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Harmful algal blooms (HABs) have always been a worldwide environmental issue. The methods based on the principle of allelopathy provide a novel direction for controlling HABs; however, there are a few studies on the application of allelopathic algaecides to control harmful algae in marine environments. Here we examined the algicidal capacity of 15 fruit peel extracts with biological activity on Alexandrium catenella. The results displayed that the mango peel extracts (MPE) showed efficient inhibition on species growth. The algicidal rate reached 93.32 ± 0.56% at 96 h after adding 5 g/L MPE to the culture medium of A. catenella. Furthermore, we found that the expression of key genes involved in PSII and PSI was downregulated as well as obstructed the electron transportation in the light reaction process and the synthesis of organic matter. The blocked photosynthetic chain induced the accumulation of substantial reactive oxygen species, resulting in severe peroxidation of the membrane lipids. Simultaneously, the expression pattern of key genes involved in the fatty acid, amino acid, and peroxisome breakdown pathways was upregulated, which suggested that the synthesis and decomposition of intracellular organic matter may be in an imbalanced state. The results above indicated that oxidative damage and energy metabolism disequilibrium are two key pathways by which MPE induced algal cell death. Furthermore, several kinds of active substances and their proportion in MPE had been identified by liquid chromatography quadrupole time-of-flight mass spectrometry. It is speculated that esters may be the important component playing an algicidal effect. However, the specific substance that plays a key role in inhibiting the growth of A. catenella and the algicidal mechanism remain to be further studied. This study might provide a new direction in the management of HABs in the future. [ABSTRACT FROM AUTHOR]

The blooms of Alexandrium catenella, the main producer of paralytic shellfish toxins worldwide, have become the main threat to coastal activities in Southern Chile, such as artisanal fisheries, aquaculture and public health. Here, we explore retrospective data from an intense Paralytic Shellfish Poisoning outbreak in Southern Chile in Summer–Autumn 2016, identifying environmental drivers, spatiotemporal dynamics, and detoxification rates of the main filter-feeder shellfish resources during an intense A. catenella bloom, which led to the greatest socio-economic impacts in that area. Exponential detoxification models evidenced large differences in detoxification dynamics between the three filter-feeder species surf clam (Ensis macha), giant barnacle (Austromegabalanus psittacus), and red sea squirt (Pyura chilensis). Surf clam showed an initial toxicity (9054 µg STX-eq·100 g−1) around 10-fold higher than the other two species. It exhibited a relatively fast detoxification rate and approached the human safety limit of 80 µg STX-eq·100 g−1 towards the end of the 150 days. Ecological implications and future trends are also discussed. Based on the cell density evolution, data previously gathered on the area, and the biology of this species, we propose that the bloom originated in the coastal area, spreading offshore thanks to the resting cysts formed and transported in the water column. [ABSTRACT FROM AUTHOR]

Consuming poisoned shellfish can lead to severe health problems and even death. Alexandrium catenella (Group I) and A. pacificum (Group IV) cause paralytic shellfish poisoning (PSP) in Korea, and PSP is detected more in a wider area. However, the association between toxic dinoflagellates and shellfish poisoning is unclear. Therefore, it is necessary to understand the toxicity, detoxification, and compositional differences in PSP in Mytilus edulis exposed to PSP caused by A. catenella and A. pacificum. High-performance liquid chromatography with post-column oxidation was used to analyze PSP toxicity in poisoned M. edulis. PSP in M. edulis increased as the A. catenella and A. pacificum cell density increased. However, the cell density of A. catenella peaked faster than that of A. pacificum, and a high level of toxicity was detected. In the detoxification experiment, PSP in M. edulis decreased rapidly within 24 h in filtered seawater. However, PSP was continuously detected without decreasing below the detection limit until the last day of the experiment. In addition, the carbamate composition (GTX1+4) was detected as the main toxic composition in poisoned M. edulis, unlike in vegetative cells. GTX1+4 can poison shellfish quickly when toxic dinoflagellates appear in the marine environment. However, poisoned shellfish take a long time to be completely detoxified. Moreover, if shellfish continuously feed on poisonous dinoflagellates, their toxicity can increase rapidly due to biotransformation. Our results can help identify the mechanisms of shellfish toxicity and detoxification after PSP caused by toxic dinoflagellates. [ABSTRACT FROM AUTHOR]

Harmful algal blooms (HABs) have always been a worldwide environmental issue. The methods based on the principle of allelopathy provide a novel direction for controlling HABs; however, there are a few studies on the application of allelopathic algaecides to control harmful algae in marine environments. Here we examined the algicidal capacity of 15 fruit peel extracts with biological activity on Alexandrium catenella. The results displayed that the mango peel extracts (MPE) showed efficient inhibition on species growth. The algicidal rate reached 93.32 ± 0.56% at 96 h after adding 5 g/L MPE to the culture medium of A. catenella. Furthermore, we found that the expression of key genes involved in PSII and PSI was downregulated as well as obstructed the electron transportation in the light reaction process and the synthesis of organic matter. The blocked photosynthetic chain induced the accumulation of substantial reactive oxygen species, resulting in severe peroxidation of the membrane lipids. Simultaneously, the expression pattern of key genes involved in the fatty acid, amino acid, and peroxisome breakdown pathways was upregulated, which suggested that the synthesis and decomposition of intracellular organic matter may be in an imbalanced state. The results above indicated that oxidative damage and energy metabolism disequilibrium are two key pathways by which MPE induced algal cell death. Furthermore, several kinds of active substances and their proportion in MPE had been identified by liquid chromatography quadrupole time-of-flight mass spectrometry. It is speculated that esters may be the important component playing an algicidal effect. However, the specific substance that plays a key role in inhibiting the growth of A. catenella and the algicidal mechanism remain to be further studied. This study might provide a new direction in the management of HABs in the future.

The marine dinoflagellate Alexandrium catenella (Group I) produces saxitoxins (STXs) in marine environments, causing paralytic shellfish poisoning (PSP). The toxins are synthesized by STXs biosynthesis genes (sxt); of them, sxtA and sxtG are considered the most essential. The promoter region and 3'-untranslated region (3'-UTR) are known to regulate gene transcription; however, they have not been sufficiently elucidated in the sxt genes. Herein, we determined and characterized the genomic DNA sequence of the proximal regions of sxtA and sxtG in A. catenella. The promoter regions of sxtA and sxtG in A. catenella were found to contained distinct cis-regulatory elements (CREs), CpG islands, and TTTT-motif (instead of TATA-box), showing a typical eukaryotic promoter. Especially, a number of light-responsive CREs (G-box, SORLIP1AT/2AT, and Sp1) were found, and the gene expression level showed a significant relationship with photosynthesis. MiSeq sequencing of the long PCR amplicons revealed that sxtA was encoded as a single exonic structure that is discrete (> 5 kbp) from other core sxt. In addition, we reported for the first time two types of 3'-UTR in sxtG mRNA, and their lengths and sequences varied among strains and species. Such results suggest that sxt may be regulated by post-transcription. Our results provide insights into the toxin dynamics of toxic dinoflagellates from a molecular perspective. [ABSTRACT FROM AUTHOR]

The blooms of Alexandrium catenella, the main producer of paralytic shellfish toxins worldwide, have become the main threat to coastal activities in Southern Chile, such as artisanal fisheries, aquaculture and public health. Here, we explore retrospective data from an intense Paralytic Shellfish Poisoning outbreak in Southern Chile in Summer–Autumn 2016, identifying environmental drivers, spatiotemporal dynamics, and detoxification rates of the main filter-feeder shellfish resources during an intense A. catenella bloom, which led to the greatest socio-economic impacts in that area. Exponential detoxification models evidenced large differences in detoxification dynamics between the three filter-feeder species surf clam (Ensis macha), giant barnacle (Austromegabalanus psittacus), and red sea squirt (Pyura chilensis). Surf clam showed an initial toxicity (9054 µg STX-eq·100 g−1) around 10-fold higher than the other two species. It exhibited a relatively fast detoxification rate and approached the human safety limit of 80 µg STX-eq·100 g−1 towards the end of the 150 days. Ecological implications and future trends are also discussed. Based on the cell density evolution, data previously gathered on the area, and the biology of this species, we propose that the bloom originated in the coastal area, spreading offshore thanks to the resting cysts formed and transported in the water column.

