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• 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]

• 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]

• 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]

• The abundance of A. catenella cysts in sediments from Patagonia is normally low. • The low abundance of A. catenella cysts contrasts with the high intensity of PSP events. • The sediment textures are not good predictors of the abundance of A. catenella resting cysts. • The A. catenella cysts abundance matches with the northward geographical expansion. The frequency of harmful algal blooms (HABs) has increased over the last two decades, a phenomenon enhanced by global climate change. However, the effects of climate change will not be distributed equally, and Chile has emerged as one important, vulnerable area. The Chilean Patagonian region (41‒56°S) hosts two marine ecoregions that support robust blue economies via wild fisheries, aquaculture, and tourism. However, the harmful algal bloom-forming dinoflagellate Alexandrium catenella , a causative agent of paralytic shellfish poisoning outbreaks, threatens the viability of blue industries in this region and others worldwide. Despite the proliferation of A. catenella blooms over the last few decades, the role of sedimentary resting cysts in the recurrence of harmful algal blooms and the species' northward expansion across Chilean Patagonia is not well understood. As a resting cyst-producing species, the sediment-cyst dynamics of A. catenella likely contribute to the geographical expansion and bloom recurrence of this species. For this purpose, we analyzed a decade of A. catenella surface sediment cyst records across the two ecoregions of the Chilean Patagonian System that were further stratified into five subregions based on water temperature, salinity, dissolved oxygen, and nutrient characteristics. We also analyzed spatio-temporal cyst dynamics in a pre-, during-, and post-bloom scenario of the Chiloense ecoregion (more northern) of the Magellanic province. Our results indicated highly variable A. catenella resting cyst abundances, with a maximum of 221 cysts cm−3 recorded in 2002 after an intense bloom. Generalized linear mixed models and linear mixed models found that sampling season, subregion, and Total Organic Matter (%) explained resting cyst presence and density. The results also demonstrated the presence of A. catenella cysts in northern subregions, evidencing the northward geographical expansion observed during the last few decades. The risks of A. catenella bloom recurrence from small, patchy resting cyst distributions across broad geographical areas and under changing environmental conditions are discussed. [ABSTRACT FROM AUTHOR]

• Three distinct allelopathic HABs were paired with 15N–labeled Akashiwo sanguinea , allowing the release and transfer to N to be experimentally traced. • Allelopathic inhibition of Akashiwo was accompanied by increases in cell densities, growth rates, and the δ15N content of all HAB species. • HAB species obtained 60–100% of their cellular N from lysed Akashiwo cells. • Allelopathy may induce a positive feedback loop, whereby competitors are lysed, N is regenerated, and HABs are intensified. Allelopathy is a biological mechanism that can promote harmful algal blooms (HAB) via the inhibition of sympatric phytoplankton. While nutrient loading can also promote HABs, the ability of allelopathy to stimulate HABs via the regeneration of nutrients has yet to be explored. To examine the impacts of allelopathically liberated N on HAB species, a series of experiments were performed using multiple allelopathic HAB species including the dinoflagellates Alexandrium catenella and Margalefidinium polykrikoides , and the pelagophyte, Aureoumbra lagunensis. These HAB species were paired with the cosmopolitan dinoflagellate, Akashiwo sanguinea , that was labeled with 15NO 3 - or 15NH 4 +, allowing the release and transfer of N to be traced as a time course during allelopathic interactions. During all experiments, the allelopathic inhibition of Akashiwo was accompanied by increases in cell densities, growth rates, and the δ15N content of the HAB species, evidencing the transfer of N liberated from Akashiwo. The cellular transfer of 15N and release of dissolved N was higher when Akashiwo was grown with 15NO 3 - compared to 15NH 4 + suggesting a differential subcellular-compartmentalization of N sources. Regardless of the type of N, HAB species obtained 60 – 100% of their cellular N from lysed Akashiwo cells and there was an enrichment of the δ15N content of the dissolved NH 4 + pool post-lysis of Akashiwo. Collectively, the results demonstrate that beyond facilitating species succession, allelopathy can supply HABs with N and, therefore, is likely important for promoting and sustaining HABs. Given that allelopathy is known to be a dose-dependent process, allelopathy may induce a positive feedback loop, whereby competitors are lysed, N is liberated, HABs are intensified and, in turn, become more strongly allelopathic. [ABSTRACT FROM AUTHOR]

Highlights • Novel use of the Microelectrode Ion Flux Estimation (MIFE) technique to assess gill cell ion fluxes against ficotoxins. • Development of an effective Basic Solution Medium (BSM) to test the RTgill-W1 ion fluxes. • Ichthyotoxic metabolites from A. catenella result in net K+ efflux from gill cells. • Paralytic Shellfish Toxins (PST) alone do not alter key gill cell-membrane ion fluxes. Abstract Harmful algal blooms (HAB) are responsible for massive mortalities of wild and aquacultured fish due to noticeable gill damage, but the precise fish-killing mechanisms remain poorly understood. A non-invasive microelectrode ion flux estimation (MIFE) technique was successfully applied to assess changes in membrane-transport processes in a model fish gill cell line exposed to harmful microplankton. Net Ca2+, H+, K+ ion fluxes in the rainbow trout cell line RTgill-W1 were monitored before and after addition of lysed cells of this Paralytic Shellfish Toxins (PST) producer along with purified endocellular dinoflagellate PST. It was demonstrated that PST alone do not play a role in fish gill damage during A. catenella outbreaks as previously thought, but that other ichthyotoxic metabolites from lysed algal cells (i.e. lipid peroxidation products or other unknown metabolites) result in net K+ efflux from fish gill cells and thereby gill cell death. [ABSTRACT FROM AUTHOR]

