Paralytic Shellfish Poisoning Toxin Content Research Articles

Impact of elevated pCO2 on paralytic shellfish poisoning toxin content and composition in Alexandrium tamarense

Ocean acidification is considered a major threat to marine ecosystems and may particularly affect primary producers. Here we investigated the impact of elevated pCO2 on paralytic shellfish poisoning toxin (PST) content and composition in two strains of Alexandrium tamarense, Alex5 and Alex2. Experiments were carried out as dilute batch to keep carbonate chemistry unaltered over time. We observed only minor changes with respect to growth and elemental composition in response to elevated pCO2. For both strains, the cellular PST content, and in particular the associated cellular toxicity, was lower in the high CO2 treatments. In addition, Alex5 showed a shift in its PST composition from a non-sulfated analogue towards less toxic sulfated analogues with increasing pCO2. Transcriptomic analyses suggest that the ability of A. tamarense to maintain cellular homeostasis is predominantly regulated on the post-translational level rather than on the transcriptomic level. Furthermore, genes associated to secondary metabolite and amino acid metabolism in Alex5 were down-regulated in the high CO2 treatment, which may explain the lower PST content. Elevated pCO2 also induced up-regulation of a putative sulfotransferase sxtN homologue and a substantial down-regulation of several sulfatases. Such changes in sulfur metabolism may explain the shift in PST composition towards more sulfated analogues. All in all, our results indicate that elevated pCO2 will have minor consequences for growth and elemental composition, but may potentially reduce the cellular toxicity of A. tamarense.

The combined effect of salinity and temperature on the growth and toxin content of four Chilean strains of Alexandrium catenella (Whedon and Kofoid) Balech 1985 (Dinophyceae) isolated from an outbreak occurring in southern Chile in 2009

The toxic dinoflagellate Alexandrium catenella has been detected in the southern Chile since 1972, causing severe negative impacts on public health and aquaculture activities. Several environmental factors have been determined to affect growth and toxin production in Alexandrium strains. The aim of this study was to determine the effect of four combined conditions of two temperatures (10 and 15°C) and two salinities (15 and 35 psu) on the growth and the Paralytic Shellfish Poisoning (PSP) toxin content and composition in four Chilean strains of A. catenella (PFB41, PFB42, PFB37 and PFB38), isolated during a summer outbreak occurred in southern Chile in 2009. The growth curves showed a higher effect of the salinity in strains PFB41 and PFB42 than in strains PFB37 and PFB38. The values of growth rates and maximum cell densities ranged from 0.25 to 0.73 div day−1 and 1.1×104 to 5.2×104 cells mL−1, respectively. All of the strains showed the highest values for both growth parameters at 15°C and 35 psu. In general, growth parameters were higher at 35 psu independently of the temperature. On the other hand, the total PSP toxin content ranged widely from 3.99 to 239fmol cell−1. The highest values of PSP toxin content were attained at 10°C and 35 psu for all of the strains, at both stages of growth. All of the strains displayed different toxin compositions, with neoSTX, GTX4-1, GTX3-2 and GTX5 being the main toxins detected. The results showed significant differences in the absolute values of growth and toxin production parameters among the strains grown under the same culture conditions, and for each strain grown under different combined conditions of temperature and salinity. These findings demonstrate that abiotic factors can differentially affect the population dynamics of the A. catenella toxic genotypes, thus making it extremely difficult to predict the ecological behavior of this species in the field in terms of the intensity of a potential outbreak.

Molecular discrimination of toxic and non-toxic Alexandrium species (Dinophyta) in natural phytoplankton assemblages from the Scottish coast of the North Sea

