Screening of ApDOT1.9 interacting proteins and the potential function of interactor ApSNARE in the rapid growth regulation of Alexandrium pacificum.

Alexandrium pacificum is a toxic dinoflagellate resulting in harmful algal blooms (HABs). ApDOT1.9 is a methyltransferase involved in the rapid growth regulation of A. pacificum, but its protein interaction information is still limited. In this study, 14 candidate interacting proteins of ApDOT1.9, which were involved in metabolism, genetic information processing, environmental information processing and cellular processes, were screened. The interaction between candidate interactor ApSNARE (Soluble N-ethylmaleimide-sensitive factor attachment protein receptors of Alexandrium pacificum) and ApDOT1.9 was further validated by molecular docking and GST (Glutathione S transferase) pull-down. The relevant biological functional information and gene expression of ApSNARE were also analyzed and detected. These results indicate that ApSNARE was an interactor of ApDOT1.9 and it may also participate in A. pacificum rapid growth regulation under high light or high nitrogen conditions, which will provide preliminary information on the interaction proteins of ApDOT1.9 and molecular regulation mechanisms of growth in A. pacificum.

The identity of Centrodinium elongatum, type species of the dinoflagellate genus Centrodinium (Dinophyceae), and a review on the synonymy of allied species.

The planktonic dinoflagellate genus Centrodinium has been understudied, with the type species C. elongatum remaining undocumented since the original description. Here, we report C. elongatum isolated from Mazatlán, Mexican Pacific. In the chains, the posterior daughter cell with an incomplete apical horn shows the morphology of C. elongatum, while the anterior daughter cell with complete epitheca corresponds to C. pulchrum. For the first time, a species of Centrodinium sensu stricto (highly laterally flattened species with horns) was cultured. An unarmored life stage, known as Murrayella ovalis, derived from the spheroplast after ecdysis. In the rDNA molecular phylogenies, C. elongatum (=C. pulchrum) nested as basal to morphologically similar species (C. eminens and C. intermedium) and as a sister group of a former Murrayella species, C. punctatum. C. elongatum differs from C. eminens and C. intermedium in the chain formation, second apical (2') plate not being divided, horns with coarse poroid ornamentation, and missing prominent distal spinules. The taxonomy of Centrodinium sensu stricto is revised, with a discussion in the identities of C. complanatum, C. eminens, and C. maximum. The name C. deflexum is restored as a senior synonym of C. intermedium and C. ovale.

Allelochemicals determine competition and grazing control in Alexandrium catenella.

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.

Toxic effects of the emerging Alexandrium pseudogonyaulax (Dinophyceae) on multiple trophic levels of the pelagic food web.

The dinoflagellate Alexandrium pseudogonyaulax, a harmful algal bloom species, is currently appearing in increasing frequency and abundance across Northern European waters, displacing other Alexandrium species. This mixotrophic alga produces goniodomins (GDs) and bioactive extracellular substances (BECs) that may pose a threat to coastal ecosystems and other marine resources. This study demonstrated the adverse effects of A. pseudogonyaulax on four marine trophic levels, including microalgae (Rhodomonas salina), microzooplankton (Polykrikos kofoidii) and mesozooplankton (Acartia tonsa), as well as fish gill cells (RTgill-W1, Oncorhynchus mykiss), ultimately leading to enhanced mortality and cell lysis. Furthermore, cell-free supernatants collected from A. pseudogonyaulax cultures caused complete loss of metabolic activity in the RTgill-W1 cell line, indicating ichthyotoxic properties, while all tested GDs were much less toxic. In addition, cell-free supernatants of A. pseudogonyaulax led to cell lysis of R. salina, while all tested GDs were non-lytic. Finally, reduced egg hatching rates of A. tonsa eggs exposed to cell-free supernatants of A. pseudogonyaulax and impaired mobility of P. kofoidii and A. tonsa exposed to A. pseudogonyaulax were also observed. Altogether, bioassay results suggest that the toxicity of A. pseudogonyaulax is mainly driven by BECs and not by GDs, although further research into factors modulating the lytic activity of Alexandrium spp. are needed.

