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Due to the increase in nanoplastics (NPs) abundance in aquatic environments, their effects on phytoplankton have aroused large research attention. In this study, 100 nm sized polystyrene NPs were chosen to investigate their effecting performance and mechanisms on a typical dinoflagellates Alexandrium tamarense. The results indicated the population growth and photosynthetic efficiencies of A. tamarense were significantly inhibited by NPs exposure, as well as the increase in cellular total carotenoids and paralytic shellfish toxins (PSTs). Meanwhile, the cellar ROS levels increased, corresponding to the increased activities or contents of multiple antioxidant components, including SOD, CAT, GPX, GR, GSH and GSSG. The transcriptional results support the physiological–biochemical results and further revealed the down-regulation of genes encoding the light reaction centers (PSI and PSII) and up-regulation of genes encoding the antioxidant components. Up-regulation of genes encoding key enzymes of the Calvin cycle and glycolytic pathway together with the TCA cycle could accelerate organic carbon and ATP production for A. tamarense cells resistant to NPs stress. Finally, more Glu and acetyl-CoA produced by the enhanced GSH cycle and the glycolytic pathway, respectively, accompanied by the up-regulation of Glu and Arg biosynthesis genes supported the increase in the PST contents under NPs exposure. This study established a data set involving physiological–biochemical changes and gene information about marine dinoflagellates responding to NPs, providing a data basis for further evaluating the ecological risk of NPs in marine environments. [ABSTRACT FROM AUTHOR]

Due to the increase in nanoplastics (NPs) abundance in aquatic environments, their effects on phytoplankton have aroused large research attention. In this study, 100 nm sized polystyrene NPs were chosen to investigate their effecting performance and mechanisms on a typical dinoflagellates Alexandrium tamarense. The results indicated the population growth and photosynthetic efficiencies of A. tamarense were significantly inhibited by NPs exposure, as well as the increase in cellular total carotenoids and paralytic shellfish toxins (PSTs). Meanwhile, the cellar ROS levels increased, corresponding to the increased activities or contents of multiple antioxidant components, including SOD, CAT, GPX, GR, GSH and GSSG. The transcriptional results support the physiological–biochemical results and further revealed the down-regulation of genes encoding the light reaction centers (PSI and PSII) and up-regulation of genes encoding the antioxidant components. Up-regulation of genes encoding key enzymes of the Calvin cycle and glycolytic pathway together with the TCA cycle could accelerate organic carbon and ATP production for A. tamarense cells resistant to NPs stress. Finally, more Glu and acetyl-CoA produced by the enhanced GSH cycle and the glycolytic pathway, respectively, accompanied by the up-regulation of Glu and Arg biosynthesis genes supported the increase in the PST contents under NPs exposure. This study established a data set involving physiological–biochemical changes and gene information about marine dinoflagellates responding to NPs, providing a data basis for further evaluating the ecological risk of NPs in marine environments.

Among all the paralytic shellfish toxins (PSTs)-producing algae, Alexandrium tamarense is one of the most widespread harmful species posing a serious threat to marine resources and human health. Therefore, it is extremely important to establish a rapid and accurate monitoring method for A. tamarense that can provide early warnings of harmful algal blooms (HABs) caused by this alga and limit the contamination due to PSTs. In this study, an ssDNA library was first obtained by whole cell systematic evolution of ligands by exponential enrichment after 18 consecutive rounds of iterative screening. After sequencing in combination with subsequent multiple alignment of sequences and secondary structure simulation, the library could be classified into 2 families, namely, Family1 and Family2, according to sequence similarity. Flow cytometry was used to test the affinity and cross-reactivity of Ata19, Ata6, Ata25 and Ata29 belonging to Family2. Ata19 was selected to be modified by truncation, through which a new resultant aptamer named as Ata19-1-1 was obtained. Ata19-1-1 with a KD of 75.16 ± 11.10 nM displayed a much higher affinity than Ata19. The specificity test showed that Ata19-1-1 has the same discrimination ability as Ata19 and can at least distinguish the target microalga from other microalgae. The observation under a fluorescence microscopy showed that the A. tamarense cells labeled with Ata19-1-1 are exhibiting bright green fluorescence and could be easily identified, factually confirming the binding of the aptamer with target cells. In summary, the aptamer Ata19-1-1 produced in this study may serve as an ideal molecular recognition element for A. tamarense, which has the potential to be developed into a novel detection method for this harmful alga in the future. [ABSTRACT FROM AUTHOR]

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The content and composition of paralytic shellfish toxins (PSTs) in Japanese basket clam (Corbicula japonica) and mussels (Mytilus galloprovincialis) from Osaka Bay, Japan, were investigated using a mouse bioassay (MBA) and hydrophilic interaction liquid chromatography-tandem mass spectrometry (HILIC-MS/MS), and the association between toxicity values of MBA and HILIC-MS/MS was verified based on research data. The overall toxicity in Japanese basket clam was lower than that in the mussel. The PSTs of Japanese basket clam and mussel consisted mainly of C1, C2, and gonyautoxins 1–4 (GTX1–4) taking toxins compositional differences as mol%. When multiplying the content of different toxins by the toxic equivalent factor (TEF), C2 and GTX1–4 accounted for more than 90% of total toxicity (MU TEF/g) based on the MU TEF score converted by TEF for the two species. The total content of C2 and GTX1–4 converted to toxicity was significantly correlated with the toxicity determined by MBA for the two species (r2 > 0.983). This study provides a suitable and ethical monitoring method to investigate toxicity in bivalves contaminated with A. tamarense by analysis of only predominant toxins, along with reducing use of MBA. [ABSTRACT FROM AUTHOR]