Molecular methods provide promising tools for routine detection and quantification of toxic microalgae in plankton samples. To this end, novel TaqMan minor groove binding probes and primers targeting the small (SSU) or large (LSU) ribosomal subunit (rRNA) were developed for two species of the marine dinoflagellate genus Alexandrium (A. minutum, A. tamutum) and for three groups/ribotypes of the A. tamarense species complex: Group I/North American (NA), Group II/Mediterranean (ME) and Group III/Western European (WE). Primers and probes for real-time quantitative PCR (qPCR) were species-specific and highly efficient when tested in qPCR assays for cross-validation with pure DNA from cultured Alexandrium strains. Suitability of the qPCR assays as molecular tools for the detection and estimation of relative cell abundances of Alexandrium species and groups was evaluated from samples of natural plankton assemblages along the Scottish east coast. The results were compared with inverted microscope cell counts (Utermöhl technique) of Alexandrium spp. and associated paralytic shellfish poisoning (PSP) toxin concentrations. The qPCR assays indicated that A. tamarense (Group I) and A. tamutum were the most abundant Alexandrium taxa and both were highly positively correlated with PSP toxin content of plankton samples. Cells of A. tamarense (Group III) were present at nearly all stations but in low abundance. Alexandrium minutum and A. tamarense (Group II) cells were not detected in any of the samples, thereby arguing for their absence from the specific North Sea region, at least at the time of the survey. The sympatric occurrence of A. tamarense Group I and Group III gives further support to the hypothesis that the groups/ribotypes of the A. tamarense species complex are cryptic species rather than variants belonging to the same species.

Grazer-induced toxin formation in dinoflagellates: a transcriptomic model study

The cosmopolitan marine dinoflagellate Alexandrium minutum is known to produce toxins causing paralytic shellfish poisoning (PSP). As has recently been shown, this toxicity can be enhanced by the presence of certain copepod species. Inducible defences are known from a variety of marine organisms, but the associated transcriptomic changes have only been investigated in model angiosperms. Here, we use a microarray approach to investigate the changes in gene expression during the copepod-provoked induction of higher toxicity in A. minutum. We found a limited set of 14 genes (0.35% of the tested sequences) to be affected by copepod presence; gene expression changed by a factor of 1.9 to 11.4 when compared with control samples. We compared the differentially expressed genes after one day – before the onset of augmented toxicity – and after 3 days, when cellular toxin content in the induced samples was five times higher than in control cultures. Comparison with differences previously found between toxic and non-toxic strains revealed two sequences apparently associated with PSP toxin content. Our study is the first investigation of transcriptomic effects associated with grazer-induced induction of elevated toxicity in a protist. This provides a novel approach to the ecology of predator–prey interactions, as well as a highly targeted method for investigating the genes associated with toxin production and regulation in toxic dinoflagellates.

Implications of life‐history transitions on the population genetic structure of the toxigenic marine dinoflagellate Alexandrium tamarense

Genotypic or phenotypic markers for characterization of natural populations of marine microalgae have typically addressed questions regarding differentiation among populations, usually with reference to a single or few clonal isolates. Based upon a large number of contemporaneous isolates from the same geographical population of the toxigenic species Alexandrium tamarense from the North Sea, we uncovered significant genetic substructure and low but significant multilocus linkage disequilibrium (LD) within the planktonic population. Between the alternative molecular genotyping approaches, only amplified fragment length polymorphism (AFLP) revealed cryptic genetic population substructure by Bayesian clustering, whereas microsatellite markers failed to yield concordant patterns. Both markers, however, gave evidence for genetic differentiation of population subgroups as defined by AFLP. A considerable portion of multilocus LD could be attributed to population subdivision. The remaining LD within population subgroups is interpreted as an indicator of frequency shifts of clonal lineages during vegetative growth of planktonic populations. Phenotypic characters such as cellular content and composition of neurotoxins associated with paralytic shellfish poisoning (PSP) and allelochemical properties may contribute to intra- or inter-annual differentiation of planktonic populations, if clonal lineages that express these characters are selectively favoured. Nevertheless, significant phenotypic differentiation for these characters among the genetically differentiated subgroups was only detected for PSP toxin content in two of the four population subgroups. By integrating the analysis of phenotypic and genotypic characteristics, we developed a conceptual population genetic model to explain the importance of life-cycle dynamics and transitions in the evolutionary ecology of these dinoflagellates.

