Intra-population clonal variability in allelochemical potency of the toxigenic dinoflagellate Alexandrium tamarense

Clonal variability in exponential growth rate and production of secondary metabolites was determined from clonal isolates of Alexandrium tamarense originating from a single geographical population from the east coast of Scotland. To assess variability in the selected phenotypic characteristics over a wide spectrum, 10 clones were chosen for experimentation from 67 clonal isolates pre-screened for their lytic capacity in a standardized bioassay with the cryptophyte Rhodomonas salina. Specific growth rates (μ) of the 10 clonal isolates ranged from 0.28 to 0.46 d⁻¹ and were significantly different among clones. Cell content (fmol cell⁻¹) and composition (mol%) of paralytic shellfish toxins (PSTs), analyzed by liquid chromatography with fluorescence detection (LC–FD), varied widely among these isolates, with total PST quotas ranging from 20 to 89 fmol cell⁻¹. Except for strain 3, the toxins C1/C2, neosaxitoxin (NEO), saxitoxin (STX), and gonyautoxins-1 and -4 (GTX1/GTX4), were consistently the most relatively abundant, with lesser amounts of GTX2/GTX3 evident among all isolates. Only clone 3 contained >20 mol% of toxin B1, with C1/C2, GTX2/GTX3 and NEO in almost equimolar ratios.Eight of the 10 clones caused cell lysis of both R. salina and the heterotrophic dinoflagellate Oxyrrhis marina, as quantified from the dose–response curves from short-term (24 h) co-incubation bioassays. For two clones, no significant mortality even at high Alexandrium cell concentrations (ca. 10⁴ mL⁻¹) was observed. Allelochemical activity expressed as EC₅₀ values, defined as the Alexandrium cell concentration causing lysis of 50% of target cells, varied by about an order of magnitude and was significantly different among clones. No correlation was observed between growth rate und allelochemical potency (as EC₅₀) indicating that at least under non-limiting growth conditions no obvious growth reducing costs are associated with the production of allelochemically active secondary metabolites.     

Interactions between Prorocentrum donghaiense Lu and Scrippsiella trochoidea (Stein) Loeblich III under laboratory culture

Interactions between Prorocentrum donghaiense Lu and Scrippsiella trochoidea (Stein) Loeblich III, two species of causative bloom dinoflagellates in China, were investigated using bi-algal cultures under controlled laboratory conditions. The growth of P. donghaiense and S. trochoidea were significantly suppressed when the initial cell densities were set at 1.9 × 10⁴ cells mL⁻¹ or 1.9 × 10⁵ cells mL⁻¹ for P. donghaiense and 1.0 × 10⁴ cells mL⁻¹ for S. trochoidea when the initial size/density ratio was 1:1 or 10:1, respectively, but no out-competement was observed in either bi-algal culture by the end. The simultaneous assay on the culture filtrate showed that P. donghaiense filtrate prepared at a lower initial density (1.9 × 10⁴ cells mL⁻¹) stimulated the co-cultured S. trochoidea at a density of 1.0 × 10⁴ cells mL⁻¹, but filtrate at a higher density (1.9 × 10⁵ cells mL⁻¹) depressed its growth. Differently, the filtrate of S. trochoidea at a density of 1.0 × 10⁴ cells mL⁻¹ significantly suppressed the growth of P. donghaiense at a density of 1.9 × 10⁴ cells mL⁻¹, but had little stimulatory effect on P. donghaiense at a density of 1.9 × 10⁵ cells mL⁻¹compared to the control (P > 0.05). It is likely that these two species of microalgae interact with each other mainly by releasing allelochemical substance(s) into the culture medium, and a direct cell-to-cell contact was not necessary for their mutual interaction. We then quantify their interactions in the bi-algal culture by using a mathematical model. The estimated parameters from the model showed that the inhibition exerted by S. trochoidea on P. donghaiense was about 43 and 24 times stronger than the inhibitory effect that P. donghaiense exerted on S. trochoidea when the initial size/density were 1:1 and 10:1, respectively. S. trochoidea seemed to have a survival strategy that was superior to P. donghaiense in the bi-algal culture under controlled laboratory conditions. We also observed a closely positive relationship between the initial cell density and its effect on the co-cultured microalga by measuring the fluorenscence: filtrate prepared from higher initial cell density had stronger interference on the co-cultured microalga. Moreover, pre-treated under different temperature conditions (30 °C, 60 °C and 100 °C) would significantly changed the effect of culture filtrate on the co-cultured microalga. Result inferred that P. donghaiense or S. trochoidea would release allelochemicals into the bi-algal culture medium and the allelochemicals might be a mixture with temperature-sensitive components in it.     