Variability in Paralytic Shellfish Toxin Profiles and Dinoflagellate Diversity in Mussels and Seawater Collected during Spring in Korean Coastal Seawater.

Paralytic shellfish toxins (PSTs) are potent neurotoxins produced by certain microalgae, particularly dinoflagellates, and they can accumulate in shellfish in coastal seawater and thus pose significant health risks to humans. To explore the relationship between toxicity and PST profiles in seawater and mussels, the spatiotemporal variations in PST concentrations and profiles were investigated along the southern coast of Korea under peak PST levels during spring. Seawater and mussel samples were collected biweekly from multiple stations, and the toxin concentrations in the samples were measured. Moreover, the dinoflagellate community composition was analyzed using next-generation sequencing to identify potential PST-producing species. The PST concentrations and toxin profiles showed substantial spatiotemporal variability, with GTX1 and GTX4 representing the dominant toxins in both samples, and C1/2 tending to be higher in seawater. Alexandrium species were identified as the primary sources of PSTs. Environmental factors such as water temperature and salinity influenced PST production. This study demonstrates that variability in the amount and composition of PSTs is due to intricate ecological interactions. To mitigate shellfish poisoning, continuous monitoring must be conducted to gain a deeper understanding of these interactions.

The ciliate Euplotes balteatus is resistant to Paralytic Shellfish Toxins from Alexandrium minutum (Dinophyceae).

Research on interactions between grazers and toxigenic algae is fundamental for understanding toxin dynamics within aquatic ecosystems and developing biotic approaches to mitigate harmful algal blooms. The dinoflagellate Alexandrium minutum is a well-known microalga responsible for paralytic shellfish toxins (PSTs) contamination in many coastal regions worldwide. This study investigated the impact of the ciliate Euplotes balteatus on cell density and PSTs transfer in simulated A. minutum blooms under controlled conditions. E. balteatus exhibited resistance to the PSTs produced by A. minutum with a density of up to 10,000 cells/mL, sustaining growth and reproduction while eliminating algal cells within a few days. The cellular PSTs content of A. minutum increased in response to the grazing pressure from E. balteatus. However, due to the substantial reduction in density, the overall toxicity of the algal population decreased to a negligible level. Most PSTs contained within algal cells were temporarily accumulated in E. balteatus before being released into the water column, suggesting unclear mechanisms for PSTs excretion in unicellular grazers. In principle, the grazing of E. balteatus on A. minutum promotes the transfer of the majority of intracellular PSTs into extracellular portions, thereby mitigating the risk of their accumulation and contamination through marine trophic pathways. However, this process also introduces an increase in the potential environmental hazards posed by extracellular PSTs to some extent.

Recovering sedimentary ancient DNA of harmful dinoflagellates accumulated over the last 9000 years off Eastern Tasmania, Australia.

Harmful algal blooms (HABs) have had significant adverse impacts on the seafood industry along the Tasmanian east coast over the past 4 decades. To investigate the history of regional HABs, we performed analyses of sedimentary ancient DNA (sedaDNA) in coastal sediments up to ~9000 years old collected inshore and offshore of Maria Island, Tasmania. We used metagenomic shotgun sequencing and a hybridisation capture array ("HABbaits1") to target three harmful dinoflagellate genera, Alexandrium, Gymnodinium, and Noctiluca. Bioinformatic and DNA damage analyses verified the authenticity of the sedaDNA sequences. Our results show that dinoflagellates of Alexandrium genera have been present off eastern Tasmania during the last ~8300 years, and we sporadically detected and unambiguously verified sequences of Gymnodinium catenatum that were present offshore up to ~7600 years ago. We also recovered sedaDNA of the fragile, soft-bodied Noctiluca scintillans with increased relative abundance since 2010, consistent with plankton surveys. This study enabled us to identify challenges of sedaDNA sequence validation (in particular for G. catenatum, a microreticulate gymnodinoid species) and provided guidance for the development of tools to monitor past and present HAB species and improvement of future HAB event predictions.

Biogeochemical conditions controlling the intensity of paralytic shellfish poisoning (PSP) outbreak caused by Alexandrium blooms: Results from 6-year field observations in Jinhae-Masan Bay, Korea.