Herbicides enter the ocean through surface runoff and have become a serious pollution threat in the coastal areas. As photosynthetic inhibitors, triazine herbicides are more harmful to marine photosynthetic organisms, especially phytoplankton, than to marine animals. Herbicide pollution may also cause other environmental issues such as algal blooms by altering the structure of phytoplankton community and toxin production in bloom-forming species. To explore whether triazine herbicides will cause similar effects, we analyzed the effects of a common triazine herbicide pollutant, terbutryn, on the physiological responses and gene expression of Alexandrium catenella, a harmful algal bloom forming dinoflagellate. Our results show that A. catenella maintained a normal growth rate when exposed to terbutryn at the concentrations of 0.1 and 1 μg L−1. Treatment with terbutryn at 10 μg L−1 significantly reduced the growth rate by inhibiting photosynthesis in A. catenella, which may also affect the repair of photosystem II by repressing the expression of the D1 protein gene. Exposure to terbutryn at 1 μg L−1 also impacted the photosystem II, but no changes were observed on the maximum relative electron transport rate of photosynthesis. Additionally, expression of genes involved in saxitoxin was also analyzed in this study, which were not affected by the terbutryn treatment. Our study provides critical information about the potential threat of a herbicide in affecting a harmful algal bloom species. [ABSTRACT FROM AUTHOR]

Paralytic shellfish toxins (PSTs) are phycotoxins widely distributed worldwide and pose serious marine ecosystems and human health threats. In China's Yellow Sea and the Bohai Sea, Alexandrium spp. has been certified as the major causative dinoflagellate of PSTs, especially in Qinhuangdao, Hebei Province where several poisoning events have been reported, with tens of consumers suffering and some even dying. In terms of these events, mussels contaminated with PSTs were the major cause of consumer poisoning. Therefore, it is vital to reveal the risk of PSTs in these shellfish, which requires scientific opinions on the formation of terminal components of PSTs in mussels. Generally, mussels are not sensitive to PSTs, resulting in the high accumulation of PSTs in their tissues. The PSTs distribute, bio-transfer, and metabolism, and the terminal metabolites pose a risk to consumers. From 2016 to the present, several serious events have occurred in Qinhuangdao caused by PSTs contamination in mussels, which resulted in poisoning by tens of consumers and a huge loss of regional economy. This study exposed purple mussel Mytilus galloprovincialis, the key cultured species of bivalves in Qinhuangdao, to Alexandrium catenella (GY-H25), the predominant producer of PSTs in this area. The accumulation and biotransformation process of PSTs in visceral mass and edible tissue and the accumulation metabolism kinetics were analyzed under exposure to different cell densities by liquid chromatography-tandem mass spectrometry. Our results showed that the growth and toxin production of GY-H25 was stable. The main components of PSTs were N-sulfocarbamoylgonyautoxin (C1 and C2), with the highest algal cell density of 3.5×107 cells/L and the highest production capacity of 2.96 pg STXeq/cell. Toxin-producing algae with the highest production of PSTs per unit volume and algal cell density (22 days) were selected for the exposure experiment. After exposure, the mussel accumulated a high content of PSTs in both visceral mass and edible tissue, with a similar trend in both exposure densities. The whole exposure experiment could be divided into four periods: 0~2 days as the initial exposure period (period Ⅰ); 2~7 days as the period of rapid accumulation (period Ⅱ); 7~12 days as the period of rapid metabolism (period Ⅲ); 12~30 days as the stable period (period Ⅳ). However, the level of PSTs in the visceral mass of mussels in both exposure groups exceeded the maximum residue limit (MRL) of EU (800 μg STX EQ/kg) at the end of the experiment, while that of PSTs in edible flesh was below the MRL. Comparatively, the highest concentration of PSTs in the visceral mass reached 6815.36 μg/kg in the high exposure group, which was 2.61 times that of the low exposure group, with an average accumulation rate of 17.89%, which was significantly higher than that of the low exposure group (13.06%). The results showed that the accumulation of PSTs in the visceral mass and edible tissue of Mytilus galloprovincialis was harmful to its tissues and organs. The toxin excretion rates of the high-and low-concentration groups of PSTs were 74.39 % and 59.15 %, respectively, after 23 days. The average daily elimination rate was 14.4 %, the metabolism rate of Mytilus edulis slowed down in the stable period (period Ⅳ), and some toxins remained in the visceral mass after 23 days of metabolism, which easily formed long-term toxin retention and threatened human life safety. In addition, PSTs in Mytilus galloprovincialis showed a strong biotransformation ability, mainly occurring among C1, C2, and GTX5. According to this research, the transformation pathway from C2 to GTX5 was the main pathway for GTX5 formation during rapid metabolism and stable periods. At the same time, the transformation of C1 to GTX5 was higher than that of C2 to C1, leading to a reduction in the overall proportion of C1. According to the comprehensive evaluation of the metabolic products and toxic equivalence factor (TEF) of GST components in mussels, the metabolic transformation of PSTs in mussels will further promote the formation and proportion of highly toxic GTX5, which will increase the terminal toxicity of PSTs in mussels and may also pose a higher risk to consumers. In this study, compared with the natural conditions, the concentration of toxic algae was far below the harmful algal bloom exposure. The total accumulation of toxins was far lower than the total accumulation of toxins in mussels exposed to harmful algal blooms. Even so, the total amount of residual toxins in the mussel viscera of the high concentration group was still enough to threaten human life safety at the end of the experiment. It was found that the toxin content in the visceral tissue tended to be stable over 12 to 30 days without an obvious downward trend. Even the toxin content increases due to the toxin transformation phenomenon, which means the difficulty in predicting the time required for PSTs in the visceral mass reduced to below the MRLs. Therefore, this study contributes to the scientific assessment of PSTs risk in mussels and provides a basis for establishing regional PSTs monitoring programs.

In the framework of a monitoring program of harmful microalgae from the marine coastal waters of the Buenos Aires Province, seven strains were isolated and characterized by morphological and molecular analysis (LSU rDNA partial sequencing, D1–D3 regions). Established strains belonged to Alexandrium catenella, Protoceratium reticulatum and Pseudo-nitzschia multiseries. The toxinological profile of the target strains were determined by UHPLC-FLD equipment for paralytic shellfish toxins (PSTs) and LC-MS/MS for lipophilic (LSTs) and amnesic toxins (ASTs). The toxin profile varied in the four strains of A. catenella, the predominant compounds were gonyautoxins (GTXs) GTX2,3 and GTX1,4 for strains LPCc001 and LPCc004, and N-sulfocarbamoyl toxins (Cs) C1,2 and GTX1,4 for strains LPCc002 and LPCc008. The obtained cellular toxicity values were moderate-to-high (12.38–46.40 pg saxitoxin equiv. cell−1). The toxin profile of P. reticulatum was dominated by yessotoxins (YTXs) (up to 94.40 pg cell−1) accompanied by homo-yessotoxin (Homo-YTX) traces. In P. multiseries, the toxin profiles were dominated by domoic acid (DA) (1.62 pg cell−1 and 1.09 pg cell−1) and secondarily by Isomer A (Iso-A), Epi-domoic acid (Epi-DA), Isomer-E (Iso-E) and Isomer-D (Iso-D). This study provides detailed information about representative HAB species in the area, useful for resource management, risk evaluation and related research on toxic dinoflagellates and diatoms.

Saxitoxin (STX) is a neurotoxin produced by certain toxic cyanobacteria and dinoflagellates. It is biochemically synthesized by the stepwise involvement of eight core enzymes encoded in sxt genes. Of them, sxtB, a cytidine deaminase (CDA)-like enzyme, may participate in the third step (after sxtA and sxtG) in the toxin synthesis pathway; however, it is insufficiently elucidated in toxic dinoflagellates. In the present study we determined a novel sxtB gene from the STX-producing dinoflagellate Alexandrium catenella, characterized structural motifs and phylogenetic origin, and evaluated transcriptional responses of the gene under different temperatures. cDNA of the AcsxtB was 1,146 bp in length from dinoflagellate spliced leader (Dino-SL) to poly (A) tail. It comprised a 972 bp open reading frame, encoding a 323 aa protein with a molecular weight of 34.20 kDa and isoelectric point of 6.36. In addition, we identified a putative mitochondrial transfer peptide and no introns in genomic coding regions. The AcsxtB was phylogenetically close to toxic cyanobacteria, but distant to non-toxic dinoflagellates. The transcription levels of AcsxtB were significantly up-regulated when cultured at 16 °C (3.36-fold) and exposed to cold stress (20 → 12 °C; 1.88-fold, 20 → 16 °C; 2.07-fold), which showed a high correlation with increased STX contents. These suggest that the sxtB should participate in STX synthesis, but its involvement may be low or regulated in very early stages before 72 h. This is the first report on sxtB characterization and transcriptional regulation with relevance to STX production in the toxic dinoflagellates. [ABSTRACT FROM AUTHOR]