Despite the long history of paralytic shellfish poisoning (PSP) events in Alaska, little is known about the seasonal distribution and abundance of the causative organism, Alexandrium , or the environmental factors that govern toxic bloom development. To address this issue, a five year study (2012–2017) was undertaken in Kachemak Bay and lower Cook Inlet Alaska to determine how the occurrence of Alexandrium catenella , the dominant PSP-causing Alexandrium species, was influenced by temperature, salinity, nutrient concentrations, and other environmental factors. Cell concentrations from 572 surface water samples were estimated using quantitative PCR. Monthly sampling revealed a seasonal pattern of A. catenella bloom development that was positively correlated with water temperature. Prevailing salinity conditions did not significantly affect abundance, nor was nutrient limitation a direct factor. Elevated cell concentrations were detected in 35 samples from Kachemak Bay (100-3050 cell eq. L −1 ) while a maximum abundance of 67 cell eq. L −1 was detected in samples from lower Cook Inlet sites. Monitoring data showed average water temperatures in Kachemak Bay increased by ∼2 °C over the course of the study and were accompanied by an increase in Alexandrium abundance. Based on these findings, 7–8 °C appears to represent a temperature threshold for significant bloom development in Kachemak Bay, with the greatest risk of shellfish toxicity occurring when temperatures exceed 10–12 °C. The role of temperature is further supported by time series data from the Alaska Coastal Current (station GAK1), which showed that summertime shellfish toxicity events in Kachemak Bay generally followed periods of anomalously high winter water temperatures. These data indicate monitoring changes in water temperatures may be used as an early warning signal for subsequent development of shellfish toxicity in Kachemak Bay. [ABSTRACT FROM AUTHOR]

Harmful algal blooms (HABs) have affected salmon farms in Chile since the early 1970′s, causing massive losses in fish. Two large HABs occurred in 2002 and 2009, during which Alexandrium catenella blooms killed tons of salmon over an extended geographic area in southern Chile. At the beginning of 2016, high and persistent densities of Pseudochattonella cf. verruculosa and A . catenella were detected in the estuarine and marine ecosystems of southern Chile. Mortality for this latter event reached 27 million salmon and trout (i.e. 39,000 tons). Unfortunately, the threshold concentrations of algae that could be harmful to the health of farmed salmon in southern Chile have not yet been quantified. Here, to protect fish farms from HABs, critical concentration levels, i.e. thresholds at which the behavior of farmed Salmo salar is affected by harmful algae were quantified using generalized linear mixed models (GLMM). An extensive database from southern Chile covering the period from 1989 to 2016 was analyzed. The database included salmon behavior, cell abundance of microalgae and oceanographic factors. For both species analyzed, the higher the cell abundance, the greater the probability of detecting anomalous behavior. A threshold of 397 cells/mL was estimated for A. catenella , although it can increase up to ca. >975 cells/mL at a Secchi depth >6 m and up to 874 cells/mL during flood tide. A threshold value <1 cell/mL for Pseudochattonella cf. verruculosa was found to be associated with anomalous salmon behavior, which significantly increased at a water temperature of 11 °C. Evidence for a relationship between fish behavior and mortality is provided. [ABSTRACT FROM AUTHOR]

The dinoflagellate genus Alexandrium Halim has frequently been associated with harmful algal blooms. Although a number of species from this genus are known to produce paralytic shellfish toxins (PST) and/or cyclic imines (CI), studies on comprehensive toxin profiling using techniques capable of detecting the full range of PST and CI analogues are limited. Isolates of Alexandrium spp. from Atlantic Canada were analyzed by targeted and untargeted liquid chromatography-tandem mass spectrometry (LC–MS). Results showed a number of distinct profiles and wide ranging cell quotas of PST and spirolides (SPX) in both A. catenella (Whedon & Kofoid) Balech and A. ostenfedii (Paulsen) Balech & Tangen. The concentration of PST in A. catenella ranged from 0.0029 to 54 fmol cell −1 with the major components being C2 and GTX4. In addition, putative PST metabolites were confirmed for the first time in A. catenella by high resolution MS/MS. By comparison, A. ostenfeldii isolates showed much lower concentrations of PST (

• Cellular toxin content of the dinoflagellate Alexandrium catenella during a laboratory-simulated bloom increased with time and plateaued after the stationary phase. • There was a significantly positive relationship between growth rate and constitutive toxin production, predominantly in the exponential phase. • Grazer-induced toxin production was evident throughout the bloom and greatest in the exponential phase. • Cell growth rate was negatively related to grazer-induced toxin production, consistent with a defense-growth trade-off. • The dynamics of cell growth vs toxin production are fundamentally different for constitutive and inducible defense. Although the typical framework for studies and models of bloom dynamics in toxigenic phytoplankton is predominantly based on abiotic determinants, there is mounting evidence of grazer control of toxin production. We tested for the effect of grazer control of toxin production and cell growth rate during a laboratory-simulated bloom of the dinoflagellate Alexandrium catenella. We measured cellular toxin content and net growth rate when cells were exposed to copepod grazers (direct exposure), copepod cues (indirect exposure), and no copepods (control) throughout the exponential, stationary, and declining phases of the bloom. During the simulated bloom, cellular toxin content plateaued after the stationary phase and there was a significantly positive relationship between growth rate and toxin production, predominantly in the exponential phase. Grazer-induced toxin production was evident throughout the bloom, but highest during the exponential phase. Induction was greater when cells were directly exposed to grazers rather than their cues alone. In the presence of grazers toxin production and cell growth rate were negatively related, indicating a defense-growth trade-off. Further, a fitness reduction associated with toxin production was more evident in the presence than the absence of grazers. Consequently, the relationship between toxin production and cell growth is fundamentally different between constitutive and inducible defense. This suggests that understanding and predicting bloom dynamics requires considering both constitutive and grazer-induced toxin production. [ABSTRACT FROM AUTHOR]

