Toxic Alexandrium Research Articles

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 (6.31fmolcell−1STXequiv.) and Alexandrium fundyense (9.56fmolcell−1STXequiv.) isolated from Wickford Cove; the toxicity of a Narragansett Bay Alexandrium peruvianum isolate (1.79fmolcell−1STXequiv.) was also determined. Combined Alexandrium tamarense/Alexandrium fundyense abundance in Wickford Cove reached a peak abundance of 1280cellsL−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.

Termination of a toxic Alexandrium bloom with hydrogen peroxide

The dinoflagellate Alexandrium ostenfeldii is a well-known harmful algal species that can potentially cause paralytic shellfish poisoning (PSP). Usually A. ostenfeldii occurs in low background concentrations only, but in August of 2012 an exceptionally dense bloom of more than 1millioncellsL−1 occurred in the brackish Ouwerkerkse Kreek in The Netherlands. The A. ostenfeldii bloom produced both saxitoxins and spirolides, and is held responsible for the death of a dog with a high saxitoxin stomach content. The Ouwerkerkse Kreek routinely discharges its water into the adjacent Oosterschelde estuary, and an immediate reduction of the bloom was required to avoid contamination of extensive shellfish grounds. Previously, treatment of infected waters with hydrogen peroxide (H2O2) successfully suppressed cyanobacterial blooms in lakes. Therefore, we adapted this treatment to eradicate the Alexandrium bloom using a three-step approach. First, we investigated the required H2O2 dosage in laboratory experiments with A. ostenfeldii. Second, we tested the method in a small, isolated canal adjacent to the Ouwerkerkse Kreek. Finally, we brought 50mgL−1 of H2O2 into the entire creek system with a special device, called a water harrow, for optimal dispersal of the added H2O2. Concentrations of both vegetative cells and pellicle cysts declined by 99.8% within 48h, and PSP toxin concentrations in the water were reduced below local regulatory levels of 15μgL−1. Zooplankton were strongly affected by the H2O2 treatment, but impacts on macroinvertebrates and fish were minimal. A key advantage of this method is that the added H2O2 decays to water and oxygen within a few days, which enables rapid recovery of the system after the treatment. This is the first successful field application of H2O2 to suppress a marine harmful algal bloom, although Alexandrium spp. reoccurred at lower concentrations in the following year. The results show that H2O2 treatment provides an effective emergency management option to mitigate toxic Alexandrium blooms, especially when immediate action is required.

The influence of bloom intensity on the encystment rate and persistence of Alexandrium minutum in Cork Harbor, Ireland

Toxic Alexandrium minutum blooms recur annually in Cork Harbor, Ireland where they initiate in an inlet known as the North Channel. The dynamics of these blooms have been studied since 2003, and a high degree of inter-annual variability in the cell densities has been observed. Two intense blooms, with maximum cell densities >500,000cellsL−1, were observed in the summers of 2004 and 2011. Annual cyst surveys during winter found that cyst densities decreased after the 2004 bloom, and by 2010 an average of ca. 40 cystsg dry wt sediment−1 was recorded. The intensity of blooms was found to be independent of the cyst density measured the previous winter. The cyst input to the sediment during both intense and low density blooms was measured directly through the deployment of sediment traps in the North Channel. The data allowed an estimate of the proportion of the A. minutum vegetative cells that underwent successful encystment, which averaged at 2.5% across a range of cell densities spanning three orders of magnitude. Maturation times of fresh cysts were determined at 5, 10 and 15°C. The maturation time at 15°C was found to be approximately 5 months, a value which increased by two months for a 5° decrease in temperature. A cyst dynamics model was constructed based on the field data to simulate the temporal variation of A. minutum cysts in the oxic layer of sediment. It revealed that a degree of resuspension is required to prevent cyst stocks from becoming exhausted in the thin oxic layer at the surface of the sediment. The model also demonstrated that the cysts supplied by periodic intense blooms, which occur with a frequency of every 7–8 years, are not in themselves enough to allow the population to persist over long time scales (decades). The cyst input from interim blooms of lower density is however enough to ensure the annual inoculation of the water column with A. minutum cells.