The effects of culture filtrate of Alexandrium tamarense on Prorocentrum donghaiense and Heterosigma akashiwo were investigated, including determination of algal density, photosynthesis, intracellular enzyme content and activity. The filtrate of A. tamarense had a stronger inhibitory effect on P. donghaiense than H. akashiwo, and the inhibitory effect decreased with higher temperature treatment of the filtrate. Instantaneous fluorescence (Ft) and maximum quantum yield of photosystem II (Fv/Fm) values of both kinds of target algae were reduced as exposed to the filtrate of A. tamarense, which proved that allelopathy could inhibit the normal operation of photosynthetic system. The increase of Malondialdehyde (MDA) content of the two kinds of target algae indicated that the cell membrane was seriously damaged by allelochemicals released by A. tamarense. The different responses of Superoxide Dismutase (SOD) and Catalase (CAT) activity in two kinds of target algae demonstrated the complexity and diversity of allelopathic mechanism. The filtrate of A. tamarense also influenced the metabolic function (ATPases) of P. donghaiense and H. akashiwo, and the influence on P. donghaiense was greater. Liquid-liquid extraction was used to extract and isolate allelochemicals from the filtrate of A. tamarense. It was found that only component I with molecular weight of 424.2573 and 434.2857 could inhibit the growth of P. donghaiense by HPLC-MS., Competing Interests: Declaration of competing interest The authors declare that they have no known competing financial interests or personal relationships that could have appeared to influence the work reported in this paper., (Copyright © 2024 Elsevier Ltd. All rights reserved.)

The bloom-forming dinoflagellate Alexandrium tamarense is one of the most important producers of paralytic shellfish poisoning toxins. Annually recurrent blooms of this dinoflagellate species is associated with the incremental nitrogen influx, especially excessive nitrate input. However, limited studies have been conducted on the toxin production and underlying molecular regulation mechanisms of A. tamarense under various nitrate (N) conditions. Therefore, toxin production and transcriptomic responses of this species were investigated. The toxin profile of A. tamarense was consistently dominated by the C2-toxins, and the cellular toxicity increased with N concentrations peaking at 9.23 ± 0.03 fmol/cell in the 883 μM N-added group. Under lower N conditions, expressions of two STX-core genes, sxtA and sxtG, were significantly down-regulated, suggesting that N regulated sxt expression and triggered responses related to toxin biosynthesis. Results of this study provided valuable insights into the ecophysiology of A. tamarense, enhancing our understanding of the occurrence of toxification events in natural environments., Competing Interests: Declaration of competing interest The authors declare that they have no known competing financial interests or personal relationships that could have appeared to influence the work reported in this paper., (Copyright © 2024 Elsevier Ltd. All rights reserved.)

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

Abstract Bacteria play an important role in preventing algal blooms and reducing their harm to the environment. To improve the algicidal activity of Pseudoalteromonas SP48 which had an inhibition effect on dinoflagellate Alexandrium tamarense, its growth medium and fermentation conditions were optimized for this bacterium. In this study, we used two steps to establish the optimum conditions. First, the proper proportion of medium was selected based on an orthogonal design. Then, the fermentation conditions were further optimized through uniform design in an enlarged 5L bioreactor. To test the algicidal ability of Pseudoalteromonas SP48 under the optimum conditions, algal cell morphology was observed by transmission electron microscopy (TEM). After the orthogonal design, we found that the optimum medium was [0.7% (m/v) tryptone, 0.2% (m/v) soluble starch, 0.2% (m/v) sucrose, 0.1% (m/v) FeSO4, and 1.2% (m/v) K2HPO4] for Pseudoalteromonas SP48 growth. Based on these results, optimum fermentation conditions were further explored in a 5L fermentation cylinder using a uniform design; the influence of variables such as incubation time, carbon type, and rotation speed were tested. The optimal fermentation conditions were fermentation time (42 hr), tryptone (1.1%), seeding volume (1.4 × 1013 cells), and rotation speed (250 r/min). Under these established optimum conditions, the biomass of strain SP48 increased by 79.2% and its lethal dose 50% (LD50) decreased by 54.0%, respectively. The TEM results showed that compared with the control group, the cell wall and cell membrane of A. tamarense were significantly damaged, and the structure and shape of the organelles were destroyed by algicidal bacteria of Pseudoalteromonas SP48. Overall, our results demonstrate that the optimized culture conditions could significantly enhance the algicidal activity of Pseudoalteromonas SP48 against a harmful dinoflagellate, such as A. tamarense. It will effectively provide a scientific foundation for both production of algicidal substances and HABs control.

Algicidal bacteria play an important role in mitigating harmful algal blooms (HABs). In the study, five bacterial strains were isolated from the East China Sea. One strain of algicidal bacterium, named DH-e, was found to selectively inhibit the motor ability of Prorocentrum donghaiense, Alexandrium tamarense (ATDH-47) and Karenia mikimotoi Hansen. Both 16S rDNA sequence analysis and morphological characteristics revealed that the algicidal DH-e bacterium belonged to Halomonas. Furthermore, results showed that the metabolites in the DH-e cell-free filtrate could kill algae directly, and the minimum inhibitory concentrations (MICs) of the bacterial metabolites on the cells of the three dinoflagellate species ranged from 35.0-70.0 µg/mL. Following short-term inhibitory tests, the dinoflagellates in mixed crude extract solution (0.7 mg/mL) ceased movement after 5 min. The algicidal mechanism of the metabolites was investigated through enzyme activities, including that of catalase (CAT), alkaline phosphatase (AKP), acetone peroxide (T-ATP) synthetase and nitrite reductase (NR). Results indicated that metabolites did not disrupt the energy or nutrient routes of the algae (P>0.05), but did initiate an increase in free radicals in the algal cells, which might explain the subsequent death of sensitive algae. Thus, the metabolites of the DH-e bacterium showed promising potential for controlling HABs. [ABSTRACT FROM AUTHOR]

