Use of two-dimensional gel electrophoresis to differentiate morphospecies of Alexandrium minutum, a paralytic shellfish poisoning toxin-producing dinoflagellate of harmful algal blooms

Contamination of shellfish with paralytic shellfish poisoning toxins (PST) produced by toxic harmful algal blooms (HABs) have been negatively affecting the shellfish and aquaculture industries worldwide. Therefore, accurate and early identification of toxic phytoplankton species is crucial in HABs surveillance programs that allow fish-farmers to take appropriate preventive measures in shellfish harvesting and other aquaculture activities to overcome the negative impacts of HABs on human health. The identification of toxic dinoflagellates present in the water is currently a time-consuming operation since it requires skillful taxonomists and toxicologists equipped with optical and scanning electron microscopes as well as sophisticated equipment, for example, high-performance liquid chromotography-fluorescence detection. In this paper, a two-dimensional gel electrophoresis (2-DE)-based proteomic approach was applied to discriminate between toxic and nontoxic strains of Alexandrium minutum. Variation in morphological features between toxic and nontoxic strains was minimal and not significant. Also, variation in 2-DE protein patterns within either toxic or nontoxic strains was low, but pronounced differences were detected between toxic and nontoxic strains. The most notable differences between these strains were several abundant proteins with pIs ranging from 4.8 to 5.3 and apparent molecular masses between 17.5 and 21.5 kDa. Groups of proteins, namely NT1, NT2, NT3, and NT4, were consistently found in all nontoxic strains, while T1 and T2 were prominent in the toxic strains. These specific protein spots characteristic for toxic and nontoxic strains remained clearly distinguishable irrespective of the various growth conditions tested. Therefore, they have the potential to serve as "taxonomic markers" to distinguish toxic and nontoxic strains within A. minutum. Initial studies revealed that the expression pattern of T1 was tightly correlated to toxin biosynthesis in the examined alga and may be used to serve as a potential toxin indicator.     

High CO2 promotes the production of paralytic shellfish poisoning toxins by Alexandrium catenella from Southern California waters

In many dinoflagellates, cellular toxin levels have been demonstrated to increase when growth is limited by essential nutrients such as phosphorus. Despite the recognized importance of nutrient limitation to dinoflagellate toxicity, interactions with current and future global environmental change variables have been relatively unexplored. This is a critical question, as dissolution of anthropogenic CO₂ emissions into seawater is leading to progressively lower pH values, or ocean acidification. Sea surface temperatures are concurrently increasing, a trend that is also projected to continue in the future. We conditioned a clonal culture of paralytic shellfish poisoning toxin producing Alexandrium catenella (A-11c) isolated from coastal Southern California to factorial combinations of two temperatures, two pCO₂ levels, and two phosphate concentrations for a period of eight months. Interactions between these variables influenced growth and carbon fixation rates and although these treatments only elicited minor differences in toxin profile, total cellular toxicity was dramatically affected. Cells conditioned to high pCO₂ (levels projected for year 2075) and low phosphate at low temperature (15 °C) were the most toxic, while lower pCO₂, higher phosphate levels, and warmer temperature (19 °C) alleviated this toxicity to varying degrees. Overall increased pCO₂ generally led to enhanced potency. Our results suggest that future increased ocean acidification may exacerbate the toxic threat posed by this toxic dinoflagellate, especially when combined with nutrient limitation, but that future warmer temperatures could also offset some of this enhanced toxicity.     