Comparative studies on morphology, ITS sequence and protein profile of Alexandrium tamarense and A. catenella isolated from the China Sea

The Alexandrium tamarense species complex is a closely related cosmopolitan toxigenic group of morphology-based species, including A. tamarense, A. catenella and A. fundyense. This study investigated the morphology, internal transcribed spacer (ITS) sequence and protein profile of A. tamarense and A. catenella grown in the same culture conditions using a combination of scanning electronic microscope (SEM), molecular and proteomic approaches. The results showed that all Alexandrium strains had the plate formula of Po, 4′, 6″, 6C, 8S, 5″′, 2″″. The ventral pore, a key conventional morphological feature to discriminate A. tamarense and A. catenella, was usually present in the first apical plate of ten A. tamarense strains, however, it was found to be absent in some cells of one Alexandrium strain, ATGX01. A. tamarense and A. catenella shared an identical ITS sequence with a minor variation at intraspecific level. Protein profiles of A. catenella DH01 and A. tamarense DH01, isolated from the same region of the East China Sea, showed no significant difference, the similarity of protein profiles of the two species reached 99% with a few proteins unique to one or the other. The present results suggest that the ventral pore is not a consistent morphological feature in the Alexandrium genus, and that A. tamarense and A. catenella are conspecific and should be redesignated to one species.     

Experimental study on the impact of dinoflagellate Alexandrium species on populations of the rotifer Brachionus plicatilis

To investigate harmful effects of the dinoflagellate Alexandrium species on microzooplankton, the rotifer Brachionus plicatilis was chosen as an assay species, and tested with 10 strains of Alexandrium including one known non-PSP-producer (Alexandrium tamarense, AT-6). HPLC analysis confirmed the PSP-content of the various strains: Alexandrium lusitanicum, Alexandrium minutum and Alexandrium tamarense (ATHK, AT5-1, AT5-3, ATCI02, ATCI03) used in the experiment were PSP-producers. No PSP toxins were detected in the strains Alexandrium sp1, Alexandrium sp2.Exposing rotifer populations to the densities of 2000 cells ml⁻¹ of each of these 10 Alexandrium strains revealed that the (non-PSP) A. tamarense (AT-6) and two other PSP-producing algae: A. lusitanicum, A. minutum, did not appear to adversely impact rotifer populations. Rotifers exposed to these three strains were able to maintain their population numbers, and in some cases, increase them. Although some increases in rotifer population growth following exposures to these three algal species were noted, the rate was less than for the non-exposed control rotifer groups.In contrast, the remaining seven algal strains (A. tamarense ATHK, AT5-1, AT5-3, ATCI02, ATCI03; also Alexandrium sp1 and Alexandrium sp2) all have adverse effects on the rotifers. Dosing rotifers with respective algal cell densities of 2000 cells ml⁻¹ each, for Alexandrium sp1, Alexandrium sp2, and A. tamarense strains ATHK and ATCI03 showed mean lethal time (LT₅₀) on rotifer populations of 21, 28, 29, and 36h, respectively. The remaining three species (A. tamarense strains AT5-1, AT5-3, ATCI02) caused respective mean rotifer LT₅₀s of 56, 56, and 71 h, compared to 160 h for the unexposed “starved control” rotifers. Experiments to determine ingestion rates for the rotifers, based on changes in their Chlorophyll a content, showed that the rotifers could feed on A. lusitanicum, A. minutum and A. tamarense strain AT-6, but could graze to little or no extent upon algal cells of the other seven strains. The effects on rotifers exposed to different cell densities, fractions, and growth phases of A. tamarense algal culture were respectively compared. It was found that only the whole algal cells had lethal effects, with strongest impact being shown by the early exponential growth phase of A. tamarense. The results indicate that some toxic mechanism(s), other than PSP and present in whole algal cells, might be responsible for the adverse effects on the exposed rotifers.     

