Paralytic Shellfish Toxins Concentrations Research Articles

Paralytic shellfish toxins producing dinoflagellates cause dysbacteriosis in scallop gut microbial biofilms

Filter-feeding bivalves could accumulate paralytic shellfish toxins (PSTs) produced by harmful dinoflagellates through diet. Despite that bivalves are resistant to these neurotoxins due to possessing PST-resistant sodium channel, exposure to PSTs-producing dinoflagellates impair bivalve survival. We hypothesized that ingesting PSTs-producing dinoflagellates may influence the gut microbiota, and then the health of bivalves. To test this idea, we compared the gut microbiota of the scallop Patinopecten yessoensis, after feeding with PST-producing or non-toxic dinoflagellates. Exposure to PSTs-producing dinoflagellates resulted in a decline of gut microbial diversity and a disturbance of community structure, accompanied by a significant increase in the abundance and richness of pathogenic bacteria, represented by Vibrio. Moreover, network analysis demonstrated extensive positive correlations between pathogenic bacteria abundances and PSTs concentrations in the digestive glands of the scallops. Furthermore, isolation of a dominant Vibrio strain and its genomic analysis revealed a variety of virulence factors, including the tolC outer membrane exporter, which were expressed in the gut microbiota. Finally, the infection experiment demonstrated scallop mortality caused by the isolated Vibrio strain; further, the pathogenicity of this Vibrio strain was attenuated by a mutation in the tolC gene. Together, these findings demonstrated that the PSTs may affect gut microbiota via direct and taxa-specific interactions with opportunistic pathogens, which proliferate after transition from seawater to the gut environment. The present study has revealed novel mechanisms towards deciphering the puzzles in environmental disturbances-caused death of an important aquaculture species.

Phycotoxins in bivalves from the western Canadian Arctic: The first evidence of toxigenicity

This study presents the first evidence that a diverse suite of phycotoxins is not only being actively produced by the toxigenic algal communities in the Canadian Arctic waters, but is also entering the marine food web. We detected measurable amounts of Amnesic Shellfish Toxins (ASTs) and Paralytic Shellfish Toxins (PSTs), as well as trace amounts of other lipophilic toxin groups including pectenotoxins, yessotoxins, and cyclic imines, in bivalves collected from the Canadian Beaufort Sea in 2014 and 2018. There appear to be species-specific differences in accumulation and retention of AST by Arctic bivalves, with significantly higher concentrations recorded in Nuculanidae than Propeamussiidae, likely reflecting physiological and allometric differences. We further confirm the omnipresence of potentially toxic taxonomically-versatile phytoplankton communities in the western Canadian Arctic comprising Pseudo-nitzschia delicatissima group, P. obtusa, Dinophysis acuminata, Prorocentrum minimum, Alexandrium tamarense, and Gymnodinium spp. Although measurements of actual toxicity levels and profiles of these species at the time of sampling fall outside of the scope of this study, we show that high abundance and competitive success of known AST-producers, Pseudo-nitzschia spp., are possible in Canadian Arctic waters. In 2014, a strong dominance of Pseudo-nitzschia spp. was observed at a few shallow coastal stations, representing nearly 40% of the total phytoplankton cell abundances with > 106 cells/L at the depth of maximum chlorophyll a. We further describe oceanographic conditions conducive to high abundances of toxin-producing algae, indicating that temperature is likely a key factor. Even though measured AST and PST concentrations in bivalve tissue remained well below the Health Canada's levels at which monitored fisheries would close, i.e., 5% and 4%, respectively, their presence demonstrate that phycotoxin accumulation is occurring in food webs of the Canadian Beaufort Sea. Yet, the phycotoxin production controls and trophic transfer mechanisms remain unknown. Canadian Arctic marine ecosystems are rapidly changing and temperatures are expected to continue to increase. Given that these changes simultaneously affect multiple, and often co-occurring, species of primary producers, adaptive capacity is likely to play an important role in the structure of phytoplankton communities in the Canadian Arctic.

Transcriptome Analysis Reveals the Genes Involved in Oxidative Stress Responses of Scallop to PST-Producing Algae and a Candidate Biomarker for PST Monitoring.