Tracing the origin of paralytic shellfish toxins in scallop Patinopecten yessoensis in the northern Yellow Sea

Some dinoflagellate species within the genera Alexandrium, Gymnodinium and Pyrodinium are well-known producers of paralytic shellfish toxins (PST), which led to many poisoning incidents around the world. In the northern Yellow Sea, an important mariculture zone for scallop Patinopecten yessoensis, PST have been frequently detected from scallops. However, there is little knowledge concerning PST-producing microalgae in this region so far. In cruises carried out in 2011 and 2012, scallop and phytoplankton samples were collected from the northern Yellow Sea. PST were detected from scallops by high-performance liquid chromatography with fluorescence detection (HPLC-FLD). Toxin content and profile were remarkably different among the four tissues, i.e. viscera, adductor muscle, mantle and gonad, suggesting apparent toxin transfer and transformation in scallops. Viscera always had the highest content of PST dominated by low-potency N-sulfocarbamoyl toxins C1 and C2, which closely resembled the toxin profiles of net-concentrated phytoplankton samples in spring. Based on the morphological features, cells of Alexandrium spp. in net-concentrated phytoplankton samples were picked out and a partial sequence of the large subunit ribosomal RNA gene (LSU rDNA) was amplified using a single-cell polymerase chain reaction (PCR) method. Cells of both toxic A. tamarense species complex and non-toxic A. affine were identified from the phytoplankton samples based on the partial LSU rDNA sequence information. According to these findings, it is implied that A. tamarense species complex is the major toxic species related to PST contamination in scallops of the northern Yellow Sea. The presence of both toxic and non-toxic Alexandrium spp. in this region requires for a species-specific method to monitor the distribution and dynamics of A. tamarense species complex.

A feasibility study into the production of a freeze-dried oyster reference material for paralytic shellfish poisoning toxins

Matrix reference materials are an essential component for the validation and quality control of analytical methodologies for the quantitation of marine biotoxins in shellfish. Given the potential advantages of reference materials in powder form, a study was conducted to assess the feasibility for the production of a freeze-dried oyster tissue reference material containing a range of important paralytic shellfish poisoning toxins. One bulk sample of a wet oyster tissue homogenate was generated following mass culturing of toxic Alexandrium and oyster feeding experiments. The bulk tissue was used to prepare untreated wet frozen aliquots with the remainder being freeze-dried and processed into appropriately-sized powder samples. A pre-column oxidation LC-FLD analysis was used to confirm the absence of any chromatographic artefacts resulting from the processing and to confirm acceptable homogeneity of the tissues. Excellent stability over both the short-term (1 month) and long-term (1 year) of the freeze-dried material was demonstrated as compared with the stability of the untreated wet tissue. A post-column oxidation LC-FLD method was used to confirm the absence of toxin epimerisation in freeze-dried tissues which were observed in the wet tissues. Overall the work showed the feasibility of an approach to produce a homogenous freeze-dried oyster matrix material with enhanced stability in comparison to the untreated wet tissue. The potential for use of the process for preparation of large scale production batches of a freeze-dried CRM for paralytic shellfish poisoning toxins has therefore been demonstrated.

Abundance and distribution of toxic Alexandrium tamarense resting cysts in the sediments of the Chukchi Sea and the eastern Bering Sea