Paralytic shellfish toxins (PSTs) are the major neurotoxic contaminants of edible bivalves in Japan. Tetrodotoxin (TTX) was recently detected in bivalve shellfish around the world, drawing widespread attention. In Japan, high levels of TTX were reported in the digestive gland of the scallop, Patinopecten yessoensis, in 1993; however, no new data have emerged since then. In this study, we simultaneously analyzed PSTs and TTX in scallops cultured in a bay of east Japan using hydrophilic interaction chromatography (HILIC)-MS/MS. These scallops were temporally collected from April to December 2017. The highest concentration of PSTs (182 µmol/kg, total congeners) in the hepatopancreas was detected in samples collected on May 23, lined to the cell density of the dinoflagellate, Alexandrium tamarense, in seawater around the scallops, whereas the highest concentration of TTX (421 nmol/kg) was detected in samples collected on August 22. Contrary to the previous report, temporal variation of the PSTs and TTX concentrations did not coincide. The highest concentration of TTX in the entire edible tissues was 7.3 µg/kg (23 nmol/kg) in samples obtained on August 22, which was lower than the European Food Safety Authority (EFSA)-proposed threshold, 44 µg TTX equivalents/kg shellfish meat. In addition, 12β-deoxygonyautoxin 3 was firstly identified in scallops.

The effects and interactive effects of different nitrogen (N) sources (ammonium, nitrate, and urea) and carbon dioxide (CO 2 ) concentrations were investigated on Alexandrium tamarense , a harmful marine dinoflagellate, by measuring its growth (μ), extracellular carbonic anhydrase (CA), and its toxicity to zebrafish ( Danio rerio ) embryo. The μ and CA were influenced more strongly by CO 2 concentrations rather than by N sources; significant effects of CO 2 on μ and CA were observed under low CO 2 concentration (LC) conditions compared to high CO 2 concentration (HC) conditions. The ammonium and nitrate media under LC conditions had the maximum μ and CA, which was inhibited under HC conditions. The embryotoxic effects were influenced more strongly by the N sources than by CO 2 concentrations, thus excluding the lower deformation in urea under HC conditions. Moreover, the antioxidant enzymes superoxide dismutase (SOD), glutathione peroxidase (GPX), glutathione S-transferase (GST), and catalase (CAT) were detected in normal (untreated) zebrafish embryos, and among them, the level of SOD was the highest. In summary, this study provides a clear insight for understanding the effects and interactive effects of N sources and CO 2 concentrations on the growth and toxicity of harmful dinoflagellates. [ABSTRACT FROM AUTHOR]

The eastern Bering Sea has a vast continental shelf, which contains various endangered marine mammals and large fishery resources. Recently, high numbers of toxic A. tamarense resting cysts were found in the bottom sediment surface of the eastern Bering Sea shelf, suggesting that the blooms have recently occurred. However, little is known about the presence of A. tamarense vegetative cells in the eastern Bering Sea. This study's goals were to detect the occurrence of A. tamarense vegetative cells on the eastern Bering Sea shelf and to find a relationship between environmental factors and their presence. Inter-annual field surveys were conducted to detect A. tamarense cells and environmental factors, such as nutrients, salinity, chlorophyll a, and water temperature, along a transect line on the eastern Bering Sea shelf during the summers of 2004, 2005, 2006, 2009, 2012, and 2013. A. tamarense vegetative cells were detected during every sampling year, and their quantities varied greatly from year to year. The maximum cell densities of A. tamarense observed during the summers of 2004 and 2005 were much higher than the Paralytic shellfish poisoning warning levels, which are greater than 100–1,000 cells L-1, in other subarctic areas. Lower quantities of the species occurred during the summers of 2009, 2012, and 2013. A significant positive correlation between A. tamarense quantity and water temperature and significant negative correlations between A. tamarense quantity and nutrient concentrations (of phosphate, silicate, and nitrite and nitrate) were detected in every sampling period. The surface- and bottom-water temperatures varied significantly from year to year, suggesting that water temperatures, which have been known to affect the cell growth and cyst germination of A. tamarense, might have affected the cells' quantities in the eastern Bering Sea each summer. Thus, an increase in the Bering Sea shelf's water temperature during the summer will increase the frequency and scale of toxic blooms and the toxin contamination of plankton feeders. This poses serious threats to humans and the marine ecosystem. [ABSTRACT FROM AUTHOR]

A multitemplate molecularly imprinted polymer was synthesized using fragments similar to gonyautoxins 1 and 4 including 2,4,6-triaminopyrimidine, 4-hydroxy-2-butanone, and imidazole as dummy templates by bulk polymerization. Scanning electron microscopy and infrared spectroscopy showed a porous structure and the formation of organic groups in the imprinting process of the polymer. One millilitre of 0.1 mol L−1acetic acid and 1 mL of 95% methanol were the optimized washing and eluting solutions during the molecularly imprinted solid-phase extraction. The adsorption capacity of the multitemplate molecularly imprinted polymer was lower than for the monotemplate molecularly imprinted polymer prepared in a previous study in seawater in whichAlexandrium tamarenseandAlexandrium minutumwere cultivated. This result may due to the inappropriate combination of the dummy template fragments in the synthesis resulting in the unfitness of the imprinting sites for gonyautoxins 1 and 4. In general, it is inappropriate to use fragments with the similar parts to the analyte as the dummy templates. However, this work indicated the optimal molecularly imprinted polymer for the selective extraction of gonyautoxins 1 and 4. [ABSTRACT FROM PUBLISHER]

Following heavy precipitation, we observed an intense algal bloom in the St. Lawrence Estuary (SLE) that coincided with an unusually high mortality of several species of marine fish, birds and mammals, including species designated at risk. The algal species was identified as Alexandrium tamarense and was determined to contain a potent mixture of paralytic shellfish toxins (PST). Significant levels of PST were found in the liver and/or gastrointestinal contents of several carcasses tested as well as in live planktivorous fish, molluscs and plankton samples collected during the bloom. This provided strong evidence for the trophic transfer of PST resulting in mortalities of multiple wildlife species. This conclusion was strengthened by the sequence of mortalities, which followed the drift of the bloom along the coast of the St. Lawrence Estuary. No other cause of mortality was identified in the majority of animals examined at necropsy. Reports of marine fauna presenting signs of neurological dysfunction were also supportive of exposure to these neurotoxins. The event reported here represents the first well-documented case of multispecies mass mortality of marine fish, birds and mammals linked to a PST-producing algal bloom. [ABSTRACT FROM AUTHOR]