In the framework of a monitoring program of harmful microalgae from the marine coastal waters of the Buenos Aires Province, seven strains were isolated and characterized by morphological and molecular analysis (LSU rDNA partial sequencing, D1–D3 regions). Established strains belonged to Alexandrium catenella, Protoceratium reticulatum and Pseudo-nitzschia multiseries. The toxinological profile of the target strains were determined by UHPLC-FLD equipment for paralytic shellfish toxins (PSTs) and LC-MS/MS for lipophilic (LSTs) and amnesic toxins (ASTs). The toxin profile varied in the four strains of A. catenella, the predominant compounds were gonyautoxins (GTXs) GTX2,3 and GTX1,4 for strains LPCc001 and LPCc004, and N-sulfocarbamoyl toxins (Cs) C1,2 and GTX1,4 for strains LPCc002 and LPCc008. The obtained cellular toxicity values were moderate-to-high (12.38–46.40 pg saxitoxin equiv. cell−1). The toxin profile of P. reticulatum was dominated by yessotoxins (YTXs) (up to 94.40 pg cell−1) accompanied by homo-yessotoxin (Homo-YTX) traces. In P. multiseries, the toxin profiles were dominated by domoic acid (DA) (1.62 pg cell−1 and 1.09 pg cell−1) and secondarily by Isomer A (Iso-A), Epi-domoic acid (Epi-DA), Isomer-E (Iso-E) and Isomer-D (Iso-D). This study provides detailed information about representative HAB species in the area, useful for resource management, risk evaluation and related research on toxic dinoflagellates and diatoms. [ABSTRACT FROM AUTHOR]

Outbreaks of paralytic shellfish poisoning caused by the toxic dinoflagellate Alexandrium catenella (Dinophyceae) are currently a serious global problem both from economic and food hygiene perspectives. In Chile, A. catenella was first recorded in 1972 and is currently the main harmful algae species in the country, in terms of seafood security. As no relevant microsatellite markers for Chilean A. catenella populations were available, we isolated 23 new polymorphic microsatellite loci of A. catenella from Chile, of which 15 are applicable to both Chilean and Japanese populations. We found a strong genetic break between Chilean and Japanese A. catenella populations (FST=0.298, P<0.001), consistent with the morphological differences between them. In contrast to the Japanese population, for which genetic diversity ranged from 0.268 to 0.937, the Chilean population had a lower genetic diversity, ranging between 0.065 and 0.512. This study successfully developed A. catenella microsatellite markers that can be used to investigate its genetic spatial and temporal bloom diversity and assess further genetic connectivity between the population from southern and northern Chilean areas. [ABSTRACT FROM AUTHOR]

The dinoflagellate genus Alexandrium contains a number of species that produce paralytic shellfish toxins and have been the focus of attention as toxic plankton for harmless algal studies. Among Alexandrium species, A. catenella and A. pacificum form ellipsoidal-shaped resting cysts, which are preserved in marine sediments, and have attracted attention as potential seeds for future proliferation after favorable environmental conditions environmental improvement. However, although these cysts are preserved in surface of marine sediments, there is no record of their occurrence from solidified sediments as fossils. In order to clarify the reason for this, we investigated the differences in the chemical composition of cyst walls between colorless cyst of Alexandrium catenella/pacificum and Lingulodinium machaerophorum, Polysphaeridium zoharyi, Spiniferites spp. by measuring the thickness of cyst walls and using Fourier Transform Infrared Spectroscopy (FTIR) and Raman Spectroscopy. The results showed that the cyst wall of A. catenella/pacificum and L. machaerophorum were all composed of cellulosic organic matter. However, A. catenella/pacificum have the higher percentage of α −/β-glucosidic linkages and that the thickness of their cyst walls is about one-third of that of L. machaerophorum. Therefore, these are reasons for the cysts of A. catenella / pacificum being more easily degraded in the sediment. • Cysts of Alexandrium catenella and/or A. pacificum are ellipsoidal in shape. • Cyst wall of Alexandrium catenella/pacificum is composed of cellulose. • Due to the higher percentage of 1,4-β-glucosidic linkages in the cyst wall, the cysts of A. catenella/pacificum are more easily degraded in the sediment. [ABSTRACT FROM AUTHOR]

Alexandrium causes serious food safety and human death due to paralytic shellfish toxins (PSTs) production. The associated bacteria can affect PSTs production of Alexandrium. However, the influencing mechanism is still unclear. Here we firstly screened functional associated bacteria for affecting PSTs production of Alexandrium catenella in Yangtze Estuary and further studied their influence on physiological process and molecular regulation of A. catenella. Thirteen bacteria strains for affecting PSTs production of A. catenella were selected. The A. catenella strains co-cultured with different functional associated bacteria all produced more PSTs than axenic strain with antibiotic treatment. Compared with axenic A. catenella , the non-axenic A. catenella produced more algal cells, soluble sugar, soluble protein and neutral lipid. By RNA-seq, it was found that non-axenic A. catenella produced more upregulated functional genes than axenic A. catenella. The biosynthesis of cofactors and spliceosome were the dominant different pathways between axenic and non-axenic A. catenella strains. The sxt A expression was closely related with Arginine and proline metabolism, Arginine biosynthesis, Fatty acid biosynthesis, TCA cycle and Glutathione metabolism, which were all downregulated in axenic A. catenella. Meantime, the non-axenic A. catenella under nitrogen deprivation produced less PSTs and functional genes than non-axenic strain under common culture condition, indicating the nitrogen significance for PSTs production. The detailed signal molecular released by associated bacteria for regulating PSTs of A. catenella needs to be further studied. [Display omitted] • Thirteen bacteria affecting PSTs production were selected. • catenella strains with bacteria produced more PSTs. • The non-axenic A. catenella produced more molecular components of physiology. • Cofactors and spliceosome were the dominant signals with bacteria influence. • The sxtA and related pathways were downloaded in axenic algal stains. [ABSTRACT FROM AUTHOR]

Dinoflagellate Alexandrium catenella is a cosmopolitan bloom-forming species with complex life cycle, the formation and germination of resting cysts are critical for its bloom dynamics. In the coastal waters of Qinhuangdao, A. catenella has been identified as the major causative agent for paralytic shellfish poisoning, but there is little knowledge concerning its resting cysts in this region. In this study, three surveys were carried out along the coast of Qinhuangdao from 2020 to 2021 to map the distribution of A. catenella resting cysts, using a quantitative PCR (qPCR) assay specific for A. catenella. The resting cysts were detected in surface sediments during all the three surveys, and their distribution patterns were similar. High abundance of resting cysts (maximum 1 300 cysts/g sediment (wet weight)) were found in a region (119.62°E–119.99°E, 39.67°N–39.98°N) northeast to the coastal waters of Qinhuangdao, where surface sediments were mainly composed of clay and silt (percentage above 50%). Prior to the formation of the A. catenella bloom in March 2021, the abundance of A. catenella vegetative cells in seawater had extremely significant positive correlation with the abundance of resting cysts in surface sediments, reflecting the important role of resting cysts in the initiation of A. catenella blooms. As far as we know, this is the first report on the distribution of A. catenella cysts along the coast of Qinhuangdao. The results will offer a sound basis for the future monitoring and mitigation of toxic A. catenella blooms and paralytic shellfish poisoning events in this region. [ABSTRACT FROM AUTHOR]

• A lexandrium catenella produced paralytic shellfish toxins and allelochemicals. • Allelochemicals caused negative effects on protists and metazoan microzooplankton. • co-occurring competitors alleviated negative effects on microzooplankton. • mathematical model predicted experimental outcome of complex food web dynamics 5. The production of allelochemicals by the toxigenic dinoflagellate Alexandrium catenella is one of the suggested mechanisms to facilitate its bloom formation and persistence by outcompeting other phototrophic protists and reducing grazing pressure. In Southern California, toxic events caused by A. catenella and paralytic shellfish toxins (PSTs) regularly impact coastal ecosystems; however, the trophic interactions and mechanisms promoting this species in a food web context are still not fully understood. In the present study, we combined a dynamical mathematical model with laboratory experiments to investigate potential toxic and allelochemical effects of an A. catenella strain isolated off the coast of Los Angeles, Southern California, on competitors and a common zooplankton consumer. Experiments were conducted using three toxigenic strains of A. catenella , comparing the new Californian isolate (Alex Cal) to two strains previously described from the North Sea, a lytic (Alex2) and non-lytic (Alex5) strain, testing for donor density-dependent effects on two phytoplankton species (Rhodomonas salina, Tetraselmis sp.) and on the rotifer Brachionus plicatilis. Bioassays revealed a steep decline in competitor and consumer populations with increasing Alex Cal concentrations, indicating an intermediate lytic activity compared to the North Sea strains (lytic Alex2 and non-lytic Alex5). The rotifer fed and grew well on the PST- toxic, but non-lytic Alex5 strain, while its survival significantly decreased with increasing concentrations of the two lytic strains Alex Cal and Alex 2, indicating that negative effects on the rotifer were mediated by allelochemicals rather than PST-toxins. Mixed culture experiments including both competitors and consumers demonstrated that the intensity of allelochemical effects not only depended on the A. catenella density but also on the target density. Negative effects on grazers were alleviated by co-occurring competitors with a lower sensitivity to allelochemicals, thus reducing harmful compounds and allowing grazing control on the dinoflagellate to come into effect again. Results from mixed culture experiments were supported by the mathematical approach used in this study which was calibrated with data from simple monoculture growth, pairwise competition and predator-prey experiments, demonstrating the applicability of this model approach to predict the outcome of more complex food web dynamics at the community level. [Display omitted] [ABSTRACT FROM AUTHOR]