Abundance and distribution of the toxic dinoflagellate Alexandrium tamarense species complex resting cyst were investigated in the eastern Bering Sea and the Chukchi Sea for the first time. Sediment samples (top 0–3cm depth) were collected from the continental shelf of the eastern Bering Sea (17 stations) and the Chukchi Sea (13 stations) together with a long core sample (top 0–21cm depth) from one station in the Chukchi Sea during 2009–2012. The cysts were enumerated using the primuline staining method. Species identification of the cysts was carried out with multiplex PCR assay and the plate morphology of vegetative cells germinated from cysts in the both areas. Alexandrium cysts were widely detected in the both areas, ranging from not detected (<1cystscm−3) to 835cystscm−3 wet sediment in the eastern Bering Sea and from not detected (<1cystscm−3) to 10,600cystscm−3 in the Chukchi Sea, and all isolated cysts were genetically and morphologically identified as the North American clade A. tamarense. Their cysts were mainly distributed in the shallow continental shelf where the water depth was less than 100m in both areas. The cysts were detected from the deep layer (18–21cm depth of sediment core) of the long core sample. The present study confirmed the abundant existence of A. tamarense with wide range of distribution in these areas. This fact suggests that A. tamarense vegetative cells have appeared in the water column in the both areas. Furthermore, these abundant cyst depositions indicate that this species originally distributed in the Arctic and subarctic regions and well adapted to the environments in the marginal ice zone.

Influences of sedimentation and hydrodynamics on the spatial distribution of Alexandrium catenella/tamarense resting cysts in a shellfish farming lagoon impacted by toxic blooms

Since resting cysts are a potential seeding source for blooms, the presence of these cysts in sediments is a marker of an established population for a number of harmful algal species. The spatial patterns of cyst density in relation to sediment characteristics and hydrodynamics are still largely misunderstood. This study investigated the spatial distribution of resting cysts belonging to the Alexandrium tamarense species complex (Dinophyceae) in sediments of a Mediterranean coastal lagoon (Thau Lagoon, France). This lagoon, hosting shellfish farming, is regularly impacted by toxic Alexandrium catenella blooms. The average cyst density across the whole lagoon was rather low, <20cystsg−1 of dry sediment (DS). However, densities varied widely among sampled stations, with the highest density (∼440cystsg−1 DS) recorded in a shallow cove named Crique-de-l’Angle, which is the only area where dense blooms of A. catenella and A. tamarense have been recorded in the years preceding this survey. An analysis using spatial autoregressive models demonstrated that cyst densities were highly spatially autocorrelated (indicating that close stations tended to have more similar cyst densities) with accumulation sites. With respect to sediment characteristics (5 granulometric fractions <2mm and biochemical components), the highest densities were found in silty sediments containing high proportions of water and organic matter. Nevertheless, the linkage between cyst density and sediment structure was not always verified; this reflected the influence of hydrodynamics on the sedimentation of cysts and sediment particles, and on the dispersal of cysts away from the bloom area by wind-induced currents, suggesting that hydrodynamics was responsible for the spatially autocorrelated distribution of cyst densities.

Transformation of paralytic shellfish poisoning toxins in UK surf clams (Spisula solida) for targeted production of reference materials

The periodic occurrence of Paralytic Shellfish Poisoning (PSP) toxins in UK surf clams and the recent move away from biological assays for PSP testing resulted in the need to determine method performance characteristics for the replacement analytical method in this species. With the requirement for laboratory reference materials to aid this validation together with known issues relating to toxin transformation in live clams and homogenised tissue, there was the need to assess the toxin transformation characteristics of PSP toxins in surf clam tissue. Initial work examined the rates of toxin transformation in UK surf clam tissue incubated with toxin standards, showing rapid transformation of N-sulfocarbamoyl toxins with slower transformation of carbamate toxins. Full transformational pathways were determined using a combination of three different analytical methods and confirmed the major expected transformations involving decarbamoylation, with some evidence for additional reaction pathways. Results obtained from the analysis of surf clam and oyster tissues incubated with varying concentrations of toxic Alexandrium algae highlighted expected transformation reactions, although significant differences were observed in the extent of the transformations amongst the range of toxins studied, with less efficient transformation of N-hydroxylated toxins as compared with other carbamate and N-sulfocarbamoyl toxins. Analysis of PSP-toxic incurred oyster, scallop and mussel tissues incubated with variable proportions of surf clam tissue showed large differences in the extent of the transformations. Total conversion of N-sulfocarbamoyl toxins was confirmed at low relative proportions of surf clam tissue in all three species, whereas transformation of carbamate toxins was found to occur only in the presence of higher proportions of surf clam tissue in oysters and mussels in comparison with scallops. Results enabled the production of three laboratory reference materials prepared following incubation of incurred homogenates with optimum proportions of surf clam tissue, resulting in materials containing a large number of PSP toxins. Stability experiments provided good preliminary evidence for the stability of these targeted materials under storage conditions. The work therefore provides both additional information relating to the transformational activity in UK surf clams and highlights a good potential method for the targeted production of reference materials which include a wider range of toxins than normally present in naturally incurred shellfish.