Effect of the endoparasite Amoebophrya sp. on toxin content and composition in the paralytic shellfish poisoning dinoflagellate Alexandrium fundyense (Dinophyceae)

Members of the Amoebophrya ceratii complex are endoparasitic dinoflagellates that parasitize a number of their dinoflagellate relatives, including toxic and/or harmful algal bloom-forming species. Despite many studies on the occurrence, prevalence, biology and molecular phylogeny of Amoebophrya spp., little attention has been given to toxin dynamics of host population following parasitism. Using Amoebophrya sp. infecting the paralytic shellfish toxin (PSP)-producing dinoflagellate Alexandrium fundyense, we addressed the following questions: (1) does parasitism by Amoebophrya sp. alter toxin content and toxin profiles of the dinoflagellate A. fundyense over the infection cycle? and (2) do parasite dinospores produced at the end of the infection cycle retain host toxins and thus potentially act as a vector to convey PSP toxin through the marine microbial food-web? Toxin time-course experiments showed that the PSP toxin contents did not vary significantly over the infection cycle, but mean toxin content for infected cultures was significantly higher than that for uninfected cultures. Host toxins were not detected in the free-living, dinospore stage of the parasite. Therefore, our results indicate that Amoebophrya sp. does not function as a vector for transferring PSP toxins to higher trophic levels. Rather, Amoebophrya infections appear to play an important role in maintaining healthy ecosystems by transforming potent toxins-producing dinoflagellates into non-toxic dinospores, representing “edible food” for consumers of the marine microbial food-web during toxic algal bloom event.     

Geographic differences in paralytic shellfish poisoning toxin profiles among Japanese populations of Alexandrium tamarense and A. catenella (Dinophyceae)

To reconsider whether toxin profile could be used as a marker for populations from different geographical areas, clonal isolates of the toxic dinoflagellates Alexandrium tamarense (Lebour) Balech and Alexandrium catenella (Whedon et Kofoid) Balech from Ofunato Bay (Iwate Prefecture), Atsumi Bay (Aichi Prefecture), Tanabe Bay (Wakayama Prefecture), Harima‐Nada (Kagawa Prefecture), Uranouchi Bay (Kochi Prefecture), Hiroshima Bay (Hiroshima Prefecture) and Yamakawa Bay (Kagoshima Prefecture), which were identified on the basis of morphotaxonomy, immunological and molecular biological techniques, were subjected to analysis of paralytic shellfish poisoning toxins by high performance liquid chromatography‐fluorometric method. All the isolates except A. tamarense OF152 from Ofunato Bay contained mainly N‐sulfocarbamoyl toxins (C1 +2) with various amounts of derivatives, and a typical north‐to‐south trend of decreasing toxicity was observed. In both A. tamarense and A. catenella, toxin profiles were rather constant within a geographical area and divergent among different geographical areas. The toxin profiles of A. tamarense from Harima‐Nada were well conserved among different bloom years. Toxin profile showed that isolates of A. tamarense from Ofunato Bay, A. tamarense from Harima‐Nada isolated in 1988 and A. catenella from Uranouchi Bay were heterogeneous. However, only two or three groups of isolates with different toxin profiles were observed in a geographical region, suggesting that several representative isolates express the genotype in a given region. These observations confirmed that toxin composition could be used as a marker to discriminate different geographical populations of these species.     

Single-cell analysis of paralytic shellfish toxins in Alexandrium tamarense by HPLC with post-column fluorescent derivatization

We developed a methodology for analyzing the C-toxin (C2) content in single Alexandrium tamarense cells; this method was based on high performance liquid chromatography (HPLC). C2 is the main paralytic shellfish toxin (PST) detected in a clonal culture of A. tamarense, which is a common causative organism in cases of paralytic shellfish poisoning in Japan. This HPLC method employs post-column fluorescent derivatization (FL). Mobile phase, column size, flow rate, reagent concentrations, and lamp type for the fluorescent detector were all optimized for the detection of C2. With this improved methodology, we could measure 1 fmol of C2 with a signal to noise ratio (S/N) = 2. Clonal heterogeneity within the toxic strain, which was maintained for 13 years after re-isolation from the original clonal culture, ranged from <1 fmol to 700 fmol cell⁻¹. This report is the first to demonstrate definitively that PST content varies on a cell-by-cell basis in a clonal culture of a dinoflagellate that causes paralytic shellfish poisoning.     