Harmful algal bloom (HAB) species Alexandrium catenella and Dinophysis acuminata are associated with paralytic shellfish poisoning (PSP) and diarrhetic shellfish poisoning (DSP) in humans, respectively. While PSP and DSP have been studied extensively, less is known about the effects of these HAB species or their associated toxins on shellfish. This study investigated A. catenella and D. acuminata toxicity in a larval oyster (Crassostrea virginica) bioassay. Larval activity and mortality were examined through 96-h laboratory exposures to live HAB cells (10–1000 cells/mL), cell lysates (1000 cells/mL equivalents), and purified toxins (10,000 cells/mL equivalents). Exposure to 1000 cells/mL live or lysed D. acuminata caused larval mortality (21.9 ± 7.0%, 10.2 ± 4.0%, respectively) while exposure to any tested cell concentration of live A. catenella, but not lysate, caused swimming arrest and/or mortality in >50% of larvae. Exposure to high concentrations of saxitoxin (STX) or okadaic acid (OA), toxins traditionally associated with PSP and DSP, respectively, had no effect on larval activity or mortality. In contrast, pectenotoxin-2 (PTX2) caused rapid larval mortality (49.6 ± 5.8% by 48 h) and completely immobilized larval oysters. The results indicate that the toxic effects of A. catenella and D. acuminata on shellfish are not linked to the primary toxins associated with PSP and DSP in humans, and that PTX2 is acutely toxic to larval oysters. [ABSTRACT FROM AUTHOR]

Microalgae host varied microbial consortium harboring cross-kingdom interactions with fundamental ecological significance in aquatic ecosystems. Revealing the complex biofunctions of the cultivable bacteria of phycosphere microbiota is one vital basis for deeply understanding the mechanisms governing these dynamic associations. In this study, a new light-yellow pigmented bacterial strain LZ-28 was isolated from the highly-toxic and harmful algal bloom-forming dinoflagellate Alexandrium catenella LZT09. Collective phenotypic and genotypic profiles were obtained to confidently identify this strain as a new Mameliellaalba member. Comparative genomic analysis showed that strain LZ-28 shared highly similar functional features with other four marine algae-derived M. alba strains in spite of their distinctive isolation sources. Based on the bioactivity assaying, the mutual growth-promoting effects between bacterial strain LZ-28 and algal strain LZT09 were observed. After the culture conditions were optimized, strain LZ-28 demonstrated an extraordinary production ability for its bioflocculanting exopolysaccharides (EPS). Moreover, the portions of two monosaccharides glucose and fucose of the EPS were found to positively contribute to the bioflocculanting capacity. Therefore, the present study sheds light on the similar genomic features among the selected M. alba strains, and it also reveals the potential pharmaceutical, environmental and biotechnological implications of active EPS produced by this new Mameliella alba strain LZ-28 recovered from toxic bloom-forming marine dinoflagellate. [ABSTRACT FROM AUTHOR]

Two yellow-pigmented bacterial strains, LZ-14 T and ABI-LZ29, were isolated from the cultivable phycosphere microbiota of the highly toxic marine dinoflagellate Alexandrium catenella LZT09 and demonstrated obvious microalgae growth-promoting potentials toward the algal host. To elucidate the taxonomic status of the two bioactive bacterial strains, they were subjected to a polyphasic taxonomic characterization. Both strains were found to be Gram-negative, aerobic, rod-shaped and motile; to contain Q-10 as the predominant ubiquinone; summed feature 8, C16:0, C18:1ω7c 11-methyl and summed feature 3 as the major fatty acids; and diphosphatidylglycerol, phosphatidylcholine, phosphatidylethanolamine, phosphatidylglycerol and two unidentified phospholipids as the predominant polar lipids. Based on the phylogenetic analysis, phylogenomic inferences and phenotypic characteristics, the strains could be clearly distinguished from phylogenetically closely related species and formed a distinct monophyletic lineage in the family Rhodobacteraceae. The size of the draft genome of strain LZ-14 T is 4.615 Mb, with a DNA G + C content of 63.3 mol%. It contains ten predicted secondary metabolite biosynthetic gene clusters and core genes for bacterial exopolysaccharide biosynthesis. Therefore, strain LZ-14 T (= CCTCC AB 2017230 T = KCTC 62342 T) represents a novel species of a new genus, for which the name Alexandriicola marinus gen. nov., sp. nov., is proposed. [ABSTRACT FROM AUTHOR]

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 brought severe ecological and economic impacts to Chile, especially in the southern region, where the shellfish and salmon industries are world-famous. The mechanisms of such HABs have been intensively studied but are still unclear. Nutrient overloading is one of the often-discussed drivers for HABs. The present study used the A. catenella strain isolated from southern Chile to investigate how iron conditions could affect their growth and toxin production as related to HAB. Our results showed that an optimum concentration of iron was pivotal for proper A. catenella growth. Thus, while excess iron exerted a toxic effect, low iron media led to iron insufficiency and growth inhibition. In addition, the study shows that the degree of paralytic shellfish toxin production by A. catenella varied depending on the iron concentration in the culture media. The A. catenella strain from southern Chile produced GTX1-4 exclusively in the fmol cell−1 scale. Based on these findings, we suggest that including iron and paralytic shellfish toxin measurements in the fields can improve the current HAB monitoring and contribute to an understanding of A. catenella bloom dynamics in Chile. [ABSTRACT FROM AUTHOR]

Harmful algal blooms, in particular recurrent blooms of the dinoflagellate Alexandrium catenella, associated with paralytic shellfish poisoning (PSP), frequently limit commercial shellfish harvests, resulting in serious socio-economic consequences. Although the PSP-inducing species that threaten the most vulnerable commercial species of shellfish are very patchy and spatially heterogeneous in their distribution, the spatial and temporal scales of their effects have largely been ignored in monitoring programs and by researchers. In this study, we examined the spatial and temporal dynamics of PSP toxicity in the clam (Ameghinomya antiqua) in two fishing grounds in southern Chile (Ovalada Island and Low Bay). During the summer of 2009, both were affected by an intense toxic bloom of A. catenella (up to 1.1 × 106 cells L−1). Generalized linear models were used to assess the potential influence of different environmental variables on the field detoxification rates of PSP toxins over a period of 12 months. This was achieved using a four parameter exponential decay model to fit and compare field detoxification rates per sampling site. The results show differences in the spatial variability and temporal dynamics of PSP toxicity, given that greater toxicities (+10-fold) and faster detoxification (20% faster) are observed at the Ovalada Island site, the less oceanic zone, and where higher amounts of clam are annually produced. Our observations support the relevance of considering different spatial and temporal scales to obtain more accurate assessments of PSP accumulation and detoxification dynamics and to improve the efficacy of fisheries management after toxic events.

