfectly matched with the majority of other Mediterranean A. taylorii strains from the databases. Based on both ion pair chromatography coupled to post-column derivatization and fluorescence detection (LC-FLD) and liquid chromatography coupled to tandem mass spectrometry (LC-MS/MS) analysis it is shown that A. taylorii AY7T does not produce paralytic shellfish toxins (PST) above a detection limit of ca. 1 fg cell−1, and also lacks any traces of spirolides and gymnodimines. The strain caused cell lysis of protistan species due to poorly characterized lytic compounds, with a density of 185 cells mL−1 causing 50% cell lysis of cryptophyte bioassay target cells (EC50). As shown here for the first time A. taylorii AY7T produced goniodomin A (GDA) at a cellular level of 11.7 pg cell−1. This first report of goniodomin (GD) production of A. taylorii supports the close evolutionary relationship of A. taylorii to other identified GD-producing Alexandrium species. As GD have been causatively linked to fish kills, future studies of Mediterranean A. taylorii blooms should include analysis of GD and should draw attention to potential links to fish kills or other environmental damage.

Toxic dinoflagellate blooms of Gymnodinium catenatum and their cysts in Taiwan Strait and their relationship to global populations

Gymnodinium catenatum is able to produce paralytic shellfish toxins (PSTs) and was responsible for a massive bloom in the Taiwan Strait, East China Sea, in June 2017, which resulted in serious human poisoning and economic losses. To understand the origin of the bloom and determine the potential for blooms in subsequent years, water and sediment samples collected in the Taiwan Strait from 2016 to 2019 were analyzed for cells and cysts using light microscopy (LM) and/or quantitative polymerase chain reaction (qPCR). The morphology of both cells and cysts from the field and cultures was examined with LM and scanning electron microscopy (SEM). Large subunit (LSU) and/or internal transcribed spacer (ITS)-5.8S rRNA gene sequences were obtained in 13 isolates from bloom samples and five strains from cysts. In addition, cells of strains TIO523 and GCLY02 (from the Taiwan Strait and Yellow Sea of China, respectively) were subjected to growth experiments, and cysts from the field were used for germination experiments under various temperatures. Our strains shared identical LSU and ITS-5.8S rRNA gene sequences with those from other parts of the world, and therefore belonged to a global population. A low abundance of G. catenatum cells were detected during most of the sampling period, but a small bloom was encountered in Quanzhou on June 8, 2018. Few cysts were observed in 2016 but a marked increase was observed after the bloom in 2017, with a highest density of 689 cysts cm−3. Cysts germinated at temperatures between 14 and 23 °C with a final germination rate over 93%. Strains TIO523 and GCLY02 displayed growth at temperatures between 17 and 26 °C and 14 and 26 °C, respectively, with both strains displaying the highest growth rate of ca. 0.5 divisions d–1 at 23 °C. The PSTs of the three strains and cysts from the sediments were analyzed by liquid chromatography with tandem mass spectrometry (LC-MS/MS). All strains were able to produce PSTs, which were dominated by N-sulfocarbamoyl C toxins (C1/2, 53.0–143.5 pg cell−1) and decarbamoyl gonyautoxins (dcGTX2/3, 26.7–52.1 pg cell−1), although they were not detected in cysts. However, hydroxybenzoyl (GC) toxins were detected in both cells and cysts. Our results suggested that the population in the Taiwan Strait belonged to a warm water ecotype and has a unique toxin profile. Our results also suggested that the persistence of cells in the water column may have initiated the bloom.