Paralytic shellfish poisoning (PSP) poses a serious health threat in Alaska and prevents effective utilization of shellfish resources by subsistence and recreational harvesters. Substantial economic losses also affect shellfish growers during PSP events. The toxins responsible for PSP are produced by dinoflagellates in the genus Alexandrium. Despite the persistent threat posed by PSP and the long history of shellfish toxicity research, there is still confusion concerning the Alexandrium species that cause PSP in Alaska. The primary objective of this study was to identify the toxic Alexandrium species present in Alaska and to develop polymerase chain reaction (PCR) assays for use in screening phytoplankton and sediment samples. Before developing the PCR assays for this study, we evaluated published assays and many were not adequate because of primer dimer formation or because of cross-reactivity. Rather than continue to grapple with the uncertainty and inadequacy of published assays, we developed new assays for the Alexandrium species most likely to be present in Alaska. Only Alexandrium fundyense Group I and A. ostenfeldii were identified from four sampling regions from southeast Alaska to Kodiak Island, indicating that these two species are widely distributed. PCR assays for these two species were converted to quantitative (q)PCR format for use in monitoring programs. During the course of this study, we realized that a systematic evaluation of all published (∼150) Alexandrium species-specific assays would be of benefit. Toward this objective, we collated published Alexandrium PCR, qPCR, and in situ hybridization assay primers and probes that targeted the small-subunit (SSU), internal transcribed spacer (ITS/5.8S), or D1-D3 large-subunit (LSU) (SSU/ITS/LSU) ribosomal DNA genes. Each individual primer or probe was screened against the GenBank database and Alexandrium gene sequence alignments constructed as part of this study. These data were used to identify a suite of species-specific Alexandrium assays that can be recommended for evaluation by the global harmful algal bloom community. [ABSTRACT FROM AUTHOR]

Quorum sensing (QS) systems play important roles in regulating many physiological functions of microorganisms, such as biofilm formation, bioluminescence, and antibiotic production. One marine algicidal bacterium, Ponticoccus sp. PD-2, was isolated from the microalga Prorocentrum donghaiense, and its N-acyl-homoserine lactone (AHL)-mediated QS system was verified. In this study, we analyzed the AHLs profile of strain PD-2. Two AHLs, 3-oxo-C8-HSL and 3-oxo-C10-HSL, were detected using a biosensor overlay assay and GC-MS methods. Two complete AHL-QS systems (designated zlaI/ R and zlbI/ R) were identified in the genome of strain PD-2. When expressed in Escherichia coli, both zlaI and zlbI genes could each produce 3-oxo-C8-HSL and 3-oxo-C10-HSL. Algicidal activity was investigated by evaluating the inhibitory rate (IR) of microalgae growth by measuring the fluorescence of viable cells. We found that the metabolites of strain PD-2 had algicidal activity against its host P. donghaiense (IR 84.81%) and two other red tide microalgae, Phaeocystis globosa (IR 78.91%) and Alexandrium tamarense (IR 67.14%). β-cyclodextrin which binds to AHLs and inhibits the QS system reduced the algicidal activity more than 50%. This indicates that inhibiting the QS system may affect the algicidal metabolites production of strain PD-2. Our study indicated that a QS-regulated algicidal system may play a potential role in the process of red tides disintegration. QS might be a potential way to control red tides. [ABSTRACT FROM AUTHOR]

Alexandrium tamarense is a toxic harmful algal blooms (HABs) causing species, which poses great threat to human health and marine economy. In this study, we isolated an algicidal bacterium Altererythrobacter sp. LY02 towards to A. tamarense and later investigated the algicidal activity, algicidal mode, characteristics of algicidal active substance and algicidal procedure. The results indicated that the cell-free filtrate of strain LY02 showed high algicidal effect on algal growth, however, bacterial cells almost lost algicidal activity. The algicidal active substance was temperature- and pH-stability, and its molecular weight was less than 1000 Da, and was a non-proteinaceous material or non-polysaccharide, mid-polar substance. Under the algicidal effect of active substance, the morphology and structure of A. tamarense cells were seriously damaged as well as organelles. Our study confirmed that the algicidal active substance could be used as an excellent bio-agent for controlling HABs caused by A. tamarense. [ABSTRACT FROM AUTHOR]

A small flagellate alga was isolated from the phycosphere of a toxic red tide dinoflagellate Alexandrium tamarense. Phylogenetic analysis and ultrastructural observations demonstrated that the samll flagellate alga is a species belong to Ochrophyte Ochromonas sp. The process of ingesting bacteria by Ochromonas sp. was recorded by a time lapse capture under a light microscope. Through the use of different assemblages in the co-culture experiment, the species interactions in this phycosphere microenvironment were analyzed. We demonstrated that the growth of Ochromonas sp. was supported by bacteria. Three strains of bacteria ingested by Ochromonas sp. were isolated and identified to belong to α-, δ- and γ-Proteobacteria. The growth of A. tamarense was suppressed when co-cultured with bacteria. In contrast, Ochromonas sp. triggered the growth of A. tamarense by inhibiting the growth of algicidal bacteria. This result firstly demonstrated a positive effect of a flagellate on a dinoflagellate in the phycosphere of A. tamarense. Combined with other negative effects between dinoflagellates and bacteria or bacteria and flagellates, this study showed a series of clear interactions among dinoflagellate, bacterium, and flagellate in the dinoflagellate microenvironment.