At the southern tip of South America, evidence of shellfish toxicity has been recorded in the accounts of early explorers and shipwreck survivors since the late 16th Century. Blooms of the toxic dinoflagellate Alexandrium catenella were described in the western Magellan Strait in the early 1970s and have since shown a northward progression through Chilean Patagonia, culminating in a catastrophic toxic event around Chiloé Island in 2016. This shift has taken place through coastal areas of extremely sparse human population density, and anthropogenically driven eutrophication is therefore unlikely to be significantly involved, at least in the south. However, human activities – such as salmon cultivation – may play a role in the intensification of blooms in the more densely populated areas of northern Patagonia. In the fjords and channels of Chilean Patagonia, phytoplankton assemblages are shaped by complex interactions between freshwater (FW) run-off and intrusions of subantarctic surface water (SASW). In the context of blooms of A. catenella, we review the properties of SASW – transformed in coastal waters into modified subantarctic water (MSAW). FW input is characterized by very low concentrations of dissolved inorganic nitrogen (DIN) and phosphorus (DIP), but relatively high concentrations of silicic acid (DSi); DIN and DIP are instead supplied predominantly by SASW which is severely deficient in DSi. These waters therefore show strong vertical gradients in DIN, DIP and DSi, but also potentially in dissolved trace metals and CO2. Large scale shifts in the relative inputs of SASW or FW can modify these vertical gradients, potentially forcing competitive changes in phytoplankton assemblages with latitude, with implications for growth and toxicity of A. catenella and other harmful species. The northward shift of blooms of A. catenella could be associated with anomalies in the Southern Annular Mode (SAM) that modify the influence of MSAW through variations in FW input to coastal waters. The historical presence of blooms in southern Patagonia and Tierra del Fuego, combined with the strongly contrasting conditions with latitude and depth, mean that southern Chile represents an ideal natural laboratory to study climatic and oceanographic influences on dynamics of A. catenella populations.

Blooms of the dinoflagellate Alexandrium catenella , which produces Paralytic Shellfish Toxins (PST), generate serious socio-economic consequences for mariculture in Chile, especially for the production of Mytilus chilensis and other bivalves. Palliative strategies, such as the depuration of mussels in enriched water with chitosan offer encouraging prospects against the advance of contaminated areas and toxin persistence. Adult mussels were fed with A. catenella for 20 days and then were allowed to depurate using chitosan as facilitator, for the same period. Intoxicated mussels showed a reduction in feeding activity and rapid PST accumulation in 20 days (C = 451.5t + 1,673.6, R2 = 0.55 p = 0.008). Not enough evidence was found to indicate a positive effect of chitosan in mussel depuration after 20 days (C = −311.1t + 8,462.4, R2 = 0.8 p = 0.001). At the end of the study, toxicity was higher than 800 μg STX eq kg−1. C2 and GTX4 analogues were the most abundant in the dinoflagellate strain, while C2 and C1 were the most accumulated in mussels. The presence of C1 was notorious during depuration, as the persistence of GTX2,3. GTX5 was only detected in A. catenella , while STX was only present in mussels. Mussel sensitivity to the presence of the toxic dinoflagellate was observed in the present study. The biotransformation, selective elimination and epimerization processes were deduced from intoxication and depuration experiments. • Chitosan was evaluated as chemical strategy to reduce depuration time of PST in mussel M. chilensis. • Not enough evidence to state that chitosan facilitates toxin depuration after 20 days. • Alexandrium catenella in diet reduced some physiological parameters of M. chilensis. [ABSTRACT FROM AUTHOR]

Alexandrium catenella produces paralytic shellfish toxins that affect marine fisheries and aquaculture as well as ecosystem and human health worldwide. This harmful algal species is extremely sensitive to environmental conditions and potentially to future climate change. Using a generalized additive mixed model (GAMM) we studied the potential effects of changing salinity and temperatures on A. catenella bloom (≥1000 cells L–1) occurrence along Canada’s East Coast throughout the 21st century. Our GAMM was applied to two high greenhouse gas emissions scenarios (RCP 8.5) and one mitigation scenario (RCP 4.5). Under present-day conditions, our model successfully predicted A. catenella’s spatio-temporal distribution in Eastern Canada. Under future conditions, all scenarios predict increases in bloom frequency and spatial extent as well as changes in bloom seasonality. Under one RCP 8.5 scenario, A. catenella bloom occurrences increased at up to 3.5 days per decade throughout the 21st century, with amplified year-to-year variability. Blooms expanded into the Gulf of St. Lawrence and onto the Scotian Shelf. These conditions could trigger unprecedented bloom events in the future throughout our study region. In all climate scenarios, the bloom season intensified earlier (May–June) and ended later (October). In some areas of the Gulf of St. Lawrence, the thermal habitat of A. catenella was exceeded, thereby locally reducing bloom risk during the summer months. We conclude that an increase in A. catenella’s environmental bloom window could further threaten marine fauna including endangered species as well as fisheries and aquaculture industries on Canada’s East Coast. Similar impacts could be felt in other coastal regions of the globe where warming and freshening of waters are intensifying.

Harmful algal bloom (HAB) species Alexandrium catenella and Dinophysis acuminata are associated with paralytic shellfish poisoning (PSP) and diarrhetic shellfish poisoning (DSP) in humans, respectively. While PSP and DSP have been studied extensively, less is known about the effects of these HAB species or their associated toxins on shellfish. This study investigated A. catenella and D. acuminata toxicity in a larval oyster (Crassostrea virginica) bioassay. Larval activity and mortality were examined through 96-h laboratory exposures to live HAB cells (10–1000 cells/mL), cell lysates (1000 cells/mL equivalents), and purified toxins (10,000 cells/mL equivalents). Exposure to 1000 cells/mL live or lysed D. acuminata caused larval mortality (21.9 ± 7.0%, 10.2 ± 4.0%, respectively) while exposure to any tested cell concentration of live A. catenella, but not lysate, caused swimming arrest and/or mortality in >50% of larvae. Exposure to high concentrations of saxitoxin (STX) or okadaic acid (OA), toxins traditionally associated with PSP and DSP, respectively, had no effect on larval activity or mortality. In contrast, pectenotoxin-2 (PTX2) caused rapid larval mortality (49.6 ± 5.8% by 48 h) and completely immobilized larval oysters. The results indicate that the toxic effects of A. catenella and D. acuminata on shellfish are not linked to the primary toxins associated with PSP and DSP in humans, and that PTX2 is acutely toxic to larval oysters.

Microalgae host varied microbial consortium harboring cross-kingdom interactions with fundamental ecological significance in aquatic ecosystems. Revealing the complex biofunctions of the cultivable bacteria of phycosphere microbiota is one vital basis for deeply understanding the mechanisms governing these dynamic associations. In this study, a new light-yellow pigmented bacterial strain LZ-28 was isolated from the highly-toxic and harmful algal bloom-forming dinoflagellate Alexandrium catenella LZT09. Collective phenotypic and genotypic profiles were obtained to confidently identify this strain as a new Mameliellaalba member. Comparative genomic analysis showed that strain LZ-28 shared highly similar functional features with other four marine algae-derived M. alba strains in spite of their distinctive isolation sources. Based on the bioactivity assaying, the mutual growth-promoting effects between bacterial strain LZ-28 and algal strain LZT09 were observed. After the culture conditions were optimized, strain LZ-28 demonstrated an extraordinary production ability for its bioflocculanting exopolysaccharides (EPS). Moreover, the portions of two monosaccharides glucose and fucose of the EPS were found to positively contribute to the bioflocculanting capacity. Therefore, the present study sheds light on the similar genomic features among the selected M. alba strains, and it also reveals the potential pharmaceutical, environmental and biotechnological implications of active EPS produced by this new Mameliella alba strain LZ-28 recovered from toxic bloom-forming marine dinoflagellate.

To evaluate how the decay of bloom-forming algae affect the coastal dissolved oxygen, a laboratory simulation was conducted in terms of three typical harmful algae, Alexandrium catenella, Prorocentrum donghaiense, and Skeletonema costatum. Algae of same biomass (55 μg/mL) were conducted in lightproof columns, and the cell density, dissolved oxygen (DO), and ammonia nitrogen of different layers were monitored at certain time series. Results show that the decomposition of algae significantly decreased the DO, and increased the ammonia nitrogen in all layers; and significant deference between different species was observed. The A. catenella treatment showed the lowest DO (average concentration of 3.4 mg/L) and the highest ammonia nitrogen (average concentration of 0.98 mg/L) at the end of test, followed by P. donghaiense; and the S. costatum showed relatively high DO and low ammonia nitrogen due to slow decay rate. Results indicate that decomposition of harmful bloom algae, especially dinoflagellate, would cause significantly DO depletion and toxic ammonia nitrogen increase, which will detrimentally affect both pelagic and benthic ecosystem. [ABSTRACT FROM AUTHOR]