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

BioMetals - 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...     

Determination of paralytic shellfish poisoning (PSP) toxins in UK shellfish

A study was conducted to aid the interpretation of data generated by parallel testing of the qualitative Jellett Rapid Test (JRT) and the mouse bioassay (MBA) for detection of paralytic shellfish poisoning (PSP) toxins within the UK statutory shellfish biotoxin monitoring programme. A selection of stored sample extracts subjected to testing by MBA and/or JRT were further analysed by liquid chromatography with fluorescence detection (LC–FLD) to provide additional information on the concentrations of PSP toxins and toxin profiles.Results, from this study, demonstrate the potential of the JRT to effectively screen out PSP toxin negative shellfish samples and samples containing low concentrations of toxins from UK monitoring programmes. Additionally, data generated using LC–FLD highlights the potential of introducing alternative analytical techniques to completely replace the requirement for the MBA.     

A multi-phylum study of grazer-induced paralytic shellfish toxin production in the dinoflagellate Alexandrium fundyense: A new perspective on control of algal toxicity

The present study surveyed grazer-induced stimulation of paralytic shellfish toxin (PST) production by the marine dinoflagellate Alexandrium fundyense. The survey included species, known to graze upon A. fundyense, from five phyla: the protists, Polykrikos kofoidii (Dinoflagellata) and Tiarina fusus (Ciliophora), the bivalve molluscs Mytilus edulis and Mya arenaria (Mollusca), the ascidians, Molgula manhattensis and Botrylloides violaceus (Chordata), and the copepod, Eurytemora herdmani (Arthropoda). Direct (grazers in contact with cells of A. fundyense) and indirect (grazers not in contact with cells of A. fundyense) induction assays were carried out with protists and copepods. Only indirect assays were carried out with molluscs and ascidians. Indirect assays also tested whether induction of PST production occurred via kairomones or feeding-related cues. All metazoan grazers induced PST production. By contrast, neither of the two species of protistan grazer induced PST production. Direct and indirect inductions of PST production were evident for the copepod, with direct induction being significantly higher than indirect induction. Effects upon PST production by phylum, species (nested within phylum), and interactions of phylum by diet, and species by diet, were evident. When induction of PST production occurred, a kairomone effect was apparent, except for M. edulis. Similarly, feeding-related cues were evident, except for E. herdmani. An asymptotic relationship between the magnitude of indirect induction of PST production and total cell ingestion by the grazers suggests a saturation response of grazer-induced PST production.     

Characterization of saxitoxin production and release and phylogeny of sxt genes in paralytic shellfish poisoning toxin-producing Aphanizomenon gracile

The freshwater cyanobacterium Aphanizomenon gracile is one of the most widely distributed producers of the potent neurotoxins saxitoxin (STX) and its derivatives (paralytic shellfish poisoning toxins, PSP toxins). However, the phylogeny of STX biosynthesis genes and the regulation of STX production and release remain poorly studied in the genus Aphanizomenon. In this study, two A. gracile strains from Spanish freshwaters were grown in semi-continuous cultures under three temperatures (15, 20 and 28 °C) and their STX production and release were determined by Enzyme-Linked ImmunoSorbent Assay (ELISA). STX production was stable along the temperature range, with 1.4–2.3-fold shifts in biomass-standardized STX contents, and maxima of 0.22 μg equivalent STX mg⁻¹ dry weight 15.3 fg equiv STX cell⁻¹ and 15.1 μg equiv STX mg⁻¹ Chl a. The extracellular fraction was remarkably high (13.6–35.3%), not clearly affected by temperature but with nitrate-depleted medium (BG11₀) inducing a 2-fold increase in extracellular content. STX production and release were not directly related to growth rates. The 16S rRNA phylogenetic analyses in sixteen A. gracile strains from Spanish and German freshwaters showed that PSP-producing A. gracile grouped within a monospecific and highly supported cluster, together with PSP-producing Aphanizomenon sp. NH-5 and clearly separated from a monospecific Aphanizomenon flos-aquae cluster. The sixteen A. gracile strains formed also monospecific and highly supported clusters for PSP-biosynthesis genes (sxtG, sxtI, sxtH and sxtX) together with Aphanizomenon sp. NH-5. This study evidences an elevated extracellular proportion of STX in A. gracile with importance for risk assessment, and supports the identification of Aphanizomenon sp. NH-5 as A. gracile.     