Blooms of the dinoflagellate Alexandrium tamarense have become worldwide phenomena and have detrimental impacts on aquatic ecosystems and human health. In this study, a culture supernatant of the marine actinomycete BS01 exerted a strong algicidal effect on A. tamarense (ATGD98-006). The target algicide from BS01 was separated by adsorption chromatography and identified by MALDI-TOF-MS and NMR analysis. The results suggested that the purified algicidal component corresponded to a hydrophobic compound (2-isobutoxyphenyl)amine (C10H15NO) with a molecular weight of 165 Da, which exhibited a significant algicidal effect (64.5%) on A. tamarense. After incubation in 5 μg/mL of (2-isobutoxyphenyl)amine for 24 h, the algae lost mobility and sank to the bottom of the flasks, and 56.5% of the algae cells lost vitality at a concentration of 20 μg/mL (p < 0.01) despite having intact cell profiles. Morphological analysis revealed that the cell structure of A. tamarense was altered by (2-isobutoxyphenyl)amine resulting in cytoplasm degradation and the loss of organelle integrity. The images following propidium iodide staining suggested that the algal nucleus was also severely damaged and eventually degraded due to exposure to the algicidal compound. All of the results indicate that (2-isobutoxyphenyl)amine from the actinomycete might be a candidate for the control of bloom-forming A. tamarense.

Members of the marine dinoflagellate genus Alexandrium are known to exude allelochemicals, unrelated to well-known neurotoxins (PSP-toxins, spirolides), with negative effects on other phytoplankton and marine grazers. Physico/chemical characterization of extracellular lytic compounds of A. tamarense, quantified by Rhodomonas salina bioassay, showed that the lytic activity, and hence presumably the compounds were stable over wide ranges of temperatures and pH and were refractory to bacterial degradation. Two distinct lytic fractions were collected by reversed-phase solid-phase extraction. The more hydrophilic fraction accounted for about 2% of the whole lytic activity of the A. tamarense culture supernatant, while the less hydrophilic one accounted for about 98% of activity. Although temporal stability of the compounds is high, substantial losses were evident during purification. Lytic activity was best removed from aqueous phase with chloroform-methanol (3:1). A “pseudo-loss” of lytic activity in undisturbed and low-concentrated samples and high activity of an emulsion between aqueous and n-hexane phase after liquid-liquid partition are strong evidence for the presence of amphipathic compounds. Lytic activity in the early fraction of gel permeation chromatography and lack of activity after 5 kD ultrafiltration indicate that the lytic agents form large aggregates or macromolecular complexes.

Harmful algal blooms occur throughout the world, threatening human health and destroying marine ecosystems. Alexandrium tamarense is a globally distributed and notoriously toxic dinoflagellate that is responsible for most paralytic shellfish poisoning incidents. The culture supernatant of the marine algicidal bacterium BS02 showed potent algicidal effects on A. tamarense ATGD98-006. In this study, we investigated the effects of this supernatant on A. tamarense at physiological and biochemical levels to elucidate the mechanism involved in the inhibition of algal growth by the supernatant of the strain BS02. Reactive oxygen species (ROS) levels increased following exposure to the BS02 supernatant, indicating that the algal cells had suffered from oxidative damage. The levels of cellular pigments, including chlorophyll a and carotenoids, were significantly decreased, which indicated that the accumulation of ROS destroyed pigment synthesis. The decline of the maximum photochemical quantum yield (Fv/Fm) and relative electron transport rate (rETR) suggested that the photosynthesis systems of algal cells were attacked by the BS02 supernatant. To eliminate the ROS, the activities of antioxidant enzymes, including superoxide dismutase (SOD) and catalase (CAT), increased significantly within a short period of time. Real-time PCR revealed changes in the transcript abundances of two target photosynthesis-related genes (psbA and psbD) and two target respiration-related genes (cob and cox). The transcription of the respiration-related genes was significantly inhibited by the treatments, which indicated that the respiratory system was disturbed. Our results demonstrate that the BS02 supernatant can affect the photosynthesis process and might block the PS II electron transport chain, leading to the production of excessive ROS. The increased ROS can further destroy membrane integrity and pigments, ultimately inducing algal cell death.

A kind of new molecularly imprinted polymer (MIP) was synthesized by bulk polymerization using guanosine as dummy template molecule, α-methacrylic acid as functional monomer and ethylene glycol dimethyl acrylic ester as crosslinker. Fourier transform infrared spectroscopy (FT-IR) and scanning electron microscopy (SEM) showed that the MIP had homogenous and uniform-sized cavities. It was confirmed that the MIP had higher binding affinity and selectivity towards gonyautoxins 1,4 (GTX 1,4) than the non-imprinted polymer (NIP) according to the static equilibrium adsorption. An off-line molecularly imprinted solid-phase extraction (MISPE) method followed by high-performance liquid chromatography with fluorescence detection (HPLC-FLD) was established for the analysis of GTX 1,4. 0.1 mol/L acetic acid and 95:5 (v:v) methanol/water were optimized as the washing and elution solutions, respectively. The recoveries of spiked cultured seawater samples were satisfactory, as high as 88 %. Using this method, the concentrations of GTX 1,4 from cultured seawater samples of Alexandrium minutum and Alexandrium tamarense were detected to be 1.10 μg/L and 0.99 μg/L, respectively. [ABSTRACT FROM AUTHOR]

The accumulation of paralytic shellfish toxins (PSTs) in bivalves is species specific. We compared the physiological responses and the toxin profiles in tissues of the burrowing surf clam, Paphies donacina, and the greenlipped mussel, Perna canaliculus, exposed to the toxic dinoflagellate Alexandrium tamarense. Bivalves were supplied with the toxic algae for 10 days, then allowed a detoxification period of 8 days. Clearance rates of mussels and clams were similar when fed either with toxic A. tamarense or non-toxic A. margalefi. Byssus production in the mussel was inhibited and exhalent siphon activity in clams was erratic following exposure to A. tamarense. There were considerable differences in the toxic profile between the dinoflagellate A. tamarense, and tissues of the mussel and the surf clam, indicating that bioconversion of the PSTs had taken place. Toxin profiles of the tissues were both species and tissue specific. Following an 8-day detoxification period, total PSTs in mussels had fallen to safe concentrations below 50 μg per 100 g, whereas concentrations in clams remained high, with an average value greater than 600 μg STX di-HCL equivalents per 100 g. The results confirmed that mussels and clams are important monitoring organisms for toxic algal blooms and can be used to minimise the health risk of PSTs to humans. [ABSTRACT FROM AUTHOR]