Previous field observations from 2018 to 2019 revealed that paralytic shellfish poisoning (PSP) caused by the blooms of toxic dinoflagellate Alexandrium species occurred under low concentrations of dissolved inorganic nitrogen (DIN) and high concentrations of dissolved organic nitrogen (DON) and humic-like fluorescent dissolved organic matter (FDOMH) in Jinhae-Masan Bay, Korea. In this study, we obtained more data for DIN, DON, FDOMH, and Alexandrium cell density from 2020 to 2023 to further validate environmental conditions for the PSP outbreak. We also measured total hydrolyzed amino acids (THAA) to determine the bioavailability of DON fueling the PSP outbreak. Over the 6-year observations, there was a consistent pattern of low DIN concentrations and high DON and FDOMH concentrations during the PSP outbreak periods. The Alexandrium cell densities, together with the PSP toxin concentrations, increased rapidly under this environmental condition. The PSP outbreak occurs when a large amount of DIN originating from the stream waters near the upstream sites is transformed into DON by biological production before entering the PSP outbreak area. The produced DON is characterized by high bioavailability based on the various AA-derived indices (enantiomeric ratio, degradation index, non-protein AA mole%, and nitrogen-normalized AA yield). In addition, the intensities of PSP outbreaks are mainly dependent on the conversion stage of DIN to DON and enhanced FDOMH. We found that the strong PSP outbreak occurred consistently under a low level of DIN (<1.0 µM) and high levels of DON (>9.0 µM) and FDOMH (>1.5 R.U.). Thus, our results suggest that the monitoring data of environmental conditions can be used to predict the PSP outbreak in the coastal oceans.

The effects of the toxic dinoflagellate Alexandrium on feeding, reproduction and mortality of the copepod Acartia: A systematic review employing weighted linear models.

The study of interactions between copepods of the genus Acartia and toxic dinoflagellates of the genus Alexandrium has been an important topic during the last four decades. Feeding behavior and physiological responses of copepods have been studied in laboratory and field experiments, sometimes with contradictory results. More recently, an evolutionary adaptive mechanism leading to enhanced tolerance of Alexandrium toxins in a population of Acartia experiencing chronic exposure to these dinoflagellates has been reported. In the present work, we collected data from the existing studies on the effects of Alexandrium on feeding, reproduction and mortality of Acartia. With these data, we performed a systematic review consisting of a secondary analysis employing general or generalized linear models, weighting data from different studies by the reciprocal of their standard deviation. Our first aim was to overcome shortcomings of individual studies: limited ranges of the variables and overlooked variables (experiment length, population adaptation). These shortcomings could have led to inconsistent conclusions by missing heterogeneous patterns in copepod responses and in the interactions between variables. Our second aim was to test the enhanced physiological performance of chronically exposed relative to naïve copepod populations over a wide geographic range. We found that the feeding rate is enhanced by increased food biomass, irrespective of the food type. Toxins do not have a clear effect on egg production and have a bi-phasic effect on egg hatching success, which was negative above a specific threshold. Toxins also increased mortality. Experiment length had a positive effect on egg production and negative on egg hatching. Naïve copepod populations showed consistently lower ingestion of Alexandrium and egg hatching rates, thereby supporting the spread of the aforementioned mechanism across populations over a wide geographic range.

The Effecting Mechanisms of 100 nm Sized Polystyrene Nanoplastics on the Typical Coastal Alexandrium tamarense.

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.

Climate Change Stressors, Phosphate Limitation, and High Irradiation Interact to Increase Alexandrium minutum Toxicity and Modulate Encystment Rates.