Extensive genetic diversity and rapid population differentiation during blooms of Alexandrium fundyense (Dinophyceae) in an isolated salt pond on Cape Cod, MA, USA.

In Massachusetts, paralytic shellfish poisoning (PSP) is annually recurrent along the coastline, including within several small embayments on Cape Cod. One such system, the Nauset Marsh System (NMS), supports extensive marshes and a thriving shellfishing industry. Over the last decade, PSP in the NMS has grown significantly worse; however, the origins and dynamics of the toxic Alexandrium fundyense (Balech) populations that bloom within the NMS are not well known. This study examined a collection of 412 strains isolated from the NMS and the Gulf of Maine (GOM) in 2006–2007 to investigate the genetic characteristics of localized blooms and assess connectivity with coastal populations. Comparisons of genetic differentiation showed that A. fundyense blooms in the NMS exhibited extensive clonal diversity and were genetically distinct from populations in the GOM. In both project years, genetic differentiation was observed among temporal samples collected from the NMS, sometimes occurring on the order of approximately 7 days. The underlying reasons for temporal differentiation are unknown, but may be due, in part, to life-cycle characteristics unique to the populations in shallow embayments, or possibly driven by selection from parasitism and zooplankton grazing; these results highlight the need to investigate the role of selective forces in the genetic dynamics of bloom populations. The small geographic scale and limited connectivity of NMS salt ponds provide a novel system for investigating regulators of blooms, as well as the influence of selective forces on population structure, all of which are otherwise difficult or impossible to study in the adjacent open-coastal waters or within larger estuaries.

Sensitive and rapid detection of two toxic microalgae Alexandrium by loop-mediated isothermal amplification

A loop-mediated isothermal amplification (LAMP) assay was designed and evaluated for rapid detection of the toxic microalgae Alexandrium catenella and A. minutum, which can produce paralytic shellfish poisoning (PSP). Two sets of four specific primers targeting these two species were derived from the sequence of internal transcribed spacer (ITS) of ribosomal DNA. The method worked well in less than an hour under isothermal conditions of 65°C. LAMP specificity was validated in closely related algae as a comparison, suggesting the strict specificity of the LAMP primers. Two visual inspection approaches were feasible to interpret the positive or negative results. The detection limits of A. catenella and A. minutum samples using the LAMP assay were found to be 5.6 and 4.5 pg DNA, respectively. The sensitivity of this LAMP assay was 10 or 100-fold higher than Polymerase Chain Reaction (PCR) method in detecting the two microalgae. These characteristics of species specificity, sensitivity, and rapidity suggest that this method has the potentiality in the monitoring of red tide caused by A. catenella and A. minutum.

Development of theca specific antisera for the profiling of cell surface proteins in the marine toxic dinoflagellate genus Alexandrium Halim

Revealing the profile of dinoflagellate cell surface proteins (CSPs) is a crucial step for developing molecular probes for effective identification, separation and enumeration of toxic and non-toxic dinoflagellates. This study aimed to develop theca-specific antibodies against the dinoflagellates Alexandrium affine (non-toxic) and A. tamarense (toxic) to distinguish the two species, and verify if these antibodies would enable the analysis of multiple CSPs for probing phytoplankton's nutrient physiology, and facilitating rapid detection and enumeration of these harmful algal species. Using 2-DE immunoblots, we evaluated the specificity and effectiveness of the theca-specific polyclonal antisera against two types of antigens generated from fresh or fixed whole cell and insoluble cellular fractions, respectively. Our results showed that, of the four cell surface antigens, paraformaldehyde fixed whole cell antigen derived antiserum specifically recognized weakly bound theca-associated CSPs in toxic Alexandrium strain. Using the optimized theca-specific antisera, about 187 and 110 cell surface associated antigenic spots were identified on the 2-DE immunoblots of A. affine and A. tamarense, respectively. This immunoproteomic approach is proven to be very useful for phytoplankton CSP studies, permitting a more in-depth elucidation of the relationship among nutrient condition, bloom dynamic and toxin production of the harmful algae in the marine environment.