Paralytic shellfish toxins (PST) could be accumulated in bivalves and cause safety problems. To protect public health, bivalves are examined for PST contamination before entering the market, usually by high-performance liquid chromatography (HPLC) or LC-tandem mass spectrometry (LC-MS/MS) in the lab, which needs PST standards not all available and is time-consuming for large sample sizes. To detect PST toxicity in bivalves rapidly and sensitively, a biomarker gene is highly demanded, but the related study is very limited. In this study, we fed a commercially important bivalve, Patinopecten yessoensis, with the PST-producing dinoflagellate Alexandrium catenella. After 1, 3, and 5 days of exposure, both PST concentrations and toxicity levels in the digestive gland continuously increased. Transcriptome analysis revealed that the differentially expressed genes were significantly enriched in oxidation-reduction process, which included the cytochrome P450 genes (CYPs), type I iodothyronine deiodinase (IOD1s), peroxidasin (PXDN), and acyl-Coenzyme A oxidase 1 (ACOX1) at day 1 and a superoxide dismutase (SOD) at day 5, highlighting the crucial roles of these genes in response to oxidative stress induced by PST. Among the 33 continuously upregulated genes, five showed a significant correlation between gene expression and PST concentration, with the highest correlation present in PyC1QL4-1, the gene encoding Complement C1Q-like protein 4, C1QL4. In addition, the correlation between PyC1QL4-1 expression and PST toxicity was also the highest. Further analysis in another aquaculture scallop (Chlamys farreri) indicated that the expression of CfC1QL4-1, the homolog of PyC1QL4-1, also exhibited significant correlations with both PST toxicity and concentration. Our results reveal the gene expression responses of scallop digestive glands to PST-producing algae and indicate that the C1QL4-1 gene might be a potential biomarker for PST monitoring in scallops, which may provide a convenient way for the early warning and sensitive detection of PST contamination in the bivalves.

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.

Occurrence and spatial distribution of paralytic shellfish toxins in seawater and marine organisms in the coastal waters of Qinhuangdao, China

In recent years, paralytic shellfish toxins (PSTs) have been prevalent in the coastal waters of Qinhuangdao, the west coast of the Bohai Sea, China. The content of PSTs in shellfish often exceeded the regulatory limit of 800 μg STX equivalent (eq.) kg−1, which poses a serious threat to human health. In this study, two surveys were conducted in May 2021 and May 2022 to investigate the distribution of PSTs in the coastal waters of Qinhuangdao. Seawater, surface sediment, phytoplankton, zooplankton, and other marine organism samples were collected, and the composition and concentration of PSTs were analyzed by liquid chromatography with tandem mass spectrometry (LC-MS/MS). Results showed that multiple PST components were detected in all seawater samples collected at different depths, mainly including GTX1/4, GTX2/3, dcGTX2, STX and C1/2, and the highest concentration of PSTs reached 244 ng STX eq. L−1. The sediment samples also contained low levels of C1/2 and GTX2/3. Trace amounts of C1/2 and GTX1-4 were detected in phytoplankton and zooplankton. Moreover, all bivalve shellfish samples were found to contain PSTs, and the scallop Azumapecten farreri and the ark clam Anadara kagoshimensis showed relatively high concentrations of 607 and 497 μg STX eq. kg−1, respectively. In addition, low levels of PSTs were also found in some non-traditional PST vectors, including whelk Rapana venosa, octopus Amphioctopus ovulum, goby Ctenotrypauchen chinensis, and greenling Hexagrammos agrammus. Results of this study improve the understanding of the distribution of PSTs in seawater and marine organisms and the potential risk of persistent PSTs in seawater to marine ecosystems and human health.

Changes in Toxin Production, Morphology and Viability of Gymnodinium catenatum Associated with Allelopathy of Chattonella marina var. marina and Gymnodinium impudicum.