Uptake of paralytic shellfish poisoning and spirolide toxins by paddle crabs ( Ovalipes catharus) via a bivalve vector

The uptake of paralytic shellfish poisoning (PSP) toxins and spirolides by the paddle crab ( Ovalipes catharus) was investigated in two laboratory feeding trials using Greenshell™ mussels ( Perna canaliculus), which had been fed toxic strains of either Alexandrium catenella or A. ostenfeldii, as a vector. Toxin uptake by crabs occurred in both feeding trials and was limited to the visceral tissue; no toxins were detected in the body meat or the gills. The first trial utilized a strain of A. catenella that had high total PSP toxin content, 442.3 ± 91.6 fmol/cell, that was dominated by low toxicity N-sulfocarbamoyl toxins resulting in a low cellular toxicity, 5.5 ± 1.6 pg STXequiv./cell. In this trial, toxin accumulation in the crabs was highly variable and ranged from 3.8 to 221.5 μg STXequiv./100 g, with 3/4 of the crabs exceeding the regulatory limit of 80 μg STXequiv./100 g. Eight days after feeding on toxic mussels the crabs still retained high levels of toxin suggesting that depuration rates in this species may be slow. In the second feeding trial, the A. ostenfeldii strain fed to mussels produced low levels of both PSP toxins (52.0 ± 19.5 fmol/cell; 1.4 ± 0.3 pg STXequiv./cell) and spirolides (1.8 pg/cell) and, as a result, the concentration transferred to crabs via the mussels was very low-PSP toxins ranged from 2.5 to 6.8 μg STXequiv./100 g and spirolides from 6 to 7 μg/kg. The results of our study demonstrate that paddle crabs are capable of acquiring both PSP toxins and spirolides and suggest that this may occur in the wild during a toxic shellfish event. It also highlights the need to remove the viscera before consumption.

Toxin profile of Alexandrium catenella from the Chilean coast as determined by liquid chromatography with fluorescence detection and liquid chromatography coupled with tandem mass spectrometry

The profile of tetrahydropurine neurotoxins associated with paralytic shellfish poisoning (PSP) was determined from a Chilean strain of the marine dinoflagellate Alexandrium catenella. The toxin composition was compared with that of toxic shellfish, presumably contaminated by natural blooms of A. catenella from the same region in southern Chile. Ion pair-liquid chromatography with post-column derivatization and fluorescence detection (LC-FD) was employed for relative quantitative analysis of the toxin components, whereas unambiguous identification of the toxins was confirmed by tandem mass spectrometry (LC–MS/MS). In the dinoflagellate strain from Chile, the N-sulfocarbamoyl derivatives (C1/C2, B1) and the carbamoyl gonyautoxins GTX1/GTX4 comprise >90% of the total PSP toxin content on a molar basis. This toxin composition is consistent with that determined for A. catenella populations from the Pacific coast in the northern hemisphere. The characteristic toxin profile is also reflected in the shellfish, but with evidence of epimerization and metabolic transformations of C1 and C2 to GTX2 and GTX3, respectively. This work represents the first unequivocal identification and confirmation of such PSP toxin components from the Chilean coast.

Feeding of the tintinnid ciliate Favella taraikaensis on the toxic dinoflagellate Alexandrium tamarense and the fate of prey toxins

To establish the fate of paralytic shellfish poisoning (PSP) toxins in planktonic foodwebs, feeding activities of the planktonic ciliate Favella taraikaensis on the PSP-producing alga Alexandrium tamarense were examined in laboratory experiments and the toxin content in the ciliates was quantified by fluorometric high-performance liquid chromatography. In six experiments, F. taraikaensis actively fed on A. tamarense and grew rapidly. The maximum specific growth and ingestion rates of F. taraikaensis on a diet of A. tamarense (>300 cells ml−1) ranged from 0.7 day−1 to 1.5 day−1 and from 3.1 cells individual−1 h−1 to 5.8 cells individual−1 h−1 respectively. The PSP toxin content in F. taraikaensis grown on A. tamarense was low and the individual toxin content of F. taraikaensis was lower than the cellular toxin content of A. tamarense. The PSP toxins of A. tamarense were therefore scarcely accumulated within F. taraikaensis during the feeding process.