Profiles of Alexandrium catenella cysts in Puget Sound sediments and the relationship to paralytic shellfish poisoning events

The magnitude of paralytic shellfish poisoning (PSP) toxins in shellfish and the geographical scope of shellfish closures in Puget Sound have increased in recent decades. PSP, monitored by the Washington Department of Health, has spread from Sequim Bay in the 1950s into central Puget Sound in the 1970s and throughout Puget Sound by the 1990s. Alexandrium catenella, the species responsible for PSP toxins, produces a benthic resting cyst that, upon germinating, can seed blooms. This study examined whether there is a relationship between profiles of cysts in the sediment and temporal variation in PSP in shellfish and if the history of the toxin's southward expansion through Puget Sound can be seen in the cyst record. To address this question, sediment cores were collected from three Puget Sound basins, Sequim Bay, Penn Cove, and Carr Inlet, and cyst profiles were determined. Activities of ²¹⁰Pb were fitted to a depth-dependent diagenetic model to date the sediment cores and determine mixing and sediment-accumulation rates. In order to compare historical variation in PSP with cyst profiles that have been altered by bioturbation, a depth and time-dependent diagenetic model was then used to predict vertical profiles of cysts that would occur under the assumption that cyst deposition rates are proportional to PSP concentration in shellfish measured over several decades at each site. The cyst profiles predicted by the model were compared to measured cyst profiles. These comparisons suggested that Alexandrium blooms and resulting PSP concentration in shellfish are more closely linked to cyst germination and deposition at some stations than at others. Sequim Bay had relatively large numbers of cysts and it is likely that the persistent toxicity here is the result of recurrent seeding from the cyst bed. Penn Cove and Carr Inlet had few cysts despite occasional blooms, suggesting that blooms are transported into those areas, perhaps from other sites of cyst germination. Sequim Bay and Penn Cove had cysts from top to bottom of the cores so it was not possible to determine the date when cysts were first introduced into these bays, but it is likely that A. catenella has been in Penn Cove since at least 1955 or for about two decades before the WDOH PSP toxicity data would indicate. The cyst profile in Carr Inlet suggested a first appearance date of 1985 that is consistent with the first appearance of PSP in shellfish in 1988.     

Effect of CO2 on growth and toxicity of Alexandrium tamarense from the East China Sea,a major producer of paralytic shellfish toxins