Phylogenetic analysis guided by intragenomic SSU rDNA polymorphism refines classification of “Alexandrium tamarense” species complex

We analyzed small subunit ribosomal rRNA gene (SSU rDNA) in single-cell isolated strains of “Alexandrium tamarense”, “Alexandrium fundyense” and “Alexandrium catenella” (Atama complex) by both direct and clone-based sequencing, and found 42–50 intragenomic SSU rDNA polymorphic sites (High IRP) in some of the strains but none or one (No/Low IRP) in others. Clone sequencing of the High-IRP amplicons revealed numerous variants with 0–3.5% nucleotide differences. Phylogenetic analyses including reported and new Atama complex SSU rDNA data divided this complex into two major well-separated clades. All the High-IRP sequences obtained in this study were grouped in Clade I (High-IRP clade), which also contained strains of Atama complex previously reported from northern Asia. Clade II (No/Low-IRP clade), with no evidence of IRP except for the one polymorphic site found in one of the strains, contained a subclade (IIC) exclusively of “A. catenella” from various geographic locations and several other subclades (IIA, IIB) predominantly of “A. tamarense”. Clade I corresponded to Group I in the large subunit (LSU) rDNA-based phylogenetic tree (Lilly et al., 2007), and subclades in our Clade II corresponded to the LSU-based Groups II–V. Our IRP information further unites the many seemingly different genotypes into a coherent group (Clade I) and provides delineating boundary between this and other genotypes (Clade II) in Atama complex. Based on the currently available data, we propose that Clade I and subclade IIC represent two distinct species, while the rest of Clade II represents another one or more species. Clade I should be considered one species because (1) intragenomic rDNA variants dispersed across strains in the phylogenetic tree uniting the many different genotypes to same or closely related populations, (2) their expressed SSU rDNAs (i.e. rRNAs) are almost identical (1 out of 1700nt, 0.06% difference), and (3) the corresponding Group I in the LSU-based tree also appears to be a coherent group. Subclade IIC should be considered another species because it has a long distance from the rest of Clade II as well as Clade I. Whether the rest of Clade II (mainly European “A. tamarense” subclades) represents another one or more species requires further study. Our results show that the three original morphospecies designations are invalid, and the strains do not group based on geographic locations or whether they are toxic in general, although some subclades are predominated by part of a morphotype from a region. Applying the IRP-guided analysis to a toxic Alexandrium bloom in Long Island Sound revealed that although some undocumented Alexandrium-related and other dinoflagellate lineages co-existed, the majority of the “diverse” SSU sequences detected belonged to one single population identical to “A. fundyense” in Gulf of Maine. Our result suggests that negligence of IRP could lead to incorrect recognition of the intragenomic SSU rDNA variants as distinct genotypes, thus overestimating strain diversity of a bloom of this species complex and possibly other HAB lineages.

Molecular detection and species identification of Alexandrium (Dinophyceae) causing harmful algal blooms along the Chilean coastline