Allelopathy between phytoplankton organisms is promoted by substances released into the marine environment that limit the presence of the dominating species. We evaluated the allelopathic effects and response of cell-free media of Chattonella marina var. marina and Gymnodinium impudicum in the toxic dinoflagellate Gymnodinium catenatum. Additionally, single- and four-cell chains of G. catenatum isolated from media with allelochemicals were cultured to evaluate the effects of post exposure on growth and cell viability. Cell diagnosis showed growth limitation and an increase in cell volume, which reduced mobility and led to cell lysis. When G. catenatum was exposed to cell-free media of C. marina and G. impudicum, temporary cysts and an increased concentration of paralytic shellfish toxins were observed. After exposure to allelochemicals, the toxin profile of G. catenatum cells in the allelopathy experiments was composed of gonyautoxins 2/3 (GTX2/3), decarcarbamoyl (dcSTX, dcGTX2/3), and the sulfocarbamoyl toxins (B1 and C1/2). A difference in toxicity (pg STXeq cell−1) was observed between G. catenatum cells in the control and those exposed to the filtrates of C. marina var. marina and G. impudicum. Single cells of G. catenatum had a lower growth rate, whereas chain-forming cells had a higher growth rate. We suggest that a low number of G. catenatum cells can survive the allelopathic effect. We hypothesize that the survival strategy of G. catenatum is migration through the chemical cloud, encystment, and increased toxicity.

Paralytic shellfish toxins associated with Arctic Tern mortalities in Alaska

Harmful algal blooms produce biotoxins that can injure or kill fish, wildlife, and humans. These blooms occur naturally but have intensified in many locations globally due to recent climatic changes, including ocean warming. Such changes are especially pronounced in northern regions, where the effects of paralytic shellfish toxins (PSTs) on marine wildlife are of growing concern. In Alaska, seabird mortality events have increased in frequency, magnitude, and duration since 2015 alongside anomalously high ocean temperatures. Although starvation has been implicated as the apparent cause of death in many of these die-offs, saxitoxin (STX) and other PSTs have been identified as possible contributing factors. Here, we describe a mortality event at a nesting colony of Arctic Terns (Sterna paradisaea) near Juneau, Alaska in 2019 and report elevated concentrations of PSTs in bird, forage fish, and mussel samples. Concentrations of STX and other PSTs in tern tissues (2.5–51.2 µg 100g−1 STX-equivalents [STX-eq]) were of similar magnitude to those reported from other PST-induced bird die-offs. We documented high PST concentrations in blue mussels (>11,000 µg 100g−1 STX-eq; Mytilus edulis spp.) collected from nearby beaches, as well as in forage fish (up to 494 µg 100g−1 STX-eq) retrieved from Arctic Tern nests, thereby providing direct evidence of PST exposure via the terns’ prey. At maximum concentrations measured in this study, a single 5 g Pacific Sand Lance (Ammodytes personatus) could exceed the median lethal STX dose (LD50) currently estimated for birds, offering strong support for PSTs as a likely source of tern mortality. In addition to describing this localized bird mortality event, we used existing energetics data from adult and nestling Arctic Terns to calculate estimated cumulative daily PST exposure based on ecologically relevant concentrations in forage fish. Our estimates revealed potentially lethal levels of PST exposure even at relatively low (≤30 ug 100g−1 STX-eq) toxin concentrations in prey. These findings suggest that PSTs present a significant hazard to Arctic Terns and other northern seabirds and should be included in future investigations of avian mortality events as well as assessments of population health.

Development of a Method for Detecting Alexandrium pacificum Based on the Quantification of sxtA4 by Chip-Based Digital PCR.

Alexandrium pacificum, which produces the paralytic shellfish toxin (PST) saxitoxin (STX), is one of the causative species of paralytic shellfish poisoning outbreaks in coastal areas of Korea. In this study, we developed a chip-based digital PCR (dPCR) method for A. pacificum detection and tested it for monitoring in Jinhae-Masan Bay. Using the sequence of an A. pacificum strain isolated in 2017, species-specific primers targeting sxtA4 (a STX biosynthesis-related gene) were designed and used in a dPCR, detecting 2.0 ± 0.24 gene copies per cell of A. pacificum. Cell abundance in field samples, estimated by a chip-based dPCR, was compared with the PST content, and measured using a mouse bioassay. A comparison with shellfish PST concentrations indicated that cell concentrations above 500 cells L−1, as measured using the dPCR assay, may cause shellfish PST concentrations to exceed the allowed limits for PSTs. Concordance rates between dPCR and PST results were 62.5% overall in 2018–2021, reaching a maximum of 91.7% in 2018–2019. The sensitivity of the dPCR assay was higher than that of microscopy and sxtA4-based qPCRs. Absolute quantification by chip-based dPCRs targeting sxtA4 in A. pacificum exhibits potential as a complementary approach to mouse bioassay PST monitoring for the prevention of toxic blooms.