The distribution of growth-inhibiting bacteria (GIB) against the toxic dinoflagellate Alexandrium catenella (Group I) was investigated targeting seagrass leaves and surface waters at the seagrass bed of Akkeshi-ko Estuary and surface waters of nearshore and offshore points of Akkeshi Bay, Japan. Weekly samplings were conducted from April to June in 2011. GIBs were detected from surface of leaves of the seagrass Zostera marina in Akkeshi-ko Estuary (7.5 × 105–4.7 × 106 colony-forming units: CFU g−1 wet leaf) and seawater at the stations in Akkeshi Bay (6.7 × 100–1.1 × 103 CFU mL−1). Sequence analyses revealed that the same bacterial strains with the same 16S rRNA sequences were isolated from the surface biofilm of Z. marina and the seawater in the Akkeshi Bay. We therefore strongly suggested that seagrass beds are the source of algicidal and growth-inhibiting bacteria in coastal ecosystems. Cells of A.catenella were not detected from seawaters in Akkeshi-ko Estuary and the coastal point of Akkeshi Bay, but frequently detected at the offshore point of Akkeshi Bay. It is suggested that A.catenella populations were suppressed by abundant GIBs derived from the seagrass bed, leading to the less toxin contamination of bivalves in Akkeshi-ko Estuary. [ABSTRACT FROM AUTHOR]

Globally, harmful algal blooms (HABs) are an increasing problem. In the Gulf of Maine and Bay of Fundy, blooms of the toxic dinoflagellate Alexandrium catenella are annually recurrent phenomena. As this region is one of the most rapidly warming areas of the global ocean, an improved understanding of the mechanisms driving the initiation of local A. catenella blooms, their interannual variability and the implications of future climate change is critical to local monitoring strategies and marine resources management. A 27-year (1988–2014) time series of weekly A. catenella cell counts from the Bay of Fundy and concurrent satellite-measured sea surface temperature, freshwater discharge from the St. John River and wind-driven turbulence are compared to assess their relationship to variability in bloom phenology metrics. The mean thermal habitat associated with early detection of A. catenella is 6.5 ± 1.6°C, whereas that of bloom initiation averages 9.2 ± 1.5°C. Both thermal habitats for A. catenella are trending earlier over the study period. Bloom initiations that precede the arrival of the thermal habitat mean (occur in colder water) are associated with higher spring freshwater discharge and are generally weaker blooms. Increased spring freshwater discharge is also associated with earlier bloom initiation and earlier maximum concentration dates. No significant relationship was observed with the strength of wind-driven mixing. Removal of the mean thermal seasonal cycle shows that surface temperature anomalies have a strong negative relationship to the bloom phenology metrics and arrival of thermal habitat: warmer years are linked to earlier arrival of thermal habitats (∼12 days °C–1) and earlier detection and bloom initiation dates (∼33 days °C–1). Using these relationships and present trends in Bay of Fundy surface temperature warming over the period 1982–2019, we project the arrival dates of bloom thermal habitat and bloom phenology metrics out to the middle of this century. Based on current rates of sea surface temperature change, bloom phenology metrics (e.g., bloom initiation, early detection), can be expected to shift 1–2 months earlier in the season by mid-century. Such changes in the phenology of A. catenella blooms will need to be incorporated into both monitoring strategies and forecasting models for the region.

A novel Gram-negative, aerobic, motile and short rod-shaped bacterium with exopolysaccharides production, designated as LZ-4T, was isolated from cultivable phycosphere microbiota of harmful algal blooms-causing marine dinoflagellate Alexandrium catenella LZT09 which produces paralytic shellfish poisoning toxins. Strain LZ-4T was able to use thiosulfate (optimum concentration 10 mM) as energy source for bacterial growth. Phylogenetic analysis based on 16S rRNA gene sequences showed that strain LZ-4T belonged to the genus Limnobacter, showing high 16S rRNA gene sequences similarities with L. thiooxidans DSM 13612T (99.4%), L. humi NBRC 11650T (98.2%) and L. litoralis NBRC 105857T (97.2%), respectively. The average nucleotide identity (ANI) and digital DNA-DNA hybridization (dDDH) values between LZ-4T and L. thiooxidans DSM 13612T were 78.9 and 21.9%, respectively. Both values were far lower than the thresholds (95–96% for ANI and 70% for dDDH) generally accepted for new species delineation. The respiratory quinone of strain LZ-4T was Q-8. The dominant cellular fatty acids were determined as summed feature 3 (C16:1ω6c/ω7c), summed feature 8 (C18:1ω6c/ω7c) and C16:0. Polar lipids profile consisted of diphosphatidylglycerol, phosphatidylethanolamine, phosphatidylglycerol, two unidentified aminolipids and three unidentified polar lipids. The genomic DNA G+C content of strain LZ-4T was 52.5 mol%. Based on polyphasic characterization, strain LZ-4T represents a novel species of the genus Limnobacter, for which the name Limnobacter alexandrii sp. nov. is proposed. The type strain is LZ-4T (=CCTCC AB 2019004T =KCTC 72281T). [ABSTRACT FROM AUTHOR]

The effect of an electron beam on the toxic microalgae Cochlodiniumpolykrikoides and Alexandrium catenella, which are involved in the phenomenon of seawater blooming known as red tides, has been studied. This annual disaster causes mass intoxication and death of marine life. Electron-beam irradiation at an absorbed dose of to 1 kGy leads to inactivation of more than 2/3 of the microalgae population due to disruption of cell walls, damage to chloroplasts, and cell aggregation. Radiolysis destroys paralytic toxins, including the most dangerous of them. Because of low doses and the ease of irradiation, the electron-beam treatment of water can be a promising method for the inactivation and detoxification of phytoplankton involved in red tides. [ABSTRACT FROM AUTHOR]

Alexandrium catenella , the main species associated with harmful algal blooms, has progressively increased its distribution through one of the most extensive and highly variable fjord systems in the world. In order to understand this successful expansion, we evaluated the effects of different salinities, light intensity, temperatures, nitrogen (N) forms and nitrogen/phosphate (N:P) ratio levels on the growth performance, using clones isolated from different locations across its wide geographic distribution. Results showed that the growth responses were plastic and, in some cases, different reaction norms among clones were observed. Despite plasticity, the optimal growth of A. catenella (i.e. highest growth rate and highest maximal cells density) was reached within a narrow thermal range (12–15°C), while salinity (20–30 PSU) and light intensity (20–120 μmol m−2 s−1) ranges were wider. These results are partially consistent with the highest cell densities recorded in the field. Furthermore, optimal growth was reached using reduced forms of nitrogen (i.e. urea and NH4+) and in unbalanced N:P ratios (18:1 and 30:1). These characteristics likely allow A. catenella to grow in highly variable environmental conditions and might partly explain the recent expansion of this species. [ABSTRACT FROM AUTHOR]

Individuals of Mytilus platensis were exposed to Alexandrium catenella to evaluate the accumulation and metabolization of paralytic shellfish toxins (PST) over a period of 25 days. Mussels were collected from the intertidal zone of Cerro Avanzado, Argentine Patagonia. After 16 days, the toxins in the tissues of mussels were detected by the methods of mouse bioassay and high performance liquid chromatography with fluorometric detection (HPLC-FDL). The accumulation kinetics of PST toxins in M. platensis fed with A. catenella fitted to a linear function, in which the accumulation rate was 31.2 µg STX eq kg−1 day−1. After 16 days, the PST toxin level in tissues of mussels reached 1178 µg STX eq kg−1 exceeding the safety limit for human consumption (800 µg STX eq kg−1 tissue), whereas the highest PST toxin level was reached at the end of the experimentation (1613 µg STX eq kg−1) at 25 days. Differences in the toxin profile of the dinoflagellates and the tissues of the mussels confirmed biotransformation of PST in the mussel digestive system. The toxin profile of M. platensis was dominated by the gonyautoxins GTX1 and GTX4, while the toxin profile of A. catenella was dominated by the N-sulfocarbamoyl toxin C2. To our knowledge, this is the first experimentation on a laboratory scale of PST toxins accumulation in M. platensis with a native strain of A. catenella of Argentine Patagonia. [ABSTRACT FROM AUTHOR]

Alexandrium catenella is one of the globally distributed toxic marine microalgae to cause paralytic shellfish poisoning that poses a great threat to marine fisheries, economy, and public health. Development of efficient and sensitive methods for accurate identification of A. catenella to minimize its damage is therefore necessary. In this study, a novel method referred to as loop-mediated isothermal amplification (LAMP) combined with lateral flow dipstick (LFD) (LAMP-LFD) was established for rapid and sensitive detection of A. catenella. The internal transcribed spacer (ITS) gene of A. catenella was cloned for sequencing and used as target for LAMP-LFD. Three sets of LAMP primers (AcLF1, AcLF2, and AcLF3) targeting the ITS were successfully designed, among which AcLF2 displaying the best performance was used in the subsequent tests. A specific LFD probe targeting the amplification region of AcLF2 was further designed. The LAMP-LFD detection system was established and the amplification conditions were optimized. Cross-reactivity tests with common marine microalgae showed that the LAMP-LFD was exclusively specific for A. catenella. The detection limits of LAMP-LFD for A. catenella genomic DNA and the plasmid containing the ITS were 4.63 × 10−4 ng μL−1 and 1.26 × 104 copies μL−1, displaying a sensitivity that is 10 times higher than that of SYBR Green I assay and 100 times higher than that of conventional PCR, respectively. Finally, the practicability of LAMP-LFD was confirmed by test with spiked samples. LAMP-LFD revealed a detection limit of approximately 0.1 cell mL–1, which was 100 times more sensitive than conventional PCR. The optimized LAMP-LFD protocol can be completed within 75 min. Therefore, the established LAMP-LFD is a specific, sensitive, and rapid method that is possibly applicable to the field monitoring of A. catenella. [ABSTRACT FROM AUTHOR]