A series of gramine modified montmorillonites (gramine-MMTs) were prepared and characterized by thermogravimetric analysis, X-ray diffraction and zeta potential measurements. These modified clays with various amounts of gramine all exhibited higher removal efficiencies of both Chattonella marina and Alexandrium tamarense than did natural clay. With the augmentation of gramine content in the modified clays, the removal efficiency increased. The 24 h LC values of gramine-MMT with 12.68 % of gramine on C. marina and A. tamarense were 9.16 and 10.21 mg L, respectively. These data suggest gramine-MMT might be a promising material to remediate harmful algal blooms. [ABSTRACT FROM AUTHOR]

Península Valdés ( PV) in Argentina is an important calving ground for southern right whales ( SRWs, Eubalaena australis). Since 2005, right whale mortality has increased at PV, with most of the deaths (~90%) being calves <3 mo old. We investigated the potential involvement of harmful algal blooms ( HABs) in these deaths by examining data that include: timing of the SRW deaths, biotoxins in samples from dead SRWs, abundances of the diatom, Pseudo-nitzschia spp., and the dinoflagellate, Alexandrium tamarense, shellfish harvesting closure dates, seasonal availability of whale prey at PV and satellite chlorophyll data. Evidence of the whales' exposure to HAB toxins includes trace levels of paralytic shellfish toxins ( PSTs) and domoic acid ( DA) in tissues of some dead whales, and fragments of Pseudo-nitzschia spp. frustules in whale feces. Additionally, whales are present at PV during both closures of the shellfish industry (due to high levels of PSTs) and periods with high levels of Pseudo-nitzschia spp. and A. tamarense. There is a positive statistical relationship between monthly Pseudo-nitzschia densities (but not A. tamarense) and calf deaths in both gulfs of PV. [ABSTRACT FROM AUTHOR]

A partir de un monitoreo de microalgas toxígenas y toxinas en moluscos iniciado en 2008, llevado a cabo entre San Clemente del Tuyú y Mar Azul (Zona Norte) y en la Bahía Anegada (Zona Sur) de la Provincia de Buenos Aires fueron registrados cuatro brotes de toxicidad por toxina paralizante de moluscos (PSP) y ocho brotes por toxinas lipofílicas de moluscos (DSP/no determinadas). Los resultados obtenidos fueron empleados como herramienta técnica a partir dela cual la autoridad de aplicación (Dirección Provincial de Pesca del Ministerio de Asuntos Agrarios de la Provincia de Buenos Aires) fundó la decisión de imponer once vedas y una alerta sanitaria al consumo, extracción y comercialización de moluscos. Tres de los brotes por PSP se produjeron en la Zona Norte y la especie asociada fue Gymnodinium catenatum. El cuarto episodio se produjo en Puerto Quequén, durante un muestreo eventual, asociado a Alexandrium tamarense. En esta misma zona se produjeron tres de los eventos de toxicidad por DSP, y las especies asociadas fueron Dinophysis acuminata y D. caudata. Los restantes cinco episodios de toxicidad por toxinas lipofílicas ocurrieron en la Zona Sur, en una o más de las tres zonas clasificadas sanitariamente como A por SENASA, y ninguno pudo ser asociado a alguna de las especies toxígenas conocidas.

To find new chemical modifiers and new modified clays with excellent algae removal efficiency, the removal effects of betaine and five sulfobetaines including octyl sulfobetaine, caprylyl sulfobetaine, lauryl sulfobetaine, myristyl sulfobetaine, and palmityl sulfobetaine (SB3-16) against the harmful alga Alexandrium tamarense were observed. Betaine and the five sulfobetaines exhibited certain removal effects against A. tamarense with the highest removal efficiency by SB3-16. Hence, a series of modified clay, montmorillonite (MMT) intercalated by SB3-16 were prepared, and their removal efficiencies were evaluated. The organo-clay with 11 wt% of SB3-16 exhibited the highest removal efficiency against A. tamarense cells. Combining the environmentally benign nature of SB3-16 and the high efficiency of SB3-16 modified clay against A. tamarense, SB3-16 modified clay could be a favorable choice to control harmful algal blooms (HABs) of this species. [ABSTRACT FROM AUTHOR]

Paralytic shellfish toxins (PSTs) are originated from cyanobacteria and dinoflagellates, including Alexandrium tamarense, the common dinoflagellate species. In this study, a toxic dinoflagellate strain of A. tamarense CI01 was selected for studying the PSTs' concentration and the related protein variation during the whole cell cycle under different nutrient conditions. High-performance liquid chromatography, 2-D DIGE and Western blotting were used collectively for protein profiling and identification. Results showed that the toxin content was suppressed under nitrogen limiting condition, but enhanced in phosphorous limiting medium. Based on the results of proteomics analysis, 7 proteins were discovered to be related to the PSTs biosynthesis of A. tamarense CI01, including S-adenosylhomocysteine hydrolase, ornithine cyclodeaminase, argininosuccinate synthase, methyluridine methyltransferase cystine ABC transporter, phosphoserine phosphatase, argininosuccinate synthase and acyl-CoA dehydrogenase, which corresponds to the metabolism of the methionine, cysteine, ornithine, arginine and proline. Moreover, some photosynthesis relating proteins also increased their expression during PST synthesis period in A. tamarense CI01, such as phosphoenolpyruvate carboxylase, chloroplast phosphoglycerate kinase, peridinin-chlorophyll α-binding protein, Mg transporter protein and chloroplast phosphoglycerate kinase. The above findings are in support of our hypothesis that these proteins are involved in toxin biosynthesis of A. tamarense CI01, but cause-and-effect mechanisms need to be investigated in further studies. [ABSTRACT FROM AUTHOR]