Performance of different extraction methods for paralytic shellfish toxins and toxin stability in shellfish during storage.

Accurate analysis of paralytic shellfish toxins (PSTs) in shellfish is important to protect seafood safety and human health. In this study, the performance of different extraction protocols for PSTs from scallop tissues is compared and discussed, including regular extraction solvents hydrochloric acid (HCl) and acetic acid (AcOH) followed by heating and solid-phase extraction (SPE) purification, and a novel technique of matrix solid-phase dispersion (MSPD) without heating. The possible conversion of C1/2 and GTX2/3 standards after heating, and the stability of PSTs in wet scallop tissues stored at -20°C for a 6-month period are also explored. Results showed that the MSPD technique could effectively mitigate matrix interference, but its recoveries of PSTs were significantly lower than those of the HCl and AcOH extraction methods followed by carbon SPE purification. The molar concentrations of M-toxins obtained by the MSPD method were generally lower than those analyzed by the HCl and AcOH extraction methods, which demonstrated a weak chemical conversion of C1/2 and GTX2/3 due to the heating process. Most of the PSTs were relatively stable in scallop tissues during 1-month storage at -20°C, while the concentrations of PSTs in scallop tissues obviously changed after 6 months due to the degradation and transformation of PSTs during long-term storage at -20°C. This work helps improve our understanding of the performance of different extraction methods and the stability of PSTs in scallop tissues stored at -20°C.

Paralytic shellfish toxins in Peruvian scallops associated with blooms of Alexandrium ostenfeldii (Paulsen) Balech & Tangen in Paracas Bay, Peru

In recent years, dense Alexandrium ostenfeldii blooms have been reported in different coastal areas. In this study, we report for the first time the occurrence of A. ostenfeldii blooms associated with the detection of paralytic shellfish toxins (PSTs) in the Peruvian scallop (Argopecten purpuratus) from Paracas Bay. Alexandrium ostenfeldii blooms occurred at the end of summer and early fall, after the increase of riverine input and under stratified conditions following a decrease in wind velocity. The highest abundances occurred during warm sea surface temperatures (18–27 °C). High PST concentrations that exceed the maximum permissible level (800 μg STX eq. kg−1) occurred even under low A. ostenfeldii abundances (20 × 103 cells l−1). Our results contribute to a better understanding of the dynamics of A. ostenfeldii in coastal systems influenced by riverine inputs and upwelling and can be used to improve monitoring programs and allow the implementation of mitigation measures along the Peruvian coast.

Generalized linear models to assess environmental drivers of paralytic shellfish toxin blooms (Southeast Tasmania, Australia)

Paralytic shellfish toxin (PST) events are often difficult to predict because synergistic processes, involving multiple temporal and spatial scales, are responsible for their development. In this study, we consider a suite of factors, from local to regional processes, driving paralytic shellfish toxin (PST) events associated with Alexandrium catenella on the east coast of Tasmania (Australia). PST concentrations in shellfish were compared against environmental variables, including daily wind speed and direction, satellite chlorophyll-a, sea surface temperature and river discharge. Relationships were tested using generalized linear models (GLMs). GLM coefficients confirm that high PST events were significantly favored by cooler temperatures and higher chlorophyll-a concentrations. PST events were seasonal, occurring between August and November and showed a significant relationship with sustained coastal upwelling and wind transition events after one to three months of predominantly upwelling winds. Satellite imagery shows evidence of cross shelf transport moderating PST events. Overall, the results suggest that a combination of factors (currents, upwelling conditions and changes in sea surface temperature) work together to initiate and moderate PST events. The use of GLMs provides a useful tool for understanding synergistic relationships. These techniques are of value for developing reliable predictive models to forecast and manage the effects of harmful and costly PST events.