In recent decades, the frequency and intensity of harmful algal blooms (HABs), as well as a profusion of toxic phytoplankton species, have significantly increased in coastal regions of China. Researchers attribute this to environmental changes such as rising atmospheric CO₂ levels. Such addition of carbon into the ocean ecosystem can lead to increased growth, enhanced metabolism, and altered toxicity of toxic phytoplankton communities resulting in serious human health concerns. In this study, the effects of elevated partial pressure of CO₂ (pCO₂) on the growth and toxicity of a strain of Alexandrium tamarense (ATDH) widespread in the East and South China Seas were investigated. Results of these studies showed a higher specific growth rate (0.31 ± 0.05 day⁻¹) when exposed to 1000 μatm CO₂, (experimental), with a corresponding density of (2.02 ± 0.19) × 10⁷ cells L⁻¹, that was significantly larger than cells under 395 μatm CO₂₍control). These data also revealed that elevated pCO₂ primarily affected the photosynthetic properties of cells in the exponential growth phase. Interestingly, measurement of the total toxin content per cell was reduced by half under elevated CO₂ conditions. The following individual toxins were measured in this study: C1, C2, GTX1, GTX2, GTX3, GTX4, GTX5, STX, dcGTX2, dcGTX3, and dcSTX. Cells grown in 1000 μatm CO₂ showed an overall decrease in the cellular concentrations of C1, C2, GTX2, GTX3, GTX5, STX, dcGTX2, dcGTX3, and dcSTX, but an increase in GTX1 and GTX4. Total cellular toxicity per cell was measured revealing an increase of nearly 60% toxicity in the presence of elevated CO₂ compared to controls. This unusual result was attributed to a significant increase in the cellular concentrations of the more toxic derivatives, GTX1 and GTX4.Taken together; these findings indicate that the A. tamarense strain ATDH isolated from the East China Sea significantly increased in growth and cellular toxicity under elevated pCO₂ levels. These data may provide vital information regarding future HABs and the corresponding harmful effects as a result of increasing atmospheric CO₂.     

Variability of paralytic shellfish toxin occurrence and profiles in bivalve molluscs from Great Britain from official control monitoring as determined by pre-column oxidation liquid chromatography and implications for applying immunochemical tests

As the official control monitoring laboratory in Great Britain for the analysis of marine biotoxins in shellfish, Cefas have for the past five years conducted routine monitoring for paralytic shellfish poisoning toxins (PST) using a non-animal alternative method to the mouse bioassay reference method; a refined version of the AOAC 2005.06 pre-column oxidation liquid chromatography method. Application of this instrumental methodology has enabled the generation of data not only on the occurrence and magnitude of PST events, but also the quantitation and assessment of different PST profiles. Since implementation of the method in 2008, results have shown huge variabilities in the occurrence of PSTs, with large spatial and temporal variabilities around the coastline. Mean PST profiles were not found to correlate either with total PST content of the shellfish, the year of sampling or with a few notable exceptions, the shellfish species. Toxin profiles were found to fall into one of four distinct profile types, with one relating solely to the exclusive presence of decarbamoyl toxins in surf clams. The other profile types contained variable proportions of gonyautoxins, N-sulfocarbamoyl toxins, neosaxitoxin and saxitoxin. While some indications of geographical repeatability were noted, this was not observed for all profile types. Consequently, the application of rapid immunochemical testing methods to end product testing would need to be considered carefully given the large differences in PST congener cross-reactivities.     

Recurrent vernal presence of the toxic Alexandrium tamarense/Alexandrium fundyense (Dinoflagellata) species complex in Narragansett Bay,USA

The vernal occurrence of toxic dinoflagellates in the Alexandrium tamarense/Alexandrium fundyense species complex in an enclosed embayment of Narragansett Bay (Wickford Cove, Rhode Island) was documented during 2005 and 2009–2012. This is the first report of regular appearance of the Alexandrium fundyense/Alexandrium tamarense species complex in Narragansett Bay. Thecal plate analysis of clonal isolates using SEM revealed cells morphologically consistent with both Alexandrium tamarense Lebour (Balech) and Alexandrium fundyense Balech. Additionally, molecular analyses confirmed that the partial sequences for 18S through the D1–D2 region of 28S were consistent with the identity of the two Alexandrium species. Toxin analyses revealed the presence of a suite of toxins (C1/2, B1 (GTX-5), STX, GTX-2/3. Neo, and GTX-1/4) in both Alexandrium tamarense (6.31 fmol cell⁻¹ STX equiv.) and Alexandrium fundyense (9.56 fmol cell⁻¹ STX equiv.) isolated from Wickford Cove; the toxicity of a Narragansett Bay Alexandrium peruvianum isolate (1.79 fmol cell⁻¹ STX equiv.) was also determined. Combined Alexandrium tamarense/Alexandrium fundyense abundance in Wickford Cove reached a peak abundance of 1280 cells L⁻¹ (May of 2010), with the combined abundance routinely exceeding levels leading to shellfishing closures in other systems. The toxic Alexandrium tamarense/Alexandrium fundyense species complex appears to be a regular component of the lower Narragansett Bay phytoplankton community, either newly emergent or previously overlooked by extant monitoring programs.     