Paralytic shellfish poisoning (PSP) caused the deaths of four people in coastal area of Korea, mainly Jinhae-Masan Bay and adjacent areas, in April 1986 and in 1996. The PSP outbreaks were caused by the consumption of mussels, Mytilus edulis . The organism that caused PSP was identified, from morphological data only, as Alexandrium tamarense which is recently renamed as A. catenella , however recent studies have shown that the morphological diagnostic characteristics used to identify Alexandrium species have uncertainties and molecular tools and other criteria should be considered as well. The organism that caused past PSP outbreaks and incidents in Korea therefore need to be carefully reconsidered. The aim of this study was to re-evaluate the species really responsible for past outbreaks of PSP in Jinhae-Masan Bay, Korea. The temporal production and fluxes of the resting cysts of Alexandrium species were investigated for one year (from March 2011 to February 2012) using a sediment trap, and the morphology and phylogeny of vegetative cells germinated from the resting cysts were analysed. The production of Alexandrium species peaked in August and November, when temporal discrepancies were found in the water temperature (22.4 and 22.7 °C in August, 19.1 and 19.6 °C in November) and salinity (29.5 and 26.1 psu in August, 30.5 and 31.8 psu in November). The morphological data revealed that Alexandrium species germinated from resting cysts collected in August have a ventral pore on the 1′ plate, whereas the 1′ plate in Alexandrium species germinated from resting cysts collected in November lacks a ventral pore. Molecular phylogenetic data for the vegetative cells from the germination experiments allowed the August and November peaks to be assigned to Alexandrium catenella (Group I) and A. pacificum (Group IV), respectively. This indicates that the production of resting cysts of A. catenella can be enhanced by relatively high water temperature. This result is not consistent with those of previous studies that A. catenella responsible for PSP outbreaks was found at relatively low water temperature. In addition, large subunit ribosomal sequences data revealed that A. pacificum isolates from Korea were closely related to those from Australia, Japan and New Zealand where the PSP toxicity of shellfish and blooms occurred in the 1990s, indicating that the introduction of toxic dinoflagellates were related to ballast water from bulk-cargo shipping. Based on these results, we concluded that past PSP outbreaks in Jinhae-Masan Bay of Korea could have been caused by A. pacificum rather than by A. catenella . [ABSTRACT FROM AUTHOR]

Red tides dominated by Cochlodinium polykrikoides often lead to great economic losses and some methods of controlling these red tides have been developed. However, due to possible adverse effects and the short persistence of their control actions, safer and more effective sustainable methods should be developed. The non-toxic dinoflagellate Alexandrium pohangense is known to grow well mixotrophically feeding on C. polykrikoides, and populations are also maintained by photosynthesis. Thus, compared with other methods, the use of mass-cultured A. pohangense is safer and the effects can be maintained in the long term. To develop an effective method, the concentrations of A. pohangense cells and culture filtrate resulting in the death of C. polykrikoides cells were determined by adding the cells or filtrates to cultured and natural populations of C. polykrikoides . Cultures containing 800 A. pohangense cells ml −1 eliminated almost all cultured C. polykrikoides cells at a concentration of 1000 cells ml −1 within 24 h. Furthermore, the addition of A. pohangense cultures at a concentration of 800 cells ml −1 to C. polykrikoides populations from a red-tide patch resulted in the death of most C. polykrikoides cells (99.8%) within 24 h. This addition of A. pohangense cells also lowered the abundances of total phototrophic dinoflagellates excluding C. polykrikoides , but did not lower the abundance of total diatoms. Filtrate from 800 cells ml −1 A. pohangense cultures reduced the population of cultured C. polykrikoides by 80% within 48 h. This suggests that A. pohangense cells eliminate C. polykrikoides by feeding and releasing extracellular compounds. Over time, A. pohangense concentrations gradually increased when incubated with C. polykrikoides . Thus, an increase in the concentration of A. pohangense by feeding may lead to A. pohangense cells eliminating more C. polykrikoides cells in larger volumes. Based on the results of this study, a 1 m 3 stock culture of A. pohangense at 4000 cells ml −1 is calculated to remove all C. polykrikoides cells in ca. 200 m 3 within 6 days. Furthermore, maintenance of A. pohangense populations through photosynthesis prepared A. pohangense to eliminate C. polykrikoides cells in future red-tide patches. Moreover, incubation of A. pohangense at 2000 cells ml −1 with juvenile olive flounder Paralichthys olivaceus for 3 days did not result in the death of fish. Therefore, the method developed in this study is a safe and effective way of controlling C. polykrikoides populations and can be easily applied to aqua-tanks on land. [ABSTRACT FROM AUTHOR]

• Nitrogen-enriched environments can increase the concentration of paralytic shellfish poisoning (PST) in dinoflagellates such as Alexandrium catenella. • The study characterizes the proteome of A. catenella and expression of PST in culture media with different NaNO 3 concentrations using mass spectrometry, bioinformatics, and toxicology techniques. • In this model, high nitrogen environment increases diverse cellular processes in A. catenella. However, the toxin-producing enzymes and toxin production are down-regulated. Dinoflagellates of the genus Alexandrium cause Harmful Algal Blooms (HABs) in coastal waters worldwide, damaging marine environments, aquaculture, and human health. They synthesize potent neurotoxic alkaloids known as PSTs (i.e., Paralytic Shellfish Toxins), the etiological agents of PSP (i.e., Paralytic Shellfish Poisoning). In recent decades, the eutrophication of coastal waters with inorganic nitrogen (e.g., nitrate, nitrite, and ammonia) has increased the frequency and scale of HABs. PSTs concentrations within Alexandrium cells can increase by up to 76% after a nitrogen enrichment event; however, the mechanisms that underlie their biosynthesis in dinoflagellates remains unclear. This study combines mass spectrometry, bioinformatics, and toxicology and investigates the expression profiles of PSTs in Alexandrium catenella grown in 0.4, 0.9 and 1.3 mM NaNO 3. Pathway analysis of protein expression revealed that tRNA amino acylation, glycolysis, TCA cycle and pigment biosynthesis were upregulated in 0.4 mM and downregulated in 1.3 mM NaNO 3 compared to those grown in 0.9 mM NaNO 3. Conversely, ATP synthesis, photosynthesis and arginine biosynthesis were downregulated in 0.4 mM and upregulated in 1.3 mM NaNO 3. Additionally, the expression of proteins involved in PST biosynthesis (sxt A, sxt G, sxt V, sxt W and sxt Z) and overall PST production like STX, NEO, C1, C2, GTX1-6 and dcGTX2 was higher at lower nitrate concentrations. Therefore, increased nitrogen concentrations increase protein synthesis, photosynthesis, and energy metabolism and decrease enzyme expression in PST biosynthesis and production. This research provides new clues about how the changes in the nitrate concentration can modulate different metabolic pathways and the expression of PST biosynthesis in toxigenic dinoflagellates. [ABSTRACT FROM AUTHOR]