Harmful algal blooms and their effects in coastal seas of Northern Europe

Harmful algal blooms (HAB) are recurrent phenomena in northern Europe along the coasts of the Baltic Sea, Kattegat-Skagerrak, eastern North Sea, Norwegian Sea and the Barents Sea. These HABs have caused occasional massive losses for the aquaculture industry and have chronically affected socioeconomic interests in several ways. This status review gives an overview of historical HAB events and summarises reports to the Harmful Algae Event Database from 1986 to the end of year 2019 and observations made in long term monitoring programmes of potentially harmful phytoplankton and of phycotoxins in bivalve shellfish. Major HAB taxa causing fish mortalities in the region include blooms of the prymnesiophyte Chrysochromulina leadbeateri in northern Norway in 1991 and 2019, resulting in huge economic losses for fish farmers. A bloom of the prymesiophyte Prymnesium polylepis (syn. Chrysochromulina polylepis) in the Kattegat-Skagerrak in 1988 was ecosystem disruptive. Blooms of the prymnesiophyte Phaeocystis spp. have caused accumulations of foam on beaches in the southwestern North Sea and Wadden Sea coasts and shellfish mortality has been linked to their occurrence. Mortality of shellfish linked to HAB events has been observed in estuarine waters associated with influx of water from the southern North Sea. The first bloom of the dictyochophyte genus Pseudochattonella was observed in 1998, and since then such blooms have been observed in high cell densities in spring causing fish mortalities some years. Dinoflagellates, primarily Dinophysis spp., intermittently yield concentrations of Diarrhetic Shellfish Toxins (DST) in blue mussels, Mytilus edulis, above regulatory limits along the coasts of Norway, Denmark and the Swedish west coast. On average, DST levels in shellfish have decreased along the Swedish and Norwegian Skagerrak coasts since approximately 2006, coinciding with a decrease in the cell abundance of D. acuta. Among dinoflagellates, Alexandrium species are the major source of Paralytic Shellfish Toxins (PST) in the region. PST concentrations above regulatory levels were rare in the Skagerrak-Kattegat during the three decadal review period, but frequent and often abundant findings of Alexandrium resting cysts in surface sediments indicate a high potential risk for blooms. PST levels often above regulatory limits along the west coast of Norway are associated with A. catenella (ribotype Group 1) as the main toxin producer. Other Alexandrium species, such as A. ostenfeldii and A. minutum, are capable of producing PST among some populations but are usually not associated with PSP events in the region. The cell abundance of A. pseudogonyaulax, a producer of the ichthyotoxin goniodomin (GD), has increased in the Skagerrak-Kattegat since 2010, and may constitute an emerging threat. The dinoflagellate Azadinium spp. have been unequivocally linked to the presence of azaspiracid toxins (AZT) responsible for Azaspiracid Shellfish Poisoning (AZP) in northern Europe. These toxins were detected in bivalve shellfish at concentrations above regulatory limits for the first time in Norway in blue mussels in 2005 and in Sweden in blue mussels and oysters (Ostrea edulis and Crassostrea gigas) in 2018. Certain members of the diatom genus Pseudo-nitzschia produce the neurotoxin domoic acid and analogs known as Amnesic Shellfish Toxins (AST). Blooms of Pseudo-nitzschia were common in the North Sea and the Skagerrak-Kattegat, but levels of AST in bivalve shellfish were rarely above regulatory limits during the review period. Summer cyanobacteria blooms in the Baltic Sea are a concern mainly for tourism by causing massive fouling of bathing water and beaches. Some of the cyanobacteria produce toxins, e.g. Nodularia spumigena, producer of nodularin, which may be a human health problem and cause occasional dog mortalities. Coastal and shelf sea regions in northern Europe provide a key supply of seafood, socioeconomic well-being and ecosystem services. Increasing anthropogenic influence and climate change create environmental stressors causing shifts in the biogeography and intensity of HABs. Continued monitoring of HAB and phycotoxins and the operation of historical databases such as HAEDAT provide not only an ongoing status report but also provide a way to interpret causes and mechanisms of HABs.