Erratum to Bacterial diversity in toxic Alexandrium tamarense blooms off the Orkney Isles and the Firth of Forth

The genetic diversity of the bacterial community associated with Alexandrium tamarense blooms was studied in blooms of the toxic dinoflagellates in the waters around the Orkney Isles and the Firth of Forth (Scotland). For toxin and molecular analysis of the bacterial communities associated with the toxic bloom, water samples were taken in 1998 and 1999 from A. tamarense blooms. The bacterial community structure, as determined by DGGE (denaturing gradient gel electrophoresis) showed clear differences between all three investigated size fractions (dinoflagellate-associated bacteria, attached bacteria and free-living bacteria), with high diversity within each sample. DNA sequence analysis of the dominant and most frequent DGGE bands revealed the dominance of α Proteobacteria, mainly of the Roseobacter clade, with similarities of 91–99%. Moreover, DGGE bands occurring at the same position in the gel throughout in most samples corroborate the presence of several specific α Proteobacteria of the Roseobacter clade. Overall, 500 bacteria were isolated from the bloom and partly phylogenetically analysed. They were members of two prokaryotic phyla, the Proteobacteria and the Bacteroidetes, related to Proteobacteria of the α and γ subdivisions (Alteromonas, Pseudoalteromonas and Colwellia). All bacteria were tested for the production of sodium channel blocking (SCB) toxins using mouse neuroblastoma assay. No production of SCB toxins was found and high performance liquid chromatography (HPLC) analysis confirmed these results. The content of total paralytic shellfish poisoning (PSP) toxin in the water samples, as measured within the toxic dinoflagellate blooms using HPLC, ranged from 53 to 2191 ng PSP l−1 in 1998 and from 0 to 478 ng PSP l−1 in 1999. Changes in PSP toxin content were not accompanied by changes of DGGE band patterns. We therefore presume that the bacterial groups identified in this study were not exclusively associated with toxic A. tamarense, but were generally associated with the phytoplankton.

Feeding and reproductive responses of marine copepods in South China Sea to toxic and nontoxic phytoplankton

The influences of the toxic and nontoxic dinoflagellate Alexandriumtamarense and the diatom Thalassiosiraweissflogii on the feeding and reproductive responses of the calanoid copepods Calanussinicus and Paracalanuscrassirostris were examined. Experiments were carried out to investigate the functional feeding response of copepods at different food concentrations for toxic and nontoxic A.tamarense and diatoms. Egg production of the copepods was compared at the same cell concentration (0.21 mg C l–1) for the three algal diets. The results demonstrated that the two copepods were able to feed on toxic dinoflagellates at rates similar to or higher than the rates with nontoxic dinoflagellates. Maximum ingestion rates were recorded at a food carbon concentration of 0.5 mg C l–1. The toxic and nontoxic dinoflagellates were radiolabeled with 109Cd and 75Se, respectively, and fed to the copepods as a mixture. The ratio of the two radiotracers in the copepods after feeding was comparable to the ratio measured in the cell mixture, indicating that the copepods were not able to select the particles based on the paralytic shellfish poisoning toxin content of the cells within a short exposure period. Copepods fed with the toxic dinoflagellates produced on average 17–46% fewer eggs than the animals fed with the nontoxic dinoflagellates and diatoms, although there was no significant difference among the three algal treatments due to the great variation among different individuals. The hatching success of eggs produced by copepods fed on the three diets and the larval development were, however, comparable. A field study was conducted to determine the grazing pressure of copepods on phytoplankton during a massive dinoflagellate bloom dominated by Prorocentrumdentatum in summer. Our calculations indicated that copepod grazing was able to remove <10% of the phytoplankton standing stocks under these conditions.

Copepod grazing selection and particle discrimination on the basis of PSP toxin content