The detection of sparse Alexandrium catenella -resting cysts in sediments of southern Chilean fjords has cast doubts on their importance in the recurrence of massive toxic dinoflagellate blooms in the region. The role of resting cysts and the existence of different regional Chilean populations was studied by culturing and genetic approaches to define: (1) cyst production; (2) dormancy period; (3) excystment success; (4) offspring viability and (5) strain mating compatibility. This study newly revealed a short cyst dormancy (minimum 69 days), the role of key abiotic factors (in decreasing order salinity, irradiance, temperature and nutrients) controlling cyst germination (max. 60%) and germling growth rates (up to 0.36–0.52 div. day −1 ). Amplified fragment length polymorphism (AFLP) characterization showed significant differences in genetic distances (GD) among A. catenella populations that were primarily determined by the geographical origin of isolates and most likely driven by oceanographic dispersal barriers. A complex heterothallic mating system pointed to variable reproductive compatibility (RCs) among Chilean strains that was high among northern (Los Lagos/North Aysén) and southern populations (Magallanes), but limited among the genetically differentiated central (South Aysén) populations. Field cyst surveys after a massive 2009 bloom event revealed the existence of exceptional high cyst densities in particular areas of the fjords (max. 14.627 cysts cm −3 ), which contrast with low cyst concentrations (<221.3 cysts cm −3 ) detected by previous oceanographic campaigns. In conclusion, the present study suggests that A. catenella resting cysts play a more important role in the success of this species in Chilean fjords than previously thought. Results from in vitro experiments suggest that pelagic–benthic processes can maintain year-round low vegetative cell concentrations in the water column, but also can explain the detection of high cysts aggregations after the 2009-bloom event. Regional drivers that lead to massive outbreaks, however, are still unknown but potential scenarios are discussed. [ABSTRACT FROM AUTHOR]

• Combination of state-of-the-art microbial ecology methods including Flow-CAM imaging, high-throughput sequencing, and quantitative PCR to study Alexandrium catenella. • Documentation of the expansion of a harmful algal species known to produce a neurotoxin, A. catenella , into the warming Arctic. • Examination of the role of contrasting physical processes on the distribution of A. catenella. • Discussion of the potential health and economic consequences for the Arctic of the expansion of the A. catenella habitat. The harmful algal genus Alexandrium has characteristically been found in temperate and subtropical regions; however recent evidence suggests global warming may be expanding its range into high latitude waters. Alexandrium cysts have previously been documented in the Chukchi Sea and we hypothesize that Alexandrium may be expanding further into the Arctic due to distribution by the Beaufort shelfbreak jet. Here we document the presence of Alexandrium catenella along the Alaskan Beaufort Sea shelf, marking an expansion of its known range. The observations of A. catenella were made using three different methods: FlowCAM imaging, 18S eukaryotic sequencing, and real-time quantitative PCR. Four occupations of a shelf/slope transect spanned the evolution of a strong wind-driven upwelling event over a 5-day period. A nearby mooring provided the physical context for the event, revealing that enhanced easterly winds reversed the Beaufort shelfbreak jet to the west and induced upwelling of colder, denser water onto the outer shelf. A. catenella sequences dominated the surface phytoplankton community at the onset of the upwelling event. This signal vanished during and after the event, likely due to a combination of alongstream advection, cross-stream advection, and wind mixing. These results suggest contrasting physical processes that are both subject to global warming amplification, delivery of warm waters via the Beaufort shelfbreak jet and upwelling, may control the proliferation of this potential harmful alga into the Arctic. [ABSTRACT FROM AUTHOR]

Outbreaks of the dinoflagellate Alexandrium catenella have caused significant economic losses to the salmon industry in the south of Chile. However, the precise ichthyotoxic mechanism by this paralytic shellfish toxin (PST) – producing dinoflagellate remains poorly understood. The rainbow trout cell line RTgill-W1 assay was used to investigate fish gill damage by multiple A. catenella strains under different environmental conditions, and potency of whole cells, lysed cells and culture medium were compared. Cytotoxic potency was highly variable among strains and strongly correlated to dinoflagellate cell-abundance. Lysed cells produced more gill damage than intact cells and supernatant, reducing gill cell viability down to 20% of controls at 4000 cells mL −1 . However, gill cells exposed to pure PST fractions (C1&C2, STX, GTX 1&4) exhibited only very limited loss of viability (<30%), even at cell concentrations equivalent to exceeding those detected in southern Chilean fjords. A. catenella lytic compounds rapidly (1–4 days) degraded in the light, but were pH (7–9) and temperature stable (17–85 °C), thus arguing against the involvement of proteinaceous compounds. Superoxide production by Chilean dinoflagellate strains was as high as by the raphidophyte Chattonella marina (max. 8.67 ± 0.14 pmol O 2 − cell −1 h −1 ), variable among strains and enhanced by cell disruption and environmental stress (low nutrients, low salinity, low or high pH). Fatty acid profiles included high concentrations of the long-chain (≥C 20 ) polyunsaturated fatty acids (PUFA) – docosahexaenoic acid (DHA, 22:6ω3; 16–20% of total fatty acids) in addition to several other PUFA, which were highly cytotoxic against gill cells as a purified fraction (LC 50 1.30 μg mL −1 ). In addition, potency was enhanced 9-fold in combination with superoxide anion (LC 50 0.15 μg mL −1 ). In conclusion, the present study demonstrates that fish gill damage during A. catenella fish-kill events in Chilean fjords cannot be explained by PST toxins, but can to a large extent be accounted for by the synergistic interaction between ROS (superoxide anion) and DHA, and potentially other PUFA, notably under conditions that promote cell lysis. [ABSTRACT FROM AUTHOR]