On the basis of morphological evidence, the species involved in South American Pacific coast harmful algal blooms (HABs) has been traditionally recognized as Alexandrium catenella (Dinophyceae). However, these observations have not been confirmed using evidence based on genomic sequence variability. Our principal objective was to accurately determine the species of Alexandrium involved in local HABs in order to implement a real-time polymerase chain reaction (PCR) assay for its rapid and easy detection on filter-feeding shellfish, such as mussels. For species-specific determination, the intergenic spacer 1 (ITS1), 5.8S subunit, ITS2 and the hypervariable genomic regions D1-D5 of the large ribosomal subunit of local strains were sequenced and compared with two data sets of other Alexandrium sequences. Species-specific primers were used to amplify signature sequences within the genomic DNA of the studied species by conventional and real-time PCR. Phylogenetic analysis determined that the Chilean strain falls into Group I of the tamarensis complex. Our results support the allocation of the Chilean Alexandrium species as a toxic Alexandrium tamarense rather than A. catenella, as currently defined. Once local species were determined to belong to Group I of the tamarensis complex, a highly sensitive and accurate real-time PCR procedure was developed to detect dinoflagellate presence in Mytilus spp. (Bivalvia) samples after being fed (challenged) in vitro with the Chilean Alexandrium strain. The results show that real-time PCR is useful to detect Alexandrium intake in filter-feeding molluscs. It has been shown that the classification of local Alexandrium using morphological evidence is not very accurate. Molecular methods enabled the HAB dinoflagellate species of the Chilean coast to be assigned as A. tamarense rather than A. catenella. Real-time PCR analysis based on A. tamarense primers allowed the detection of dinoflagellate DNA in Mytilus spp. samples exposed to this alga. Through the specific assignment of dinoflagellate species involved in HABs, more reliable preventive policies can be implemented.

Saxitoxin δ15N as a species-specific tracer of the source of nitrogen used by the toxin-producing dinoflagellate Alexandrium tamarense

Alexandrium tamarense Balech is a harmful, bloom-forming dinoflagellate which can produce saxitoxins (STXs), a suite of powerful neurotoxins that bioaccumulate up the food chain and can have severe economic and health impacts. Being nitrogen-rich, STX is ideal for studying the relationship between its producer and the nitrogen used for growth. In this study, we measured the δ 15 N of medium nitrate, algal cells and toxin in batch cultures of A. tamarense. In nitrate-replete conditions, cells were 1.5‰ depleted relative to the source, and STX was 1.5‰ depleted relative to the cells. In nitrogen-limiting cultures the isotopic difference between cells and STX changed as nitrate in the growth medium was depleted, indicating uncoupled toxin syn- thesis and cell growth under changing external nutrient conditions. Our results show that toxin- specific nitrogen stable isotope analysis may provide a tool that can help to identify the nitrogen sources supporting toxic Alexandrium blooms.

Dynamics of a mesoscale eddy off Cape Ann, Massachusetts in May 2005

Observations and numerical modeling indicate that a mesoscale anti-cyclonic eddy formed south of Cape Ann at the northern entrance of Massachusetts Bay (MB) during May 2005, when large river discharges in the western Gulf of Maine and two strong Nor'easters passing through the regions led to an unprecedented toxic Alexandrium fundyense bloom (red tide). Both model results and field measurements suggest that the western Maine Coastal Current separated from Cape Ann around May 7–8, and the eddy formed on around May 10. The eddy was trapped at the formation location for about a week before detaching from the coastline and moving slowly southward on May 17. Both model results and theoretical analysis suggest that the separation of the coastal current from the coast and subsequent eddy formation were initiated at the subsurface by an adverse pressure gradient between Cape Ann and MB due to the higher sea level set up by onshore Ekman transport and higher density in downstream MB. After the formation, the eddy was maintained by the input of vorticity transported by the coastal current from the north, and local vorticity generation around the cape by the horizontal gradients of wind-driven currents, bottom stress, and water density induced by the Merrimack River plume. Observations and model results indicate that the anti-cyclonic eddy significantly changed the pathway of nutrient and biota transport into the coastal areas and enhanced phytoplankton including Alexandrium abundances around the perimeter of the eddy and in the western coast of MB.