Monitoring of the toxic dinoflagellate Alexandrium catenella in Osaka Bay,Japan using a massively parallel sequencing (MPS)-based technique

Since 2002, blooms of Alexandrium catenella sensu Fraga et al. (2015) and paralytic shellfish toxicity events have occurred almost yearly in Osaka Bay, Japan. To better understand the triggers for reoccurring A. catenella blooms in Osaka Bay, phytoplankton community was monitored during the spring seasons of 2012–2015. Monitoring was performed using massively parallel sequencing (MPS)-based technique on amplicon sequences of the 18S rRNA gene. Dense blooms of A. catenella occurred every year except in 2012, however, there was no significant correlation with the environmental parameters investigated. Plankton community diversity decreased before and middle of the A. catenella blooms, suggesting that the decline in diversity could be an indicator for the bloom occurrence. The yearly abundance pattern of A. catenella cells obtained by morphology-based counting coincided with the relative sequence abundances, which supports the effectiveness of MPS-based phytoplankton monitoring.     

Assessment of sodium channel mutations in Makah tribal members of the U.S. Pacific Northwest as a potential mechanism of resistance to paralytic shellfish poisoning

The Makah Tribe of Neah Bay, Washington, has historically relied on the subsistence harvest of coastal seafood, including shellfish, which remains an important cultural and ceremonial resource. Tribal legend describes visitors from other tribes that died from eating shellfish collected on Makah lands. These deaths were believed to be caused by paralytic shellfish poisoning, a human illness caused by ingestion of shellfish contaminated with saxitoxins, which are produced by toxin-producing marine dinoflagellates on which the shellfish feed. These paralytic shellfish toxins include saxitoxin, a potent Na⁺ channel antagonist that binds to the pore region of voltage gated Na⁺ channels. Amino acid mutations in the Na⁺ channel pore have been demonstrated to confer resistance to saxitoxin in softshell clam populations exposed to paralytic shellfish toxins present in their environment. Because of the notion of resistance to paralytic shellfish toxins, the study aimed to determine if a resistance strategy was possible in humans with historical exposure to toxins in shellfish. We collected, extracted and purified DNA from buccal swabs of 83 volunteer Makah tribal members and sequenced the skeletal muscle Na⁺ channel (Nav1.4) at nine loci to characterize potential mutations in the relevant saxitoxin binding regions. No mutations of these specific regions were identified after comparison to a reference sequence. This study suggests that any resistance of Makah tribal members to saxitoxin, if present, is not a function of Nav1.4 modification, but may be due to mutations in neuronal or cardiac sodium channels, or some other mechanism unrelated to sodium channel function.     

Erratum to: Bacterial diversity in toxic<Emphasis Type="Italic"> Alexandrium tamarense</Emphasis> 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 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 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 and 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.An erratum to this article can be found at Communicated by H.-D. Franke     

Modelling paralytic shellfish toxins (PSTs) transfer and accumulation in populations of two planktonic grazers