A differential screening study using high-resolution (HR)-hydrophilic interaction chromatography (HILIC)-electrospray ionization (ESI)–quadrupole time-of-flight mass spectrometry (Q-TOF MS) was conducted to identify saxitoxin (STX) analogues in the marine dinoflagellate toxic sub-clone Alexandrium tamarense Axat-2 and the non-toxic sub-clone UAT-014-009 derived from the same Japanese isolate. One unknown compound was identified only in the toxic sub-clone and was found to have the molecular formula C 9 H 16 N 6 O 2 . This structure differed from that of decarbamoyl STX (dcSTX; C 9 H 16 N 6 O 3 ) by the loss of a single oxygen. A 12-deoxy-dcSTX standard (a mixture of 12α- and β-deoxy-dcSTX) was chemically prepared from dcSTX by reduction with sodium borohydride. The unknown compound in the toxic strain of A. tamarense was identified as 12β-deoxy-dcSTX by comparison of its HR-HILIC-LC–MS retention time and HR–MS/MS spectrum with those of the chemically prepared standard, and the identification was confirmed by high-sensitivity HPLC analysis with post-column fluorescent derivatization. Moreover, two Japanese isolates of A. catenella showing toxin profiles different from that of A. tamarense were also found to contain 12β-deoxy-dcSTX. Previously, 12β-deoxy-dcSTX was isolated from the freshwater cyanobacterium Lyngbya wollei , which produces a unique set of STX analogues. This study is the first evidence of the presence of 12β-deoxy-dcSTX in marine dinoflagellates. [ABSTRACT FROM AUTHOR]

Blooms of toxic dinoflagellates can co-occur with mass mortality events associated with herpesvirus OsHV-1 μVar infection that have been decimating Pacific oyster Crassostrea gigas spat and juveniles every summer since 2008 in France. This study investigated the possible effect of a harmful dinoflagellate, Alexandrium catenella , a producer of Paralytic Shellfish Toxins (PSTs), upon the oyster spat–herpesvirus interaction. Oyster spat from a hatchery were challenged by cohabitation with oysters contaminated in the field with OsHV-1 μVar and possibly other pathogens. Simultaneously, the oysters were exposed to cultured A. catenella . Infection with OsHV-1 μVar and PST accumulation were measured after 4 days of experimental exposure. Exposure to Alexandrium catenella modified the host–pathogen interaction by reducing prevalence of OsHV-1 μVar infection. In addition, oysters challenged with OsHV-1 μVar and possibly other pathogens from the environment accumulated smaller amounts of PSTs than unchallenged oysters. Three possible mechanisms are suggested by these results: (i) possible direct interactions between A. catenella and herpesvirus (or associated pathogens) could reduce viral transmission and algal availability for oyster consumption; (ii) oyster feeding behavior or digestive functions may have been altered, thus decreasing both uptake of viral particles and consumption or digestion of toxic algae and consequent toxin accumulation; (iii) immuno-activation by A. catenella could enhance defense efficiency against OsHV-1 μVar infection. These findings suggest further research on relationships between OsHV-1 μVar and toxic dinoflagellates and their combined effects upon disease transmission and proliferation processes, as well as on oyster physiological and immunological involvement in this complex, tripartite interaction. [ABSTRACT FROM AUTHOR]

Cell abundances and distributions of Alexandrium catenella resting cysts in recent sediments were studied along time at two locations in the Chilean Inland Sea exposed to different oceanographic conditions: Low Bay, which is much more open to the ocean than the more interior and protected Ovalada Island. The bloom began in interior areas but maximum cyst concentrations were recorded in locations more open to the ocean, at the end of the Moraleda channel. Our results showed a time lapse of around 3 months from the bloom peak (planktonic population) until the number of resting cysts in the sediments reached a maximum. Three months later, less than 10% of the A. catenella cysts remained in the sediments. Maximum cyst numbers in the water column occurred one month after the planktonic peak, when no cells were present. The dinoflagellate assemblage at both study sites was dominated by heterotrophic cysts, except during the A. catenella bloom. CCA analyses of species composition and environmental factors indicated that the frequency of A. catenella blooms was associated with low temperatures, but not with salinity, chlorophyll a concentration, and predator presence (measured as clam biomass). However, resting cyst distribution was only related to cell abundance and location. The occurrence of A. catenella cysts was also associated with that of cysts from the toxic species Protoceratium reticulatum . By shedding light on the ecological requirements of A. catenella blooms, our observations support the relevance of encystment as a mechanism of bloom termination and show a very fast depletion of cysts from the sediments (<3 months), which suggest a small role for resting cyst deposits in the recurrence of A. catenella blooms in this area. [ABSTRACT FROM AUTHOR]

Specimens of Munida gregaria were collected within and in the vicinity of a bloom of the toxic dinoflagellate Alexandrium catenella in Queen Charlotte Sound, New Zealand. The crustacean contained paralytic shellfish toxin (PST) with an analogue profile dominated by N -sulfocarbamoyl analogues (C1,2 and GTX5) and carbamate gonyautoxins (GTX1,4), similar to that of the dinoflagellate. A feeding experiment showed that M. gregaria is capable of actively grazing on A. catenella and it may play a role in controlling population growth of the dinoflagellate. This is the first account of the accumulation of PST by M. gregaria . When it is periodically abundant, M. gregaria is an important food item for fish, birds and other marine fauna and they are a vector by which PST may be transferred to higher trophic levels. [ABSTRACT FROM AUTHOR]

Molecular mechanisms regulating responses to phosphate (P) limitation in dinoflagellates are poorly understood. Here we investigated gene differential expression Alexandrium catenella strain ATHK using suppression subtractive hybridization (SSH) and reverse-transcription quantitative PCR (qRT-PCR) techniques. SSH revealed a score of genes responded to P-limitation, and comparison with EST sequencing results indicated superior efficiency of SSH in identifying differentially expressed genes. Among the P-limitation responsive genes were the up-regulated stress-related genes, the down-regulated phosphorus-transferring genes, and a number of other genes previously undocumented for dinoflagellates. Most notably, the previously undocumented cell cycle inhibitory regulator fizzy/cell division cycle 20-related protein gene was up-regulated while putatively cell cycle promoting genes (e.g. calcium-dependent protein kinase) down-regulated, consistent with the decrease in growth rate and cell cycle arrest in G1 phase under P-limitation observed microscopically and flow cytometrically respectively. Besides, both SSH and qRT-PCR showed that saxitoxin related genes, menaquinone biosynthesis methyltransferase, an unknown gene and some other genes were also significantly induced by P-limitation. Our results demonstrate the utility of SSH in gene regulation research and provide a set of promising genes for further studies to understand the regulation of cell division (and hence population growth) and toxin production by P-limitation in A. catenella and likely other dinoflagellates. [ABSTRACT FROM AUTHOR]