The changes in the cell physiology (growth rate, cell size, and cell DNA content), photosynthetic efficiency, toxicity, and sexuality under variable light and nutrient (phosphates) conditions were evaluated in cultures of the dinoflagellate Alexandrium minutum obtained from a red tide in the Ría de Vigo (NW Spain). The cells were grown at low (40 and 150 µE m-2 s-1), moderate (400 µE m-2 s-1), and high (800 µE m-2 s-1) light intensities in a medium with phosphate (P+) and without (P-). Cultures were acclimated to the irradiance conditions for one week, and the experiment was run for ~1 month. The cell size and DNA content were monitored via flow cytometry. Two different clonal strains were employed as a monoculture (in a P- or P+ medium) or, to foster sexuality and resting cyst formation, as a mixed culture (only in a P- medium). A. minutum growth was favored by increasing light intensities until 400 µE m-2 s-1. The DNA content analyses indicated the accumulation of S-phase cells at the highest light intensities (400 and 800 µE m-2 s-1) and therefore the negative effects on cell cycle progression. Only when the cells were grown in a P- medium did higher light intensities trigger dose-dependent, significantly higher toxicities in all the A. minutum cultures. This result suggests that the toxicity level is responsive to the combined effects of (high) light and (low) P stress. The cell size was not significantly affected by the light intensity or P conditions. The optimal light intensity for resting cyst formation was 150 µE m-2 s-1, with higher irradiances reducing the total encystment yield. Encystment was not observed at the lowest light intensity tested, indicative of the key role of low-level irradiance in gamete and/or zygote formation, in contrast to the stressor effect of excessive irradiance on planozygote formation and/or encystment.

Assessing the global distribution and risk of harmful microalgae: A focus on three toxic Alexandrium dinoflagellates.

Harmful dinoflagellates and their resulting blooms pose a threat to marine life and human health. However, to date, global maps of marine life often overlook harmful microorganisms. As harmful algal blooms (HABs) increase in frequency, severity, and extent, understanding the distribution of harmful dinoflagellates and their drivers is crucial for their management. We used MaxEnt, random forest, and ensemble models to map the habitats of the representative HABs species in the genus Alexandrium, including A. catenella, A. minutum, and A. pacificum. Since species occurrence records used in previous studies were solely morphology-based, potentially leading to misidentifications, we corrected these species' distribution records using molecular criteria. The results showed that the key environmental drivers included the distance to the coastline, bathymetry, sea surface temperature (SST), and dissolved oxygen. Alexandrium catenella thrives in temperate to cold zones and is driven by low SST and high oxygen levels. Alexandrium pacificum mainly inhabits the Temperate Northern Pacific and prefers warmer SST and lower oxygen levels. Alexandrium minutum thrives universally and adapts widely to SST and oxygen. By analyzing the habitat suitability of locations with recorded HAB occurrences, we found that high habitat suitability could serve as a reference indicator for bloom risk. Therefore, we have proposed a qualitative method to spatially assess the harmful algae risk according to the habitat suitability. On the global risk map, coastal temperate seas, such as the Mediterranean, Northwest Pacific, and Southern Australia, faced higher risks. Although HABs currently have restricted geographic distributions, our study found these harmful algae possess high environmental tolerance and can thrive across diverse habitats. HAB impacts could increase if climate changes or ocean conditions became more favorable. Marine transportation may also spread the harmful algae to new unaffected ecosystems. This study has pioneered the assessment of harmful algal risk based on habitat suitability.

A comparative analysis of real-time quantitative PCR and metabarcoding methods for eDNA-based detection of the toxic dinophyte Alexandrium tamiyavanichii (Dinophyceae).

The marine dinophyte Alexandrium tamiyavanichii is a toxigenic species that produces a group of neurotoxins that is responsible for paralytic shellfish poisoning in humans. Early detection of the species is essential for efficient monitoring. Harmful microalgal monitoring systems have evolved over the years with the advent of environmental DNA (eDNA)-based species detection techniques. In this study, eDNA samples were collected from a large-scale sampling covering the southern South China Sea. The sensitivity and specificity of metabarcoding of the V4 and V9 18S ribosomal DNA barcodes by high-throughput sequencing (HTS) were compared to the species-specific real-time qPCR targeting the A. tamiyavanichii ITS2 region. Environmental samples were screened for A. tamiyavanichii by qPCR (n = 43) and analyzed with metabarcoding (n = 30). Our results revealed a high occupancy profile across samples for both methods; 88% by qPCR, and 80-83% by HTS. When comparing the consistency between the two approaches, only two samples out of 30 were discordant. The V4 and V9 molecular units detected in each sample were positively correlated with the qPCR ITS2 gene copies (V4, rs = 0.67, p < 0.0001; V9, rs = 0.65, p < 0.0001), indicating that metabarcoding could be used as a useful tool for early detection of the species. Our results also revealed that the estimation of A. tamiyavanichii cell abundances based on the HTS read abundances was comparable to that of the qPCR quantification. For long-term monitoring, metabarcoding could serve as a cost-effective screening of detecting not only single HAB species but also simultaneously detecting a multitude of potentially harmful species, which is valuable in informing the subsequent implementation of species-specific monitoring strategies.