A novel approach to identifying PST tolerant copepods: An individual ingestion assay

Phenotypic tolerance of individual copepods to paralytic shellfish toxins (PSTs), as determined by fecundity, has recently been reported. In the present study, we tested whether similar phenotypic tolerance related to ingestion also exists. In short-term feeding assays, ingestion rates of individual females of the copepod Acartia hudsonica were measured on a sole diet of toxic Alexandrium fundyense as well as on a sole diet of nontoxic Alexandrium tamarense. When copepods fed on A. fundyense, the frequency distribution of ingestion rates was polymodal, consistent with the hypothesis of levels of phenotypic tolerance associated with toxin ingestion. Four distinct groups of ingestion rates on toxic algae were observed. Mean rates ranged from near zero to more than 100 cells copepod −1 h −1. In contrast, when the same individuals fed on the nontoxic A. tamarense diet, the polymodal distribution of ingestion rates was not apparent. Furthermore, group-specific mean ingestion rates, for two ingestion-defined groups, were always significantly greater on the toxic diet than they were on the nontoxic diet. These results support the hypothesis that discrete groups of grazer ingestion of toxic A. fundyense are related to PST tolerance.

Effects of toxic Alexandrium tamarense on behavior, hemocyte responses and development of brown ring disease in Manila clams

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 430:35-48 (2011) - DOI: https://doi.org/10.3354/meps09111 Effects of toxic Alexandrium tamarense on behavior, hemocyte responses and development of brown ring disease in Manila clams V. M. Bricelj1,*, Susan E. Ford2, Christophe Lambert3, Annaick Barbou3, Christine Paillard3 1Institute of Marine and Coastal Sciences, Rutgers University, New Brunswick, New Jersey 08901, USA 2Haskin Shellfish Research Laboratory, Rutgers University, Port Norris, New Jersey 08349, USA 3Institut Universitaire Européen de la Mer, LEMAR, Technopôle Brest Iroise, 29280 Plouzané, France *Email: mbricelj@marine.rutgers.edu ABSTRACT: In this study of short-term exposure of Ruditapes philippinarum from Brittany, France, to an Alexandrium tamarense isolate that produces high concentrations of paralytic shellfish toxins (PSTs), the effects of the isolate on clam fitness, toxin uptake, and the response of hemocytes (responsible for internal defense in bivalves) were examined. Adults exhibited individual variability in ­resistance to the effects of PSTs, measured by their burrowing capacity. If genetically based, this may (1) indicate that resistance is under natural selection and (2) provide a new model to advance our understanding of the molecular basis for PST resistance in bivalves, so far determined only in Mya arenaria. Juvenile clams were more vulnerable to PSTs than adults, experiencing 100% burrowing incapacitation within 1 d of exposure to A. tamarense. They also experienced growth suppression and high mortalities following 4 d of toxification. Juveniles challenged with Vibrio tapetis, the cause of brown ring disease (BRD), and exposed to A. tamarense, developed significantly fewer BRD symptoms relative to controls fed non-toxic algae, but suffered higher mortality. Adult clams exposed to A. tamarense showed a significant increase in hemocyte concentrations and a small, but significant, decrease in phagocytic activity, and no effect on hemocyte viability or other functional parameters. We speculate that the inhibitory effects on BRD progression may be attributable to toxicity of PSTs to V. tapetis, inability of juveniles to activate the shell conchiolin-deposition response, and/or an overall increase in phagocytic cells induced by A. tamarense. Harmful algae and pathogens may thus ­interact and modulate the effects of disease in bivalve populations. KEY WORDS: Ruditapes philippinarum · Paralytic shellfish toxins · Vibrio · Burrowing · Hemocytes Full text in pdf format PreviousNextCite this article as: Bricelj VM, Ford SE, Lambert C, Barbou A, Paillard C (2011) Effects of toxic Alexandrium tamarense on behavior, hemocyte responses and development of brown ring disease in Manila clams. Mar Ecol Prog Ser 430:35-48. https://doi.org/10.3354/meps09111 Export citation RSS - Facebook - Tweet - linkedIn Cited by Published in MEPS Vol. 430. Online publication date: May 26, 2011 Print ISSN: 0171-8630; Online ISSN: 1616-1599 Copyright © 2011 Inter-Research.