Uptake of Paralytic Shellfish Toxins by Blacklip Abalone (Haliotis rubra rubra Leach) from direct exposure to Alexandrium catenella microalgal cells and toxic aquaculture feed

The Tasmanian abalone fishery represents the largest wild abalone resource in the world, supplying close to 25% of the annual wild-caught global harvest. Prompted by the need to manage Paralytic Shellfish Toxin (PST) contamination of Blacklip Abalone (Haliotis rubra rubra) from east coast Tasmania, the uptake of toxins by this species is investigated in a land-based, controlled aquaculture setting. Abalone were exposed to either live Alexandrium catenella microalgal cultures or PST contaminated feed pellets during a 28 day exposure period and toxins quantified in viscera, foot muscle and epipodium tissues. PST profiles of abalone foot tissues were dominated by saxitoxin and neosaxitoxin, whilst viscera more closely resembled those of the toxin source (A. catenella cells rich in gonyautoxin 1&4 and 2&3 or feed pellets containing A. catenella extracts rich in these analogues). This indicates direct uptake of PST in the viscera via browsing/grazing on the pellet and /or sedimented microalgal cells. After exposure to A. catenella cell culture, PST concentrations in the foot (muscle + epipodium) were on average 8 times higher than in the viscera. Higher toxicity of foot tissue was caused by higher PST content of the epipodium (up to 1,085 µg STX.2HCl equiv. kg−1), which despite its small contribution to total animal weight significantly added to the overall toxin burden. Higher PST levels in the abalone foot suggest that toxin monitoring programmes may not need to routinely analyse both foot and viscera, potentially allowing for a 50% reduction of analytical costs. This option is being further investigated with continuing field studies.

Multiple New Paralytic Shellfish Toxin Vectors in Offshore North Sea Benthos, a Deep Secret Exposed.

In early 2018, a large easterly storm hit the East Anglian coast of the UK, colloquially known as the ‘Beast from the East’, which also resulted in mass strandings of benthic organisms. There were subsequent instances of dogs consuming such organisms, leading to illness and, in some cases, fatalities. Epidemiological investigations identified paralytic shellfish toxins (PSTs) as the cause, with toxins present in a range of species and concentrations exceeding 14,000 µg STX eq./kg in the sunstar Crossaster papposus. This study sought to better elucidate the geographic spread of any toxicity and identify any key organisms of concern. During the summers of 2018 and 2019, various species of benthic invertebrates were collected from demersal trawl surveys conducted across a variety of locations in the North Sea. An analysis of the benthic epifauna using two independent PST testing methods identified a ‘hot spot’ of toxic organisms in the Southern Bight, with a mean toxicity of 449 µg STX eq./kg. PSTs were quantified in sea chervil (Alcyonidium diaphanum), the first known detection in the phylum bryozoan, as well as eleven other new vectors (>50 µg STX eq./kg), namely the opisthobranch Scaphander lignarius, the starfish Anseropoda placenta, Asterias rubens, Luidia ciliaris, Astropecten irregularis and Stichastrella rosea, the brittlestar Ophiura ophiura, the crustaceans Atelecyclus rotundatus and Munida rugosa, the sea mouse Aphrodita aculeata, and the sea urchin Psammechinus miliaris. The two species that showed consistently high PST concentrations were C. papposus and A. diaphanum. Two toxic profiles were identified, with one dominated by dcSTX (decarbamoylsaxitoxin) associated with the majority of samples across the whole sampling region. The second profile occurred only in North-Eastern England and consisted of mostly STX (Saxitoxin) and GTX2 (gonyautoxin 2). Consequently, this study highlights widespread and variable levels of PSTs in the marine benthos, together with the first evidence for toxicity in a large number of new species. These findings highlight impacts to ‘One Health’, with the unexpected sources of toxins potentially creating risks to animal, human and environmental health, with further work required to assess the severity and geographical/temporal extent of these impacts.

Change in Paralytic Shellfish Toxins in the Mussel Mytilus galloprovincialis Depending on Dynamics of Harmful Alexandrium catenella (Group I) in the Geoje Coast (South Korea) during Bloom Season.