El 26 de marzo de 2001 se registró por primera vez en las aguas del Pacífico colombiano, en el área de la Ensenada de Tumaco, la aparición de una marea roja producida por la especie Alexandrium tamarense con valores que superaron las 7.5 x 10(6) céls l-1 , la marea se mantuvo hacia la región oceánica, derivando en sentido sur-norte por efecto de las corrientes, hasta ser vista por ultima vez cerca a la Isla Gorgona, casi tres semanas después. Un año mas tarde, en marzo de 2002, una segunda proliferación, conformada por la misma especie, se reportó cerca de Cabo Corrientes con una concentración de 1.6 x 10(6) céls l -1 . Ambos casos estuvieron relacionados con la presencia de bajas temperaturas en la superficie del mar y no estuvieron asociadas con fenómenos de intoxicación y mortalidadFrom April 26th to May 15th 2001, a large algae bloom was observed off Tumaco Bay on the Pacific coast of Colombia. This was the first harmful algae bloom (HAB) reported in the region, and reached Gorgona Island, about 120 km north. Ayear later, starting March 2002, an offshore HAB developed from Cabo Corrientes North to Solano Bay. The typical abundance during the blooms reached 7.5 x 10(6) cells l-1 for the 2001 event and 1.6 x 10(6) cells l-1 for the 2002 event. During both events, low temperature and high salinity were recorded. Typical measurements in the area are 27-27.5°C and 30-31.5 psu. Values observed during the two events were 24-24.6°C and 33-34 psu; 3°C below normal and more than 2.5 psu above average values. These conditions are indicative of local upwelling processes at the time of the events. On both occasions, cells corresponding to the Alexandrium catenella/fundeyense/tamarense complex represented 99-100% of the biomass. It was difficult to differentiate the cells from A. catenella, but the presence of short chains of only 4 cells (single cells represented most of the biomass) was suggestive of A. tamarense. Shape, dimensions, and detailed structure of the apical pore complex, first apical plate, posterior sulcal plate, and position of the ventral pore on plate 1’ of cells were consistent with the description of A. tamarense, which has not been reported in the tropical East Pacific. The Control Center of Pacific Contamination of the Maritime General Direction of the Colombian Navy has been monitoring the area since 1994 without finding this species or HABs. This leads us to consider the two events as caused by recently introduced species, where local upwelling processes favor permanent and cyclic HABs. However, during these two events, there were no reports of effects on marine biota or of human poisoning, probably because the blooms occurred some distance offshore and far from exploited shellfish beds

A severe outbreak of Paralytic Shellfish Poisoning (PSP) occurred in Manzanillo and Guayacán, northwestern coast of Margarita Island, Venezuela, between August and October 1991. A bloom of dinoflagellates including Prorocentrum gracile, Gymnodinium catenatum and Alexandrium tamarense seemed to be responsible for this outbreak. Levels of PSP toxins in mussels (Perna perna) exceeded the international safety limit of saxitoxin, 80 µg STX/100 g meat. PSP toxin values varied between 2 548 and 115 µg STX/100 g meat in Manzanillo, and between 1 422 and 86 µg STX/100 g meat in Guayacán. At both locations, the highest levels were detected in August, when 24 patients exhibited typical symptoms of PSP toxicity after consuming cooked mussels (16 required hospitalization). A high pressure liquid chromatographic (HPLC) procedure was recently used on the 1991 samples. The major toxin detected in samples of both locations was decarbamoyl saxitoxin (dcSTX), but low concentrations of saxitoxin were also found in Manzanillo samples. Gonyautoxins GTX1, GTX2 and GTX3 were detected only at Guayacán, while in both locations, decarbamoylgonyatouxin (dcGTX2,3) toxins were detected. These findings represent the first time that causative toxins of PSP in Venezuela have been chemically identified, and confirm the presence of dcSTX and dcGTX in mussels from the Caribbean Sea. The presence of dcSTX and dcGTX in shellfish is indicative that Gymnodinium catenatum was a causative organism for outbreak of PSPUn severo brote de intoxicación paralizante por moluscos (PSP en inglés) ocurrió en Manzanillo y Guayacán en la costa noroeste de la Isla de Margarita, Venezuela entre agosto y octubre de 1991. Una proliferación de Prorocentrum gracile, Gymnodinium catenatum y Alexandrium tamarense causó el brote. Los niveles de PSP en mejillón (Perna perna) superaron los niveles máximos permisibles de saxitoxina, 80 µg STX/100g carne. Los niveles de toxinas variaron entre 2 548 y 115 µg STX/100 g carne en Manzanillo y entre 1 422 y 86 µg STX/100g carne en Guayacán. En ambas localidades, los máximos niveles se detectaron en agosto, cuando 24 personas presentaron síntomas típicos de PSP después de consumir mejillones cocidos (16 fueron hospitalizados). Se aplicó recientemente cromatografía líquida de alta presión (HPLC) a muestras del año 1991 y la toxina más detectada fue decarbamoyl saxitoxina (dcSTX), pero también se encontró saxitoxinas en muestras de Manzanillo. Las gonyautoxinas GTX1, GTX2 y GTX3 solo se encontraron en Guayacán; en ambas localidades se detectó decarbamoylgonyatouxin (dcGTX2,3). Estos hallazgos representan la primera vez que las toxinas causantes de un brote de PSP en Venezuela han sido químicamente identificadas, confirmando la presencia de dcSTX y dcGTX en mejillones del mar Caribe. La presencia de dcSTX y dcGTX en moluscos, indica que G. catenatum fue el organismo responsable de la intoxicación