A model was developed in this paper in order to study and compare the paralytic shellfish toxin (PST) transfer and accumulation of two different potential PST vectors in the planktonic community, the heterotrophic dinoflagellate in its red form Noctiluca scintillans and the copepod Acartia clausi. Different factors that influence the toxin transfer such as toxin synthesis, grazing on toxic and non-toxic food and population size of PST producers and vectors were considered in the model. Moreover, a laboratory experiment was conducted in order to calculate the detoxification rates of Noctiluca fed on Alexandrium catenella. According to the model results, the two grazers showed a significant difference mainly in the timing of the PST accumulation. Noctiluca exhibited a rapid response to the grazing of Alexandrium with high initial toxin accumulation, followed by a reduction to zero concentration of toxins in a period of almost two days. In contrast, Acartia showed a considerable delay in comparison to Noctiluca to accumulate the same amount of toxin in the population. This delay is linked to the slower reproduction rates that characterize the copepod. The range of the initial values used for the sensitivity analysis of the model is representative of the coastal environment of a Galician ria (embayment located at the NW of Iberian Peninsula), where the two grazers and Alexandrium may co-exist. The model for Acartia showed less sensitivity to these key parameters probably due to the time delay in accumulation of significant amount of toxins. Both grazers showed a rapid (50 h) reduction of ingested toxin, suggesting inefficiency to transfer toxins through predation in the food web.     

Paralytic shellfish toxins or spirolides? The role of environmental and genetic factors in toxin production of the Alexandrium ostenfeldii complex

Dinoflagellates of the Alexandrium ostenfeldii complex (A. ostenfeldii, A. peruvianum) are capable of producing different types of neurotoxins: paralytic shellfish toxins (PSTs), spirolides and gymnodimines, depending on the strain and its geographic origin. While Atlantic and Mediterranean strains have been reported to produce spirolides, strains originating from the brackish Baltic Sea produce PSTs. Some North Sea, USA and New Zealand strains contain both toxins. Causes for such intraspecific variability in toxin production are unknown. We investigated whether salinity affects toxin production and growth rate of 5 A. ostenfeldii/peruvianum strains with brackish water (Baltic Sea) or oceanic (NE Atlantic) origin. The strains were grown until stationary phase at 7 salinities (6–35), and their growth and toxin production was monitored. Presence of saxitoxin (STX) genes (sxtA1 and sxtA4 motifs) in each strain was also analyzed. Salinity significantly affected both growth rate and toxicity of the individual strains but did not change their major toxin profile. The two Baltic Sea strains exhibited growth at salinities 6–25 and consistently produced gonyautoxin (GTX) 2, GTX3 and STX. The two North Sea strains grew at salinities 20–35 and produced mainly 20-methyl spirolide G (20mG), whereas the strain originating from the northern coast of Ireland was able to grow at salinities 15–35, only producing 13-desmethyl spirolide C (13dmC). The effects of salinity on total cellular toxin concentration and distribution of toxin analogs were strain-specific. Both saxitoxin gene motifs were present in the Baltic Sea strains, whereas the 2 North Sea strains lacked sxtA4, and the Irish strain lacked both motifs. Thus sxtA4 only seems to be specific for PST producing strains. The results show that toxin profiles of A. ostenfeldii/peruvianum strains are predetermined and the production of either spirolides or PSTs cannot be induced by salinity changes. However, changes in salinity may lead to changed growth rates, total cellular toxin concentrations as well as relative distribution of the different PST and spirolide analogs, thus affecting the actual toxicity of A. ostenfeldii/peruvianum populations.     

中国东海和南海有害赤潮高发区麻痹性贝毒素研究

用小白鼠生物检测法和高效液相色谱法对采自浙江舟山和广东深圳海域贝类的麻痹性贝毒素进行了调查和分析,结果表明,舟山海域近岸的贝类毒素检出率为14％,染毒的贝类毒素含量不高,低于小白鼠生物检测法的测定范围;深圳近岸贝类毒素检出率为30％以上,华贵栉孔扇贝是主要的染毒贝类,有1个样品毒素含量达5．1Mu·g-1,超出安全食用标准．从深圳大亚湾华贵栉孔扇贝检测出10种麻痹性贝毒素成分,消化腺的主要毒素成分为GTXl+2和GTX5,Cl+2和GTX2+3,而剔除消化腺后其余贝组织的主要成分为neoSTX和GTX5．贝毒素主要积累在扇贝的消化腺内,消化腺含有的毒素是贝肉组织的8倍．