Paralytic shellfish toxins (PSTs) produced by Alexandrium catenella (formerly A. tamarense) in Korean coastal waters caused the deaths of four people (in 1986 and 1996) who consumed contaminated mussels (Mytilus edulis). This led to more detailed consideration of the risks of PST outbreaks and incidents in Korea, including the introduction of shellfish collection bans. In this study, we investigated the relationships between A. catenella population dynamics and PST accumulation in the mussel M. galloprovincialis. Discharges from the Nakdong River affect the environmental conditions along the Geoje coast, resulting in low salinity and high nutrient levels that trigger blooms of A. catenella. At the toxin peak on 24 April 2017, the toxins detected in A. catenella cells were C1, gonyautoxin (GTX)1 and GTX2, whereas the concentrations of PSTs in M. galloprovincialis were high and in the order of GTX4 > GTX1 > GTX3 > saxitoxin (STX) > GTX2 > neoSTX > decarbamoylgonyautoxin (dcGTX)2 > dc GTX3. The PST level in mussels was also high. At 15 °C, the PSTs are constantly found to be higher (10-fold higher in 2017 and 30-fold higher in 2018) than safe levels for human consumption (80 μg STX diHCl equivalents 100 g−1).

Glutathione S-transferase genes in scallops and their diverse expression patterns after exposure to PST-producing dinoflagellates

The glutathione S-transferases (GSTs) are a superfamily of enzymes that function in cellular protection against toxic substances and oxidative stress. Bivalves could accumulate high concentration of paralytic shellfish toxins (PSTs) from harmful algae. To understand the possible involvement of GSTs in protecting bivalves during PST accumulation and metabolism, the GST genes were systemically analyzed in two cultured scallops, Azumapecten farreri and Mizuhopecten yessoensis, which were reported for PST deposition during harmful algae bloom. A total of 35 and 37 GSTs were identified in A. farreri (AfGSTs) and M. yessoensis (MyGSTs) genome, respectively, and the expansion of the sigma class from the cytosolic subfamily was observed. In both scallop species, sigma class GSTs showed higher expression than other members. The high GSTs expression was detected mainly during/after larvae stages and in the two most toxic organs, hepatopancreas and kidney. After ingesting PST-producing dinoflagellates, all the regulated AfGSTs in the hepatopancreas were from the sigma class, but with opposite regulation pattern between Alexandrium catenella and A. minutum exposure. In scallop kidneys, where PSTs transformed into higher toxicity, more AfGSTs were regulated than in the hepatopancreas, and most of them were from the sigma class, with similar regulation pattern between A. catenella and A. minutum exposure. In M. yessoensis exposed to A. catenella, MyGST-σ2 was the only up-regulated MyGST in both hepatopancreas and kidney. Our results suggested the possible diverse function of scallop GSTs and the importance of sigma class in the defense against PSTs, which would contribute to the adaptive evolution of scallops in marine environments.

The Southeast Alaska Tribal Ocean Research (SEATOR) Partnership: Addressing Data Gaps in Harmful Algal Bloom Monitoring and Shellfish Safety in Southeast Alaska.

Many communities in Southeast Alaska harvest shellfish such as mussels and clams as an important part of a subsistence or traditional diet. Harmful algal blooms (HABs) of phytoplankton such as Alexandrium spp. produce toxins that can accumulate in shellfish tissues to concentrations that can pose a hazard for human health. Since 2013, several tribal governments and communities have pooled resources to form the Southeast Alaska Tribal Ocean Research (SEATOR) network, with the goal of minimizing risks to seafood harvest and enhancing food security. SEATOR monitors toxin concentrations in shellfish and collects and consolidates data on environmental variables that may be important predictors of toxin levels such as sea surface temperature and salinity. Data from SEATOR are publicly available and are encouraged to be used for the development and testing of predictive algorithms that could improve seafood risk assessment in Southeast Alaska. To date, more than 1700 shellfish samples have been analyzed for paralytic shellfish toxins (PSTs) in more than 20 locations, with potentially lethal concentrations observed in blue mussels (Mytilus trossulus) and butter clams (Saxidomus gigantea). Concentrations of PSTs exhibit seasonality in some species, and observations of Alexandrium are correlated to sea surface temperature and salinity; however, concentrations above the threshold of concern have been found in all months, and substantial variation in concentrations of PSTs remain unexplained.

First Detection of Paralytic Shellfish Toxins from Alexandrium pacificum above the Regulatory Limit in Blue Mussels (Mytilus galloprovincialis) in New South Wales, Australia.