Diatom blooms in Thau lagoon are always related to rain events leading to inputs of inorganic nutrients such as phosphate, ammonium and nitrate through the watershed with time lags of about 1 week. In contrast, blooms of Alexandrium catenella/tamarense can occur following periods of 3 weeks without precipitation and no significant input of conventional nutrients such as nitrate and phosphate. Field results also indicate a significant drop (from 22-25 to 15-16 µM over 3 days) in dissolved organic nitrogen (DON) at the bloom peak, as well as a significant inverse relationship between A. catenella/tamarense cell density and DON concentrations that is not apparent for diatom blooms. Such dinoflagellate blooms are also associated with elevated (6-9 µM) ammonium concentrations, a curious feature also observed by other investigators, possibly the results of ammonium excretion by this organism during urea or other organic nitrogen assimilation. The potential use of DON by this organism represents short cuts in the nitrogen cycle between plants and nutrients and requires a new model for phytoplankton growth that is different from the classical diatom bloom model. In contrast to such diatom blooms that are due to conventional (nitrate, phosphate) nutrient pulses, Alexandrium catenella/tamarense blooms on the monthly time scale are due to organic nutrient enrichment, a feature that allows net growth rates of about 1.3 d-1, a value higher than that generally attributed to such organisms. [ABSTRACT FROM AUTHOR]

Highlights: [•] Conditioning of a clonal culture of Alexandrium catenella to a factorial combination of environmental change variables; CO2, temperature, phosphorus. [•] Measurements of growth and cellular toxins after eight months of conditioning. [•] Increased CO2 led to enhanced cellular toxicity. [•] Combined high CO2, low temperature and phosphate-limitation were the ‘most toxic’. [Copyright &y& Elsevier]

Highlights: [•] We studied growth of 4 mixotrophic dinoflagellate species with their prey organisms. [•] Growth of Karenia mikimotoi was notably promoted after addition of prey organisms. [•] Growth of two Prorocentrum species was only slightly improved with prey organisms. [•] Presence of prey organisms has no effects on the growth of Alexandrium catenella. [•] Mixotrophy may play different roles in growth and bloom of different dinoflagellates. [Copyright &y& Elsevier]

Highlights: [•] Wind patterns override factors controlling Alexandrium catenella blooms in Thau lagoon. [•] Surface water temperature impacts the seasonal development of blooms (spring and fall). [•] Southeasterly winds disperse the blooms, northwesterly winds terminate the blooms. [•] Maximal cell densities of Alexandrium catenella are related to calm period durations. [Copyright &y& Elsevier]

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

Abstract: In 2010 the Cawthron Institute adopted AOAC official method 2005.06 (Lawrence method) for regulatory testing of paralytic shellfish toxins. This included adapting the method to a UPLC format and developing a rapid periodate screen to eliminate the vast majority of samples with no PSTs present. The method gained New Zealand regulatory approval and has since been used to test >2000 samples. Soon after implementation a major HAB of the toxic dinoflagellate Alexandrium catenella occurred in a prime shellfish growing area of New Zealand. This event was the most serious to date in this country with extremely high cell concentrations observed in some locations (>4×106 cellsL−1). Toxin levels observed in Greenshell™ mussels (Perna canaliculus) and Flat oysters (Ostrea chilensis) exceeded the regulatory level of 0.8mg/kg shellfish meat as saxitoxin equivalents. Closures of commercial shellfish harvesting areas were enforced for a period of up to three months as toxin levels remained above the regulatory level for an extended period, even after the bloom had crashed. Analysis of several hundred positive shellfish samples during this event allowed us to better understand the technical performance of the method during a bloom event. The periodate screen substantially overestimated the true PST level in the samples because several PSTs gave co-eluting oxidation products, and it was assumed that the entire peak was due to the presence of the more toxic congener. The ratio between the screen and confirmation test results remained relatively constant throughout the bloom events. This information supports an amendment to the overly conservative regulatory control scheme employed in New Zealand for PST testing. Despite overestimation, the periodate screen has proved highly useful as it allows a quick determination of PST-free samples and provides a high level of security against harvesting contaminated products. [Copyright &y& Elsevier]

Abstract: The toxic dinoflagellate Alexandrium catenella has been detected in the southern Chile since 1972, causing severe negative impacts on public health and aquaculture activities. Several environmental factors have been determined to affect growth and toxin production in Alexandrium strains. The aim of this study was to determine the effect of four combined conditions of two temperatures (10 and 15°C) and two salinities (15 and 35 psu) on the growth and the Paralytic Shellfish Poisoning (PSP) toxin content and composition in four Chilean strains of A. catenella (PFB41, PFB42, PFB37 and PFB38), isolated during a summer outbreak occurred in southern Chile in 2009. The growth curves showed a higher effect of the salinity in strains PFB41 and PFB42 than in strains PFB37 and PFB38. The values of growth rates and maximum cell densities ranged from 0.25 to 0.73 div day−1 and 1.1×104 to 5.2×104 cells mL−1, respectively. All of the strains showed the highest values for both growth parameters at 15°C and 35 psu. In general, growth parameters were higher at 35 psu independently of the temperature. On the other hand, the total PSP toxin content ranged widely from 3.99 to 239fmol cell−1. The highest values of PSP toxin content were attained at 10°C and 35 psu for all of the strains, at both stages of growth. All of the strains displayed different toxin compositions, with neoSTX, GTX4-1, GTX3-2 and GTX5 being the main toxins detected. The results showed significant differences in the absolute values of growth and toxin production parameters among the strains grown under the same culture conditions, and for each strain grown under different combined conditions of temperature and salinity. These findings demonstrate that abiotic factors can differentially affect the population dynamics of the A. catenella toxic genotypes, thus making it extremely difficult to predict the ecological behavior of this species in the field in terms of the intensity of a potential outbreak. [Copyright &y& Elsevier]