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The producer of paralytic shellfish toxin (PST), Alexandrium catenella , is one of the main generators of HABs in the coasts of Chile. Its presence produces ecological and economic damage, directly affecting filter-feeding organisms, and indirectly to other organism through the trophic chain. The objective of this research was to identify the effect of a toxic diet on the energetic and reproductive parameters of the carnivorous snail Chorus giganteus. Two groups of snails were used, one fed with toxic prey (bivalves fed with A. catenella), and the other fed with non-toxic prey. Both treatments were maintained under these conditions for 63 days, then, elemental composition (C, N) and energy content were estimated, and fecundity parameters were analyzed. The results indicate that snails fed with toxic prey had a lower percentage of C and C/N ratio. The energy content was significantly lower in intoxicated snails. Regarding fecundity parameters, a higher number of egg-masses were produced by toxic snails, however, only 62% of these showed embryonic development, with 57% hatching success. A negative relationship was identified between the mean PST concentration, quantified in snails, and the number of egg-masses produced per aquarium. In the aquarium where the snails had highest average PST concentration (1200 ± 820 μg STX.2HCL eq. Kg−1) there was no oviposition, while egg-masses were only produced by snails in aquaria where the average concentration did not exceed 360 ± 160 μg STX.2HCL eq. Kg−1. It is likely that, with low levels of accumulated PST, C. giganteus activates its oviposition process as a response to toxin-induced stress, generating a higher energy expenditure supported by a redirection of its reserves. However, when the intoxication presents higher levels, the reproductive process could be inhibited, similar to what has been identified in other molluscs. • Ingestion of paralytic shellfish toxin reduces the energy content in marine snail. • Gastropods with low levels of saxitoxin increase their fecundity. • Paralytic shellfish toxin effect on gastropods fecundity depends on its concentration. [ABSTRACT FROM AUTHOR]

We employed a detection method to quantify Alexandrium catenella (Group I), one of the causative species for paralytic shellfish poisoning (PSP) in Jinhae-Masan Bay, Korea, targets sxtA4 , via chip-based digital PCR. Additionally, we explored the dynamics of Alexandrium during the spring of 2022 using an rDNA-based quantitative PCR (qPCR) assay to enhance the performance of the dPCR assay. In matching dPCR results with PSP monitoring reports, we optimized a cell regulatory threshold of 102 cells L−1, the maximum cell density when shellfish harvesting was permitted, for the dPCR assay. This threshold functioned similar to the PST threshold used in mouse bioassays (MBAs). Furthermore, we validated a total concordance rate of 83.8 % between the two assays for 2020–2022, reaching a maximum of 96.2 % in 2020. Thus, the result of dPCR could complement MBAs, facilitating the early detection of PSP outbreaks. • We optimized a cell regulatory threshold of 102 cells L−1. • We validated a total concordance rate of 83.8 % between dPCR assay and MBAs. • Thus, dPCR assay can complement MBAs. [ABSTRACT FROM AUTHOR]

• A highly toxic outbreak occurred in the Beagle Channel, with 197,266 µg STXeq PST kg−1 mussel tissue. • PST in birds and terrestrial fauna accompanied an unprecedented multispecies mortality event. • PST toxins as determined via HPLC may accumulate and biotransform along the pelagic food web. • The wasp-waist structure of the pelagic food web could facilitate toxin spread. The Beagle Channel is a Subantarctic semi-estuarine environment at the southern tip of South America, where intoxication events associated with harmful algal blooms have been reported since 1886, including a world record in toxicity due to Alexandrium catenella in 1992. Toxic algae affect public health and ecosystem services, particularly mussel aquaculture and fisheries management. During the austral summer of 2022, an intense bloom of A. catenella (5 × 104 cells L−1) occurred in the Beagle Channel, leading to the second most toxic event in the area, with mussel toxicity reaching 197,266 µg STXeq kg−1. This event was synchronous with the mortality of marine organisms from different trophic levels and terrestrial fauna, i.e., two Fuegian red foxes and a southern caracara. Stomach content and liver samples from dead kelp gulls (Larus dominicanus), Magellanic penguins (Spheniscus magellanicus), papua penguins (Pygoscelis papua), and imperial cormorants (Leucocarbo atriceps), presented variable paralytic shellfish toxins (PST) levels (up to 3427 µg STXeq kg−1) as measured by high performance liquid chromatography (HPLC), suggesting that deaths were associated with high PST toxicity level. The different toxin profiles found in phytoplankton, zooplankton, squat lobsters (Grimothea gregaria), Fuegian sprat (Sprattus fuegensis), and seabirds evidenced possible toxin transformation along the food web and the possible transfer vectors. The unexpected detection of PST in terrestrial fauna (up to 2707 µg STXeq kg−1) suggested intoxication by scavenging on squat lobsters, which had high toxicity (26,663 µg STXeq kg−1). PST trace levels were also detected in a liver sample of a dead false killer whale (Pseudorca crassidens), an oceanic odontocete stranded on the coast during the bloom. Overall, our results denote the exceptional nature of the toxic, multispecies mortality event and that toxins may propagate to several levels of the food web in this Subantarctic environment. [ABSTRACT FROM AUTHOR]

In southern Chile, Alexandrium catenella is the main species generating harmful algal blooms (HABs) and over time it has expanded its range since it was first recorded in the Magallanes region in 1972. In 2016 a severe bloom of an Alexandrium species occurred, which was notable for its intensity and geographical extent, extending into new areas to the north of the Patagonian fjords including areas along the open Pacific Ocean coast. Given the exceptional nature of this event, we verified the taxonomic classification of the species that generated the bloom and evaluated the influence of the range expansion process on its genetic structure and population diversity. This was achieved by isolating clones collected in 2014 from cyst beds located at the northern limit of its then known distribution, along with clones isolated from the water samples taken during the 2016 bloom. These clones were characterized genetically with LSU rDNA and AFLPs molecular markers. Phylogenetic analyses showed that all clones were aggregated in the Group I of the A. tamarense species complex, which confirmed that A. catenella was the species responsible for the 2016 bloom. High genetic diversity was observed within populations though there were no significant differences between populations. Furthermore, genetic structure showed an isolation by distance relationship among populations, and several analyses consistently indicated a high divergence among the population groups derived from both cysts and vegetative cells. Despite this study not finding the patterns expected for species range expansion (i.e., diversity gradient and/or high population divergence), the genetic diversity and structure indicated that these were influenced by the geographic distance effect and the physical restrictions to gene flow, along with the demographic processes that occurred during the distinct phases of the life cycle of A. catenella.

Toxic dinoflagellate Alexandrium spp. produce saxitoxins (STXs), whose biosynthesis pathway is affected by temperature. However, the link between the regulation of the relevant genes and STXs’ accumulation and temperature is insufficiently understood. In the present study, we evaluated the effects of temperature on cellular STXs and the expression of two core STX biosynthesis genes (sxtA4 and sxtG) in the toxic dinoflagellate Alexandrium catenella Alex03 isolated from Korean waters. We analyzed the growth rate, toxin profiles, and gene responses in cells exposed to different temperatures, including long-term adaptation (12, 16, and 20 °C) and cold and heat stresses. Temperature significantly affected the growth of A. catenella, with optimal growth (0.49 division/day) at 16 °C and the largest cell size (30.5 µm) at 12 °C. High concentration of STXs eq were detected in cells cultured at 16 °C (86.3 fmol/cell) and exposed to cold stress at 20→12 °C (96.6 fmol/cell) compared to those at 20 °C and exposed to heat stress. Quantitative real-time PCR (qRT-PCR) revealed significant gene expression changes of sxtA4 in cells cultured at 16 °C (1.8-fold) and cold shock at 20→16 °C (9.9-fold). In addition, sxtG was significantly induced in cells exposed to cold shocks (20→16 °C; 19.5-fold) and heat stress (12→20 °C; 25.6-fold). Principal component analysis (PCA) revealed that low temperature (12 and 16 °C) and cold stress were positively related with STXs’ production and gene expression levels. These results suggest that temperature may affect the toxicity and regulation of STX biosynthesis genes in dinoflagellates.