Development of a qualitative PCR method for the Alexandrium spp. (Dinophyceae) detection in contaminated mussels (Mytilus galloprovincialis)

Paralytic shellfish poisoning (PSP) is a syndrome caused by the consumption of shellfish contaminated with neurotoxins produced by organisms of the marine dinoflagellate genus Alexandrium. A. minutum is the most widespread species responsible for PSP in the Western Mediterranean basin. The standard monitoring of shellfish farms for the presence of harmful algae and related toxins usually requires the microscopic examination of phytoplankton populations, bioassays and toxin determination by HPLC. These procedures are time-consuming and require remarkable experience, thus limiting the number of specimens that can be analyzed by a single laboratory unit. Molecular biology techniques may be helpful in the detection of target microorganisms in field samples. In this study, we developed a qualitative PCR assay for the rapid detection of all potentially toxic species belonging to the Alexandrium genus and specifically A. minutum, in contaminated mussels. Alexandrium genus-specific primers were designed to target the 5.8S rDNA region, while an A. minutum species-specific primer was designed to bind in the ITS1 region. The assay was validated using several fixed seawater samples from the Mediterranean basin, which were analyzed using PCR along with standard microscopy procedures. The assay provided a rapid method for monitoring the presence of Alexandrium spp. in mussel tissues, as well as in seawater samples. The results showed that PCR is a valid, rapid alternative procedure for the detection of target phytoplankton species either in seawater or directly in mussels, where microalgae can accumulate.     

The Inhibitory Effects of Garlic (<Emphasis Type="Italic">Allium sativum</Emphasis>) and Diallyl Trisulfide on <Emphasis Type="Italic">Alexandrium tamarense</Emphasis> and other Harmful Algal Species

Using cell suspension ability as an indicator, we studied the inhibitory effect of garlic (Allium sativum) and diallyl trisulfide on six species of red tide causing algae. This included: the inhibition by 0.08% garlic solution of five algal species — Alexandrium tamarense, Scrippsiella trochoidea, Alexandrium catenella, Alexandrium minutum and Alexandrium satoanum; the effects of garlic concentration on the inhibition of A. tamarense, S. trochoidea and Chaetoceros sp.; the effects of inhibitory time on the rejuvenation of algal cells; and the effects of heating and preservation time on algal inhibition by garlic solution. In addition, whether or not the ingredients of garlic solution had a possible algicidal effect was studied by comparing inhibition of A. tamarense by garlic solution and man-made diallyl trisulfide. The results showed that 1) inhibition by garlic solution was significant on A. tamarense, A. satoanum, A. catenella and S. trochoidea, and the least effective was a concentration of 0.04% on A. tamarense and S. trochoidea. Moreover, the higher the concentration, the stronger was the inhibition, and a high inhibitory rate (IR) could be maintained for at least three days when the garlic concentration was above 0.04%. For A. tamarense, it was also found that the longer the inhibitory time and the higher the concentration, the lower was the rate of resumed cell activity. On the contrary, garlic solution could not inhibit A. minutum or Chaetoceros sp.; 2) The IR to A. tamarense was reduced slightly as the heating time of the garlic solution was lengthened, but the average IR was still above 80%. There was no significant difference between the IR of the supernatant and sediment of the garlic solution. Furthermore, no change of algal inhibition was found when the garlic solution was preserved at 20°C for several days; 3) As with garlic solution, diallyl trisulfide inhibited A. tamarense strongly; the IR was above 93% and was maintained for at least three days, as long as the concentration was 3.2–10.0 mg L⁻¹. Thus, diallyl trisulfide may have been the major ingredient in garlic solution which inhibited the algae but, in addition, more than one ingredient may have been inhibiting the algae. In conclusion, garlic was a good algal inhibitor with many advantages, such as being common, cheap, non toxic and with high efficiency, and diallyl trisulfide, one of the components of garlic, was similarly effective in algal inhibition. It would be useful, therefore, to further study garlic as an environmentally friendly algal inhibitor.     