Abstract: Juvenile Northern scallops Argopecten purpuratus were exposed to cultures of the paralytic shellfish toxin (PST) producing dinoflagellate, Alexandrium catenella, or a non-toxic microalga as a control, T-iso. After 3 and 6 days of exposure to either A. catenella or T-iso, scallops were stimulated to elicit an escape response by exposing them to the predatory sea star Meyenaster gelatinosus. We monitored the escape response of the scallops in terms of reaction time after first contact with the sea star, number of claps (burst of rapid valve closures) until exhaustion, clapping time, clapping rate, the time scallops spent closed when exhausted, and recovery from the initial number of claps, clapping time and clapping rate. Additionally, histopathological and stress responses (through heat-shock protein [hsp70] induction), as well as accumulation of Paralytic Shellfish Poisoning (PSP) toxins, were monitored on scallops after 3 and 6 days of exposure to A. catenella. After 6 days of exposure, scallops exposed to A. catenella accumulated PSTs and reacted more rapidly with a higher clapping rate, however the duration of their escape response was shorter than controls, when exposed to M. gelatinosus. Additionally, scallops exposed to A. catenella showed histopathological features, especially after 6 days of exposure, including increased melanization of the tissues and myopathy, with high levels of degeneration of the muscle fibers. A six-day exposure to A. catenella also caused an increase in prevalence of rickettsiales-like organisms within scallop tissues. This study suggests that PST accumulation can affect the interaction between the Northern scallop and both pathogens and predators, potentially increasing their susceptibility to either of them. [Copyright &y& Elsevier]

Abstract: The functional role of harmful substances (i.e. toxins) produced by marine planktonic algae is still, in many cases, unknown. This study describes a novel mechanism by which the phototrophic dinoflagellate Alexandrium pseudogonyaulax secretes a toxic mucus trap where prey items are caught and immobilized prior to ingestion. Prey cells remain entrapped and immobile in the mucus trap, but most stay intact, readily available as whole-cell prey. It is shown that food uptake by A. pseudogonyaulax increases its growth rate considerably even in nutrient-replete, high-light conditions. The increase in growth rate was more enhanced in light-limited treatments and A. pseudogonyaulax grew significantly faster when fed Heterocapsa rotundata, than when fed Teleaulax acuta under both light conditions. For comparison, strains of Alexandrium catenella and Alexandrium minutum were studied for their mixotrophic capabilities. None of these strains were mixotrophic under the conditions provided. In addition, the toxic effects on various protistan targets of these Alexandrium strains as well as Alexandrium tamarense and Alexandrium ostenfeldii were compared to that of A. pseudogonyaulax. A. tamarense and A. catenella did immobilize and lyse target cells through substances leaked directly into the water, differing from all the strains of A. pseudogonyaulax studied. Results show that the toxic effect of A. pseudogonyaulax is non-specific causing nearly 100% immobilization of a variety of protistan targets at relatively low cell concentrations (500cellsml−1 of donor cell). A critical donor cell density was not required as only one A. pseudogonyaulax cell was able to cause immobilization of target cells. For the first time, the connection between excreted toxins and phagotrophy is evident in an Alexandrium species and this particular strategy has the potential to severely impact competing phytoplankton communities. [Copyright &y& Elsevier]

Abstract: Morphological, toxicological and phylogenetic analyses, using the partial LSU gene and internal spacer (ITS) regions of the rDNA gene, were combined to evaluate the intraregional diversity of Alexandrium catenella occurring along the southern coast of Chile. Twenty-two strains isolated from different localities along the wide area of distribution of the species (from 42°S to 55°S) were examined by these three approaches. Morphologically, although the strains showed diagnostic characters according to the species definition, variations in these traits within and between strains were also observed. The absence of an apical or posterior attachment pore, for instance, was observed mainly in old isolates. Indirect connection between the apical and 1′ plates, traits normally seen in other species of the same genus, was also noted in some strains. However, the lack of a ventral pore on the 1′ plate was one of the most distinctive characteristics present in all the Chilean strains. Toxicologically, the Chilean strains were characterized by the dominance of N-sulfocarbamate (C1,2) and gonyautoxins (GTX1–4), but also by the scarcity or absence of saxitoxin. Considering the dominance of these toxins in each strain, at least two distinctive toxin patterns were distinguished. Through rDNA sequence analysis, the Chilean strains were segregated as part of Clade I (North American) of the Alexandrium tamarense species complex. Nevertheless, significant genetic diversity was also observed among the Chilean strains, especially using ITS sequences. Through these three approaches, Chilean strains of A. catenella showed significant intraregional variability, which is appropriate for a native species. However, the distribution of its genetic diversity seems to be inconsistent with the apparent northward expansion observed along the west coast of South America. [Copyright &y& Elsevier]

Abstract: The dinoflagellate Alexandrium catenella causes recurrent harmful algal blooms in southern Chile. This species belongs to the “Alexandrium tamarense/catenella/fundyense species complex” (the “tamarensis complex”), defined by morphological attributes. Ribosomal sequences serve to differentiate five evolutionary lineages (clades) in this species complex. These distinctions reflect the geographic distribution and toxicity of the populations rather than their morphological designations. Despite the social and economic impact that harmful blooms produce in Chile, few strains of A. catenella have been isolated. Moreover, physiological and/or genetic studies of the group are scarce. The aim of this work was to examine possible physiological and genetic variability among populations of A. catenella having different geographical origins but isolated from the same toxic event. Seven strains of A. catenella were isolated and established from phytoplankton samples collected in the Aysén and Los Lagos regions of southern Chile during a recent outbreak (February–March 2009). Growth, toxicity, and ITS sequences were compared among these strains. All the strains included in this study were grouped with strains belonging to the previously described “North America” clade. The genetic diversity detected among Chilean strains was 3%, a much higher value than those reported for comparisons among strains from other parts of the world. In addition, a remarkable variability of growth parameters and toxicity was detected among strains. Strain PFB45 showed the highest PSP toxin content, whereas strain PFB41 had the lowest value of this parameter but had the highest maximum cell density. In strains PFB38, PFB42, and PFB37, more than 98% of the total PSP toxin content occurred in the form of gonyautoxins (primarily GTX-4,1 and GTX-3,2). In strains PFB39, PFB36, and PFB45, neoSTX, and STX toxins were detected. These results demonstrated remarkable variability at the genetic and physiological level among strains of A. catenella isolated from the same outbreak. No correlations were found between the phenotypic traits (growth and toxicity) and the genetic affiliation of the strains studied. [Copyright &y& Elsevier]