Harmful algal blooms (HABs) cause a variety of deleterious effects on aquatic ecosystems, especially the toxic dinoflagellate Alexandrium tamarense, which poses a serious threat to marine economic and human health based on releasing paralytic shellfish poison into the environment. The algicidal bacterium Deinococcus sp. Y35 which can induce growth inhibition on A. tamarense was used to investigate the functional mechanism. The growth status, reactive oxygen species (ROS) content, photosynthetic system and the nuclear system of algal cells were determined under algicidal activity. A culture of strain Y35 not only induced overproduction of ROS in algal cells within only 0.5 h of treatment, also decrease the total protein content as well as the response of the antioxidant enzyme. Meanwhile, lipid peroxidation was induced and cell membrane integrity was lost. Photosynthetic pigments including chlorophyll a and carotenoid decreased along with the photosynthetic efficiency being significantly inhibited. At the same time, photosynthesis-related gene expression showed down-regulation. More than, the destruction of cell nuclear structure and inhibition of proliferating cell nuclear antigen (PCNA) related gene expression were confirmed. The potential functional mechanism of the algicidal bacterium on A. tamarense was investigated and provided a novel viewpoint which could be used in HABs control. [ABSTRACT FROM AUTHOR]

The dinoflagellate Alexandrium tamarense produces paralytic shellfish toxins that cause foodborne illnesses via bivalve consumption. The toxin profiles produced by A. tamarense and the toxins in the blue mussel ( Mytilus edulis) were compared by high-performance liquid chromatography. Whereas the 11β-epimer toxins C2 and gonyautoxin GTX4 were predominant toxins produced by A. tamarense, but the 11α-epimer toxin GTX1 was predominant, followed by C1, C2, and GTX in blue mussel. In the early period (March 24 to April 7) of toxin accumulation in blue mussels, the levels of the C2 and GTX4 were higher than those of C1 and GTX1. However, with an increase in intoxication time, the relative proportions of 11β-epimer toxins and 11α-epimer toxins were reversed, so that the ratios of C1 and GTX1 were 2.1-2.4 times higher than those of C2 and GTX4, respectively, until May 31. This shifting pattern of predominance indicates that toxicity increases through a conversion process during accumulation. [ABSTRACT FROM AUTHOR]

To explore the genetic diversity and paralytic shellfish poisoning (PSP) toxin profile of the Alexandrium tamarense species complex along the coast of China, 67 strains of A. tamarense from the China Sea were collected and genetic diversity were analyzed based on the rDNA sequences. In addition, PSP toxin compositions and contents were detected by HPLC. According to the 5.8S rDNA and ITS, and LSU rDNA D1–D2 sequence, A. tamarense in the China Sea comprises at least Group IV and Group I ribotypes. In these Chinese strains, the toxins with the highest concentration in the profile were C1/2, gonyautoxins 1/4 (GTX1/4) and neosaxitoxin (NEO). However, the toxin profiles were atypical and C1/2 toxins were not detected in some strains. No strict correlation was observed between the PSP toxins profile and the geographical distribution. [ABSTRACT FROM AUTHOR]

The Alexandrium tamarense species complex is one of the most studied marine dinoflagellate groups due to its ecological, toxicological and economic importance. Several members of this complex produce saxitoxin and its congeners - potent neurotoxins that cause paralytic shellfish poisoning. Isolates from this complex are assigned to A. tamarense , A. fundyense , or A. catenella based on two main morphological characters: the ability to form chains and the presence/absence of a ventral pore between Plates 1′ and 4′. However, studies have shown that these characters are not consistent and/or distinctive. Further, phylogenies based on multiple regions in the rDNA operon indicate that the sequences from morphologically indistinguishable isolates partition into five clades. These clades were initially named based on their presumed geographic distribution, but recently were renamed as Groups I-V following the discovery of sympatry among some groups. In this study we present data on morphology, ITS/5.8S genetic distances, ITS2 compensatory base changes, mating incompatibilities, toxicity, the sxtA toxin synthesis gene, and rDNA phylogenies. All results were consistent with each group representing a distinct cryptic species. Accordingly, the groups were assigned species names as follows: Group I, A. fundyense; Group II, A. mediterraneum; Group III, A. tamarense ; Group IV, A. pacificum; Group V, A. australiense . [ABSTRACT FROM AUTHOR]

This study investigated the inductive effect of Alexandrium tamarense, a toxic dinoflagellate producing paralytic shellfish poison, on oxidative stress and apoptosis in hepatopancreas of Chinese shrimp, Fenneropenaeus chinensis. The individuals of F. chinensis were exposed to 200 and 1000 cells mL of A. tamarense with their superoxide dismutase (SOD), glutathione S-transferase (GST) activities, malonyldialdehyde (MDA) concentration, and caspase gene ( FcCasp) expression in hepatopancreas determined at 12, 24, 48, 72 and 96 h. In addition, apoptosis in hepatopancreas of F. chinensis at 96 h after exposure was determined through terminal deoxynucleotidyl transferase-mediated dUTP nick end labeling (TUNEL) assay. The hepatopancreatic SOD and GST activities of F. chinensis exposed to 1000 cells mL of A. tamarense showed a bell-shaped response to exposure time. The hepatopancreatic MDA concentration of F. chinensis exposed to 1000 cells mL of A. tamarense increased gradually from 48 to 96 h, and such a trend corresponded to the decrease of GST activity. The hepatopancreatic FcCasp transcript abundance of F. chinensis exposed to 1000 cells mL of A. tamarense was positively and linearly correlated to MDA concentration. Results of TUNEL assay showed that exposure to 1000 cells mL of A. tamarense induced apoptosis in the hepatopancreas of F. chinensis. Our study revealed that A. tamarense exposure influenced the antioxidative status of F. chinensis and caused lipid peroxidation and apoptosis in the hepatopancreas of shrimp. [ABSTRACT FROM AUTHOR]