Environmental Factors Modulate Saxitoxins (STXs) Production in Toxic Dinoflagellate Alexandrium: An Updated Review of STXs and Synthesis Gene Aspects.

The marine dinoflagellate Alexandrium is known to form harmful algal blooms (HABs) and produces saxitoxin (STX) and its derivatives (STXs) that cause paralytic shellfish poisoning (PSP) in humans. Cell growth and cellular metabolism are affected by environmental conditions, including nutrients, temperature, light, and the salinity of aquatic systems. Abiotic factors not only engage in photosynthesis, but also modulate the production of toxic secondary metabolites, such as STXs, in dinoflagellates. STXs production is influenced by a variety of abiotic factors; however, the relationship between the regulation of these abiotic variables and STXs accumulation seems not to be consistent, and sometimes it is controversial. Few studies have suggested that abiotic factors may influence toxicity and STXs-biosynthesis gene (sxt) regulation in toxic Alexandrium, particularly in A. catenella, A. minutum, and A. pacificum. Hence, in this review, we focused on STXs production in toxic Alexandrium with respect to the major abiotic factors, such as temperature, salinity, nutrients, and light intensity. This review informs future research on more sxt genes involved in STXs production in relation to the abiotic factors in toxic dinoflagellates.

Toxin production and transcriptomic response to nitrate concentrations in the toxic dinoflagellate Alexandrium tamarense.

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.

Development of saxitoxin biosynthesis gene sxtB-targeted qPCR assay for the quantification of toxic dinoflagellates Alexandrium catenella (group I) and A. pacificum (group IV) occurring in the Korean coast.

Toxic dinoflagellate Alexandrium can produce saxitoxins (STXs) and cause paralytic shellfish poisoning (PSP), and thus they are monitored for environmental safety management. Microscopic discrimination of dinoflagellates is difficult to distinguish between toxic and non-toxic species due to their similar morphology. Meanwhile, an alternative quantitative PCR (qPCR) assay is sensitive, rapid, and cost-effective for harmful species monitoring. Herein, we developed a novel qPCR assay to detect the STXs biosynthesis gene sxtB of Alexandrium catenella and A. pacificum, the leading cause of PSP outbreaks in Asian coasts and worldwide. The newly designed sxtB TaqMan probes target the species without any positive signal in other relative dinoflagellates. Deming regression analysis revealed that the sxtB copy number of A. catenella and A. pacificum was 3.6 and 4.1 copies per cell, respectively. During the blooming periods (April 13th-14th, 2020), only A. catenella cells were detected through the qPCR assay, ranging from 5.0 × 10 to 2.5 × 104 eq cells L-1. In addition, sxtB qPCR quantified more accurately compared to large subunit (LSU) rRNA targeting qPCR assay that overestimate cell density. Besides, the sensitivity of sxtB was higher compared to the microscope when the species were rarely present (5.0 × 102 cells L-1). These suggest that the sxtB qPCR assay can be applied to toxic Alexandrium monitoring in the Korean coast, even in the early stage of bloomings.

Polyphyletic origin of saxitoxin biosynthesis genes in the marine dinoflagellate Alexandrium revealed by comparative transcriptomics.