Abstract: Blooms of Alexandrium catenella were frequently found in sea areas adjacent to the estuary of the Changjiang River in spring of the last several years, often in association with large-scale blooms of Prorocentrum donghaiense. It was considered that the increasing intensity of these dinoflagellate blooms was related to the incremental nutrient influx from the Changjiang River over the last 40 years, especially the “excess” nitrogen (N) input. A nutrient-addition experiment was then designed to study the responses of a strain (ACDH) of A. catenella, isolated from the East China Sea, to the addition of different N substrates, including nitrate, ammonium and urea. It was found that N-starved cells of strain ACDH could use all the three N substrates as the sole N source to grow, with similar specific growth rate and maximum cell density. All the three N substrates could elevate cellular toxin content shortly after the addition of nutrients, but cellular toxin content of the urea group was much lower than the nitrate group and the ammonium group. In the following days, the cellular toxin content of the urea group was even lower than the N-limited control group, but cellular toxin contents of the nitrate group and ammonium group were still comparable to the control. It was presumed, based on these findings, that a relatively slower N uptake and assimilation rate occurred in the urea group, and urea–N adsorbed was preferentially utilized for growth rather than toxin synthesis. Therefore, urea in natural seawater could promote the formation of algal blooms, but decrease the toxicity, of strain ACDH. [Copyright &y& Elsevier]

Abstract: Uptake rates of three potential N-sources (ammonium, nitrate and N-urea) and two potential C-sources (HCO3 − and C-urea) were estimated during growth of Alexandrium catenella in both light and dark phases. According to the variations observed in 13C-isotopic ratio, A. catenella cells were not able to use C-urea. Furthermore, decreases in 13C cell content during darkness revealed a probably high involvement of C recently fixed in dark respiration. Dark N-uptake capacities of A. catenella were characterized by dark/light uptake ratio of 27%, 43% and 65% for NO3 −, NH4 + and N-urea, respectively. An accumulation of C-rich compounds during the light period was highlighted through strong diel variations in C:N ratio and would provide C and energy for these dark uptake processes indicating an uncoupling between N and C metabolism. Total costs in terms of C associated with dark N-uptake and assimilation were estimated and revealed that the main part of those costs may be associated with maintenance metabolism in A. catenella cells. The relatively low C-costs of biosynthesis in darkness suggest that dark uptake and C-storage strategies correspond to a benefit in terms of competitiveness for A. catenella, optimized by the migrating abilities of this species. [Copyright &y& Elsevier]

Abstract: Alexandrium catenella (Whedon et Kofoid) Balech, a toxic dinoflagellate, is a bloom-forming planktonic species in cold water coastal regions. It produces strong paralytic shellfish poisoning (PSP) toxins which are transmitted via tainted shellfish. These toxins can affect humans, other mammals, fish and birds. In this study, polyclonal antiserum against A. catenella was produced, and a competitive enzyme-linked immunosorbent assay (cELISA) was developed to detect A. catenella. The antiserum against A. catenella showed good specificity, the linear detection range was relatively large, between 38 and 600,000 cells. In addition, specific probes were designed to target the small subunit ribosomal RNA (SSU rRNA) of A. catenella, and quantitative sandwich hybridization integrated with a nuclease protection assay (NPA-SH) was established in order to identify and quantify A. catenella. The NPA-SH assay did not show good specificity as well as cELISA, by which A. catenella and A. tamarense could not be distinguished. Samples in different cell growth phases were analyzed with cELISA and NPA-SH. The results showed that the cell concentration calculated by cELISA was very similar with microscopy, while that of NPA-SH was sometimes higher than that of microscopy, especially in log phase. Comparing the two methods, both assays allow rapid identification of A. catenella without time-consuming microscopy when multiple sites need to be considered in routine monitoring. Meanwhile, cELISA was more specific and accurate in detection of A. catenella than NPA-SH. [Copyright &y& Elsevier]

Abstract: The risk of harmful algal blooms (HABs) of the dinoflagellate Alexandrium catenella in Puget Sound, Washington State, can be assessed by identifying and predicting climate and environmental conditions that are favorable for bloom development and the accumulation of paralytic shellfish toxins (PSTs) in shellfish. When these favorable conditions occur in combination, a harmful algal bloom window of opportunity (HAB-WOO) exists for A. catenella. The original HAB-WOO was identified by for the time period 1993–2007. In general, it showed that warm air and water temperatures, low streamflow, low winds, and small tidal variability precede PST events. Here, we use the HAB-WOO model to examine (i) changes in the annual HAB-WOO duration over the period from 1967 to 2006, and (ii) the potential effect of future climate change on HAB risk through the late 21st century. The annual HAB-WOO duration increased between 1978 and 2006, as did the frequency and geographic extent of PST events. Two step-like changes occurred in 1978 and 1992 with higher annual values attained by the HAB-WOO compared to previous years. The 1978 step change may be related to the 1977 reversal of the Pacific Decadal Oscillation from cool to warm phase. Climate change projections from global climate models and regionally downscaled climate models for the Pacific Northwest are used to evaluate scenarios for the future HAB-WOO. Under a moderate greenhouse gas emissions scenario (i.e., A1B), the annual HAB-WOO for A. catenella in Puget Sound is projected to increase by an average of 13 days by the end of the 21st century. Furthermore, the annual HAB-WOO may begin up to 2 months earlier in the year and persist for up to 1 month later in the year compared to the present day. This research provides managers, health authorities, and shellfish growers in Washington State with critical information for anticipating climate impacts on toxic HABs in the Pacific Northwest now and in a future warmer climate. [Copyright &y& Elsevier]