MEPS Marine Ecology Progress Series Contact the journal Facebook Twitter RSS Mailing List Subscribe to our mailing list via Mailchimp HomeLatest VolumeAbout the JournalEditorsTheme Sections MEPS 181:163-176 (1999) - doi:10.3354/meps181163 Copepod grazing selection and particle discrimination on the basis of PSP toxin content Gregory J. Teegarden* Graduate School of Oceanography, University of Rhode Island, Narragansett, Rhode Island 02882, USA *E-mail: gregt@gsosun1.gso.uri.edu ABSTRACT: Omnivorous copepods are capable of discriminatory feeding using mechano- and chemosensory mechanisms. The presence of phycotoxins in phytoplankton often results in reduced consumption of such potential prey by copepods, though it has not been clear if this is the result of discriminatory feeding by either tactile (mechanosensory) or chemosensory recognition of toxic prey, or perhaps a physiological response to ingested neurotoxic compounds. In this study, experiments were performed to determine whether 3 species of marine copepods (Acartia tonsa, Centropages hamatus, and Eurytemora herdmani) that commonly co-occur with toxic Alexandrium spp. dinoflagellates were capable of discriminating between cultured Alexandrium spp. strains on the basis of paralytic shellfish poisoning (PSP) toxin content, i.e. by chemosensory means, using live fluorescently labeled cells. Additional experiments investigated whether toxic cells in mixtures with non-toxic alternate species of dinoflagellates affected either prey selection or total carbon consumption rates of copepods, and whether daily carbon rations could be maintained on both toxic and non-toxic Alexandrium spp. monoculture diets. Results indicated that all 3 copepod species could discriminate between toxic and non-toxic Alexandrium spp. cells by chemosensory means, suggesting that selective behavior, rather than physiological effects, governs the grazing response of copepods exposed to toxic prey. Prey selection in mixtures of several dinoflagellate species depended on whether the Alexandrium spp. cells present were toxic or non-toxic. C. hamatus and E. herdmani (but not A. tonsa) maintained daily carbon rations despite the presence of toxic Alexandrium spp., chiefly through increased consumption of alternate prey. For A. tonsa and C. hamatus, carbon rations were not equivalent between toxic and non-toxic Alexandrium spp. monoculture diets, indicating strong aversions to PSP toxins, and the potential for physiological effects when no other food is available. In all experiments feeding behavior varied among copepod species, suggesting that grazing pressure on toxic Alexandrium spp. is not uniform throughout the zooplankton community. The grazer-deterrent effects observed have implications for the function of PSP toxins. KEY WORDS: Alexandrium · Copepod · Grazing behavior · PSP toxins Full text in pdf format PreviousNextExport citation RSS - Facebook - Tweet - linkedIn Cited by Published in MEPS Vol. 181. Publication date: May 18, 1999 Print ISSN:0171-8630; Online ISSN:1616-1599 Copyright © 1999 Inter-Research.

A modified HPLC method for analysis of PSP toxins in algae and shellfish from china

Improvements to an established HPLC method are introduced. The modified method is more efficient for separation and detection of the toxins responsible for paralytic shellfish poisoning (PSP). The PSP toxin content of two strains of Alexandrium tamarense and approximately forty shellfish samples collected from different locations in China have been analyzed with this HPLC method. Only one shellfish sample, collected from Lianyungang, China, contained PSP toxins.

Use of high-performance liquid chromatography to investigate the production of paralytic shellfish toxins by Protogonyaulax spp. in culture

Procedures have been developed for the extraction and high-performance liquid chromatography (HPLC) analysis of paralytic shellfish poisoning (PSP) toxins from Protogonyaulax spp. grown in batch culture. Using these procedures, the toxin content of two isolates of P. tamarensis (NEPCC 183 and 255) and one isolate of P. catenella (NEPCC 355) were examined. Total toxin and individual toxin concentrations were measured for each isolate during the exponential and stationary phases of growth in batch culture. The total toxicity of each isolate as measured by HPLC analysis was found to agree with toxicity as determined by the standard mouse bioassay. Two of the isolates (255 and 355) were found to be toxic and the third (183) was non-toxic. The toxic isolates (255 and 355) both showed higher average total PSP toxin content during the exponential phase (35 and 23 fmol toxin cell-1, respectively) than during the stationary phase (21 and 8 fmol toxin cell-1, respectively). These cultures differed dramatically in their toxin composition. P. tamarensis (255) contained a large proportion of the N(21) sulfo toxins (B1, B2, C1, C2) while P. catenella (355) contained primarily Gonyautoxins 1 through 4. The percent composition of individual toxins was found to be constant throughout the growth cycle for both toxic isolates, even though the total toxin concentration varied. Our results suggest that PSP toxin profiles might be useful as chemotaxonomic indicators.