The effects of co-occurring harmful algal blooms (HABs) on marine organisms is largely unknown. We assessed the individual and combined impacts of the toxin producing HABs, Alexandrium catenella and Dinophysis acuminata , and a non-toxin-producing HAB (Gymnodinium instriatum) on early life stages of two estuarine fish species (Menidia beryllina and Cyprinodon variegatus). Lethal (i.e. time to death) and sublethal (i.e. growth, grazing rate, and swimming activity) effects of cultured HABs were investigated for eleutheroembryo and larval life stages. Mixed algal treatments (i.e. A. catenella and D. acuminata mixtures) were often equally toxic as A. catenella monoculture treatments alone, although responses depended on the fish species and life stage. Fish exposed to toxin producing HABs died significantly sooner (i.e. <1–3 days) than controls. Significant differences in sublethal effects were also found between fed controls and toxic HAB treatments, although responses were often similar to G. instriatum or starved controls. Collectively, the results demonstrate that HABs may reduce fish productivity and fitness. [ABSTRACT FROM AUTHOR]

Alexandrium catenella is a cosmopolitan harmful algal bloom (HAB) forming dinoflagellate. To comprehend its mechanisms of bloom formation, gene expression analysis is indispensable. Quantitative real-time reverse transcription PCR (qRT-PCR) is an ideal method for swift and precise quantification of gene expression analysis that greatly relies on selection of apposite reference genes for data normalization. To date, limited studies have focused on the screening of reference genes in dinoflagellates. The unavailability of valid reference gene for normalizing poses hindrances in the appliance of qRT-PCR to the HAB forming group. The work presented here evaluated 12 reference genes for their expression stability using qRT-PCR under diverse nutritional conditions together with high light. Statistical algorithm such as RefFinder was used that analyze data using geNorm and NormFinder, comparative delta-CT method along with a combination approach that declares comprehensive ranking contingent upon the geometric mean of the results procured from other methods. Comprehensive analysis across all condition by geNorm showed ACT, IPP, and GAPDH as genes possessing highest stability followed by Tubα and ICDH. Comprehensive analysis by Normfinder declared that IPP is the most stable gene followed by ACT, CYC, GAPDH, and GTEF. Combination approach using comparative delta CT, geNorm and NormFinder analysis programs through RefFinder as well as ranking by standard deviation of delta CT declared IPP as the most stable gene followed by ACT and GAPDH. IPP, ACT, and GAPDH represented the top 4 listed stable reference genes among all the analysis methods used. The results of pairwise variation by geNorm showed that V2/V3 value under all the tested conditions was below the cut-off value of 0.15 which shows that two genes are sufficient for normalization. Our results in accord with other widely conducted studies emphasize the significance of reference gene validation in precise experimental setup before appliance to avoid serious misinterpretation of the results. It is highly expected that the procedures and outcome of the report may be helpful for future studies in the gene expression analysis of A. catenella and to screen out reference genes for other algae. [ABSTRACT FROM AUTHOR]

• Compared control of toxin production of a major dinoflagellate species by nutrients, prey alarm cues and grazers. • Neither ammonium nor urea increased cell toxin production. • Surprisingly alarm cues from interspecific prey increased cell toxin production. • Grazer-induced toxin production was an order of magnitude greater than nutrient or prey alarm cues. • Grazer-induced toxin production appears to be a key control of prey toxicity. Dinoflagellate paralytic shellfish toxin (PST) production is mediated by several abiotic and biotic factors. This study compared the relative importance of nitrogen source and concentration, prey alarm cues and grazer presence on toxin production of the marine dinoflagellate Alexandrium catenella (Group I, strain BF-5). In separate assays run under either nutrient-replete (F/2 medium) or nutrient-depleted (filtered seawater) conditions, PST production of A. catenella was measured as a function of varying concentrations of added nitrogen sources (ammonium and urea), alarm cues from lysed conspecific (A. catenella Group I strains) and interspecific (the diatom, Thalassiosira weissflogii, and the green flagellate, Tetraselmis sp.) algae, and the presence of a grazer (the copepod Acartia hudsonica). Results showed that addition of ammonium or urea did not increase PST production. Unexpectedly, interspecific alarm cues increased toxin production but conspecific ones did not. Grazer presence dramatically induced PST production in A. catenella, irrespective of nutrient conditions, and this effect was an order of magnitude greater than any of the other variables tested. These results corroborate previous studies on grazer-induced PST production, and support the hypothesis that grazer-induced toxin production is not an experimental artifact, but rather a prey defense mechanism. [ABSTRACT FROM AUTHOR]

Abstract Phosphorus (P) is a key macronutrient limiting cell growth and bloom formation of marine dinoflagellates. Physiological responses to changing ambient P have been investigated in dinoflagellates; however, the molecular mechanisms behind these responses remain limited. Here, we compared the protein expression profiles of a marine dinoflagellate Alexandrium catenella grown in inorganic P-replete, P-deficient, and inorganic- and organic-P resupplied conditions using an iTRAQ-based quantitative proteomic approach. P deficiency inhibited cell growth and enhanced alkaline phosphatase activity (APA) but had no effect on photosynthetic efficiency. After P resupply, the P-deficient cells recovered growth rapidly and APA decreased. Proteins involved in sphingolipid metabolism, organic P utilization, starch and sucrose metabolism, and photosynthesis were up-regulated in the P-deficient cells, while proteins associated with protein synthesis, nutrient assimilation and energy metabolism were down-regulated. The responses of the P-deficient A. catenella to the resupply of organic and inorganic P presented significant differences: more biological processes were enhanced in the organic P-resupplied cells than those in the inorganic P-resupplied cells; A. catenella might directly utilize G-6-P for nucleic acid synthesis through the pentose phosphate pathway. Our results indicate that A. catenella has evolved diverse adaptive strategies to ambient P deficiency and specific mechanisms to utilize dissolved organic P, which might be an important reason resulting in A. catenella bloom in the low inorganic P environment. Biological significance The ability of marine dinoflagellates to utilize different phosphorus (P) species and adapt to ambient P deficiency determines their success in the ocean. In this study, we investigated the response mechanisms of a dinoflagellate Alexandrium catenella to ambient P deficiency, and resupply of inorganic- and organic-P at the proteome level. Our results indicated that A. catenella initiated multiple adaptive strategies to ambient P deficiency, e.g. utilizing nonphospholipids and glycosphingolipids instead of phospholipids, enhancing expression of acid phosphatase to utilize organic P, and reallocating intracellular energy. Proteome responses of the P-deficient A. catenella to resupply of inorganic- and organic-P differed significantly, indicating different utilization pathways of inorganic and organic P, A. catenella might directly utilize low molecular weight organic P, such as G-6-P as both P and carbon sources. Graphical abstract Unlabelled Image Highlights • A. catenella evolved diverse adaptive mechanisms to P-deficiency. • G-6-P provided a source of P and carbon for A.catenella. [ABSTRACT FROM AUTHOR]

Abstract The neurotoxic complex saxitoxin, is a group of marine toxins that historically has significantly impacted human health and the ability to utilize marine resources. A steady increase in the distribution and intensity of Alexandrium catenella blooms in Chile, and around the world, has caused major ecological and socioeconomic impacts, putting this type of dinoflagellate, and its toxicity, in the spotlight. Ostrea chilensis is a commercially and ecologically important resource harvested from wild populations and farmed in centers of southern Chile, where it is exposed to large harmful algal blooms of the type that can cause paralysis in humans. This study contributes to our understanding about the transfer of toxins from A. catenella cells to juvenile and adult Ostrea chilensis by tracking transformations of the neurotoxic complex until it reaches its most stable molecular form in the intracellular environment of O. chilensis tissues. These biotransformations are different in O. chilensis juveniles and adults, indicating a differentiated response for these two life stages of this bivalve species. These studies can be used for similar analyses in other ecologically and commercially important species of filter feeding organisms, providing greater understanding of the specific interactions of bivalves in scenarios of toxic dinoflagellate proliferations (e.g. A. catenella blooms). Highlights • The toxin profile of Alexandrium catenella is mainly represented by C2 and GTX4. • Juvenile and adults of Ostrea chilensis present different pathways of PST biotransformation. • O. chilensis accumulates and biotransform toxins produced by A. catenella , acting as a vector towards higher trophic levels. [ABSTRACT FROM AUTHOR]