Evidence of selection for resistance to paralytic shellfish toxins during the early life history of soft‐shell clam, Mya arenaria, populations

This study identifies early, postmetamorphic soft‐shell clams, Mya arenaria, as the life‐history stage most susceptible to effects of blooms of paralytic shellfish poisoning (PSP) toxin–producing Alexandrium spp. Laboratory experiments used progeny from predominantly susceptible (naϊve) or resistant (annually exposed) NW Atlantic populations. Growth and survival of toxified veliger larvae did not differ from those fed nontoxic algae. In contrast, postlarvae (4–12‐mm shell length) from both populations exposed to a highly toxic Alexandrium tamarense isolate (~ 100 cells mL−1, 64–69 pg saxitoxin equivalents [STXeq] cell−1) suffered burrowing incapacitation, toxin accumulation, and mortalities within 1 week of toxin exposure. These effects were greater and occurred sooner in the naϊve population. Short‐term toxification in the laboratory caused a significant shift in the genotypic composition of this population, determined with a molecular marker for sodium‐channel resistance. Clams with the sensitive genotype were selectively eliminated relative to resistant or heterozygote clams. Ingestion of toxic cells (too large for larval capture) is thus required to elicit toxic effects. Exposure to mixed, toxic, and nontoxic algal suspensions demonstrated that adverse effects to fitness (survival and growth) were dose‐dependent, occurring only at ≥ 50 cells mL−1 of the isolate used (PR18b). Paralysis and thus increased predatory risk occurred even at 10 cells mL−1. Postlarvae &lt; 12 mm, which can co‐occur with red tides throughout the Atlantic range of M. arenaria, were more susceptible to PSP than large (&gt; 30 mm) juveniles. Natural selection for resistance in Atlantic populations will thus vary latitudinally with the timing, duration, and intensity of toxic blooms.

Tracing the influence of sewage discharge on coastal bays of Southern Vancouver Island (BC, Canada) using sedimentary records of phytoplankton

The impact of sewage and stormwater effluents on phytoplankton is investigated by comparing organic-walled dinoflagellate cyst abundance and diversity from 38 surface sediment samples, flanking southern Vancouver Island. Site locations include those directly adjacent to wastewater outfall at Clover and Macaulay Points and Saanich Peninsula, as well as from a variety of near-shore environments with differing tidal flow influences. Excellently preserved dinoflagellate cyst assemblages have been recovered and 36 cyst taxa were identified. Local assemblages are characterized by a high relative proportion (average 56%) of cysts produced by heterotrophic dinoflagellates, which is typical for regions of high primary production. Relative proportional increases of cysts from heterotrophic species with particular increases of Polykrikos kofoidii/schwartzii and Dubridinium species, known to reflect areas affected by eutrophication, occur directly adjacent to all three sewage outfalls, as well as in the more stagnant waters of Esquimalt and Victoria Harbours and at the mouth of Cadboro Bay. Further effects of an anthropogenic effluent can be seen in the relatively higher concentrations of organic carbon and the diatom production proxy, biogenic opal. Results from this study clearly indicate a much larger impact zone than predicted by a sewage effluent plume model or trends found in monitored benthic biota and sediment chemistry that evidence primary outfall affects <800 m eastward of Macaulay Point and about 200 m eastward of the Clover Point. Enhanced production of cysts from potentially toxic Alexandrium species is also observed near locations of sewage outfalls.

Allelopathic effects of Alexandrium spp. on Prorocentrum donghaiense

Effects of toxic Alexandrium species on Prorocentrum donghaiense were studied in co-cultures and filtrates in order to shed some light on the competition between the co-occurring harmful algae. The three live cultures of Alexandrium tamarense and Alexandrium minutum negatively affected the growth of P. donghaiense, while only A. minutum was affected by P. donghaiense when they were in 1:3 ratio of cells. Each species was affected by the co-occurring algae as could be observed by microscopy. The allelopathic effects observed in co-culture did not correlate with the measured content of the intracellular PSP toxins but matched the observed hemolytic properties of the culture filtrates, suggesting that certain hemolytic substances other than PSP toxins are responsible for the allelopathic effect of Alexandrium.