The marine dinoflagellate Alexandrium is known to form harmful algal blooms, and at least 14 species within the genus can produce saxitoxins (STXs). STX biosynthesis genes (sxt) are individually revealed in toxic dinoflagellates; however, the evolutionary history remains controversial. Herein, we determined the transcriptome sequences of toxic Alexandrium (A. catenella and A. pacificum) and non-toxic Alexandrium (A. fraterculus and A. fragae) and characterized their sxt by focusing on evolutionary events and STX production. Comparative transcriptome analysis revealed higher homology of the sxt in toxic Alexandrium than in non-toxic species. Notably, non-toxic Alexandrium spp. were found to have lost two sxt core genes, namely sxtA4 and sxtG. Expression levels of 28 transcripts related to eight sxt core genes showed that sxtA, sxtG, and sxtI were relatively high (>1.5) in the toxic group compared to the non-toxic group. In contrast, the non-toxic group showed high expression levels in sxtU (1.9) and sxtD (1.7). Phylogenetic tree comparisons revealed distinct evolutionary patterns between 28S rDNA and sxtA, sxtB, sxtI, sxtD, and sxtU. However, similar topology was observed between 28S rDNA, sxtS, and sxtH/T. In the sxtB and sxtI phylogeny trees, toxic Alexandrium and cyanobacteria were clustered together, separating from non-toxic species. These suggest that Alexandrium may acquire sxt genes independently via horizontal gene transfer from toxic cyanobacteria and other multiple sources, demonstrating monocistronic transcripts of sxt in dinoflagellates.

Antagonistic interactions of the dinoflagellate Alexandrium catenella under simultaneous warming and acidification.

There is a concern that harmful algal bloom (HAB) species may increase under climate change. Yet, we lack understanding of how ecological interactions will be affected under ocean warming and acidification (OWA) conditions. We tested the antagonistic effects of three strains of the dinoflagellate HAB species Alexandrium catenella on three target species (the chlorophyte Tetraselmis sp., the cryptomonad Rhodomonas salina, and the diatom Thalassiosira weissflogii) at various biomass ratios between species, at ambient (16 °C and 400 µatm CO2) and OWA (20 °C and 2000 µatm CO2) conditions. In these experiments the Alexandrium strains had been raised under OWA conditions for ∼100 generations. All three non-HAB species increased their growth rate under OWA relative to ambient conditions. Growth rate inhibition was evident for R. salina and Tetraselmis sp. under OWA conditions, but not under ambient conditions. These negative effects were exacerbated at higher concentrations of Alexandrium relative to non-HAB species. By contrast, T. weissflogii showed positive growth in the presence of two strains of Alexandrium under ambient conditions, whereas growth was unaffected under OWA. Contrary to our expectations, A. catenella had a slight negative response in the presence of the diatom. These results demonstrate that Alexandrium exerts higher antagonistic effects under OWA compared to ambient conditions, and these effects are species-specific and density dependent. These negative effects may shift phytoplankton community composition under OWA conditions.

Allelochemicals of Alexandrium tamarense and its algicidal mechanism for Prorocentrum donghaiense and Heterosigma akashiwo.

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.

An On-Farm Workflow for Predictive Management of Paralytic Shellfish Toxin-Producing Harmful Algal Blooms for the Aquaculture Industry.

Paralytic shellfish toxins (PSTs) produced by marine dinoflagellates significantly impact shellfish industries worldwide. Early detection on-farm and with minimal training would allow additional time for management decisions to minimize economic losses. Here, we describe and test a standardized workflow based on the detection of sxtA4, an initial gene in the biosynthesis of PSTs. The workflow is simple and inexpensive and does not require a specialized laboratory. It consists of (1) water collection and filtration using a custom gravity sampler, (2) buffer selection for sample preservation and cell lysis for DNA, and (3) an assay based on a region of sxtA, DinoDtec lyophilized quantitative polymerase chain reaction (qPCR) assay. Water samples spiked with Alexandrium catenella showed a cell recovery of >90% when compared to light microscopy counts. The performance of the lysis method (90.3% efficient), Longmire's buffer, and the DinoDtec qPCR assay (tested across a range of Alexandrium species (90.7-106.9% efficiency; r2 > 0.99)) was found to be specific, sensitive, and efficient. We tested the application of this workflow weekly from May 2016 to 30th October 2017 to compare the relationship between sxtA4 copies L-1 in seawater and PSTs in mussel tissue (Mytilus galloprovincialis) on-farm and spatially (across multiple sites), effectively demonstrating an ∼2 week early warning of two A. catenella HABs (r = 0.95). Our tool provides an early, accurate, and efficient method for the identification of PST risk in shellfish aquaculture.