In 2016, 2017 and 2018, elevated levels of the species Alexandrium pacificum were detected within a blue mussel (Mytilus galloprovincialis) aquaculture area at Twofold Bay on the south coast of New South Wales, Australia. In 2016, the bloom persisted for at least eight weeks and maximum cell concentrations of 89,000 cells L−1 of A. pacificum were reported. The identity of A. pacificum was confirmed using molecular genetic tools (qPCR and amplicon sequencing) and complemented by light and scanning electron microscopy of cultured strains. Maximum reported concentrations of paralytic shellfish toxins (PSTs) in mussel tissue was 7.2 mg/kg PST STX equivalent. Elevated cell concentrations of A. pacificum were reported along the adjacent coastal shelf areas, and positive PST results were reported from nearby oyster producing estuaries during 2016. This is the first record of PSTs above the regulatory limit (0.8 mg/kg) in commercial aquaculture in New South Wales since the establishment of routine biotoxin monitoring in 2005. The intensity and duration of the 2016 A. pacificum bloom were unusual given the relatively low abundances of A. pacificum in estuarine and coastal waters of the region found in the prior 10 years.

Paralytic Shellfish Toxins (PST)-Transforming Enzymes: A Review.

Paralytic shellfish toxins (PSTs) are a group of toxins that cause paralytic shellfish poisoning through blockage of voltage-gated sodium channels. PSTs are produced by prokaryotic freshwater cyanobacteria and eukaryotic marine dinoflagellates. Proliferation of toxic algae species can lead to harmful algal blooms, during which seafood accumulate high levels of PSTs, posing a health threat to consumers. The existence of PST-transforming enzymes was first remarked due to the divergence of PST profiles and concentrations between contaminated bivalves and toxigenic organisms. Later, several enzymes involved in PST transformation, synthesis and elimination have been identified. The knowledge of PST-transforming enzymes is necessary for understanding the processes of toxin accumulation and depuration in mollusk bivalves. Furthermore, PST-transforming enzymes facilitate the obtainment of pure analogues of toxins as in natural sources they are present in a mixture. Pure compounds are of interest for the development of drug candidates and as analytical reference materials. PST-transforming enzymes can also be employed for the development of analytical tools for toxin detection. This review summarizes the PST-transforming enzymes identified so far in living organisms from bacteria to humans, with special emphasis on bivalves, cyanobacteria and dinoflagellates, and discusses enzymes’ biological functions and potential practical applications.

Detection of Paralytic Shellfish Toxins in Southern Rock Lobster Jasus edwardsii Using the Qualitative Neogen™ Lateral Flow Immunoassay: Single-Laboratory Validation.

Paralytic shellfish toxins (PST) are a significant problem for the Tasmanian shellfish and Southern Rock Lobster (Jasus edwardsii) industries, and the introduction of a rapid screening test in the monitoring program could save time and money. The aim was to perform a single-laboratory validation of the Neogen rapid test for PST in the hepatopancreas of Southern Rock Lobster. The AOAC INTERNATIONAL guidelines for the validation of qualitative binary chemistry methods were followed. Three different PST profiles (mixtures) were used, of which two were commonly found in naturally contaminated lobster hepatopancreas (high in gonyautoxin 2&3 and saxitoxin), and the third toxin profile was observed in a few select animals (high in gonyautoxin 1&4). The Neogen test consistently returned negative results for non-target toxins (selectivity). The probability of detection (POD) of PST in the lobster hepatopancreas using the Neogen test increased with increasing PST concentrations. POD values of 1.0 were obtained at ≥0.57 mg STX-diHCl eq/kg in mixtures 1 and 2, and 0.95 and 1.0 for mixture 3 at 0.79 and 1.21 mg STX-diHCl eq/kg, respectively, with a fitted POD of 0.98 for 0.80 mg STX-diHCl eq/kg. The performance of the Neogen test when using four different production lots (ruggedness) showed no significant differences. The results of the validation study were satisfactory and the Neogen test is being trialed within the Tasmanian PST monitoring program of Southern Rock Lobster. The Neogen rapid kit was successfully validated for the detection of PST in Southern Rock Lobster hepatopancreas.