Morphogenetic diversity and biotoxin composition of Alexandrium (Dinophyceae) in Irish coastal waters

The diversity of Alexandrium spp. in Irish coastal waters was investigated through the morphological examination of resting cysts and vegetative cells, the determination of PSP toxin and spirolide profiles and the sequence analysis of rDNA genes. Six morphospecies were characterised: A. tamarense, A. minutum, A. ostenfeldii, A. peruvianum, A. tamutum and A. andersoni. Both PSP toxin producing and non-toxic strains of A. tamarense and A. minutum were observed. The average toxicities of toxic strains for both cultured species were respectively 11.3 (8.6 S.D.) and 2.3 (0.5 S.D.) pg STX equiv. cell⁻¹. Alexandrium ostenfeldii and A. peruvianum did not synthesise PSP toxins but HPLC–MS analysis of two strains showed distinct spirolide profiles. A cyst-derived culture of A. peruvianum from Lough Swilly mainly produced spirolides 13 desmethyl-C and 13 desmethyl-D whereas one of A. ostenfeldii, from Bantry Bay, produced spirolides C and D. Species identification was confirmed through the analyses of SSU, ITS1-5.8S-ITS2 and LSU rDNA genes. Some nucleotide variability was observed among clones of toxic strains of A. tamarense, which all clustered within the North American clade. However, rDNA sequencing did not allow discrimination between the toxic and non-toxic forms of A. minutum. Phylogenetic analysis also permitted the differentiation of A. ostenfeldii from A. peruvianum. Resting cysts of PSP toxin producing Alexandrium species were found in Cork Harbour and Belfast Lough, locations where shellfish contamination events have occurred in the past, highlighting the potential for the initiation of harmful blooms from cyst beds. The finding of supposedly non-toxic and biotoxin-producing Alexandrium species near aquaculture production sites will necessitate the use of reliable discriminative methods in phytoplankton monitoring.     

Detection and quantification of cultured marine Alexandrium species by real-time PCR

The occurrence of harmful algal blooms (HABs) throughout the world has increased and poses a large threat to human health, fishery resources and tourism industries. The genus Alexandrium includes a number of toxic species associated with HABs. Therefore, it is very important to rapidly detect and monitor the harmful algae, such as Alexandrium genus. In this study, a standard curve of plasmid containing 18S rDNA-28S rDNA region from Alexandrium catenella was constructed and 5.8S rDNA sequence served as the primer of the real-time PCR. Cultured A. catenella, Alexandrium affine, Alexandrium lusitanicum and Alexandrium minutum samples were analyzed by real-time PCR using the same set of primers simultaneously. Using microscopy cells counts, 5.8S rDNA copies per cell and total DNA per cell were estimated. This assay method is promising for rapid detection of large number of Alexandrium samples.     

Development of Molecular Identification Method for Genus Alexandrium (Dinophyceae) Using Whole-Cell FISH

We have developed a method to identify species in the genus Alexandrium using whole-cell fluorescent in situ hybridization with FITC-labeled oligonucleotide probes that target large subunit ribosomal rRNA molecules. The probes were designed based on the sequence of the rDNA D1-D2 region of Alexandrium species. DNA probes specific for toxic A. tamarense and A. catenella and nontoxic A. affine, A. fraterculus, A. insuetum, and A. pseudogonyaulax, respectively, were applied to vegetative cells of all above Alexandrium species to test the sensitivity of the probes. Each DNA probe hybridized specifically with vegetative cells of the corresponding Alexandrium species and showed no cross-reactivity to noncorresponding Alexandrium species. In addition, no cross-reactivity of the probes was observed in experiments using concentrated natural seawater samples. The TAMAD2 probe, which is highly specific to A. tamarense, a common toxic species in Korean coastal waters, provides a simple and reliable molecular tool for identification of toxic Alexandrium species.     

The first description of the potentially toxic dinoflagellate, Alexandrium minutum in Hunts Bay, Kingston Harbour, Jamaica

The occurrence and morphology of the potentially toxic dinoflagellate species Alexandrium minutum found for the first time in Jamaica, were examined and described by light and scanning electron microscopy. Classical morphological examinations of whole cells, the thecal plate pattern of intact cells and more importantly the structure of individual thecal plates of squashed cells, were conducted in an attempt to positively identify the species. Characteristics such as a tear-drop shaped apical pore plate with a comma-shaped apical pore and no anterior attachment pore; a narrow sixth precingular plate; a narrow anterior sulcal plate longer than or approximately as long as it is wide; and a posterior sulcal plate wider than long, confirmed the Jamaican species as A. minutum. This dinoflagellate which produces potent neurotoxins responsible for paralytic shellfish poisoning (PSP) in humans in many parts of the World, as well as mass mortality of various marine flora and fauna, was identified in water samples collected during an extensive bloom of the species in the brackish to saline water body of Hunts Bay, an estuarine arm of Kingston Harbour, Jamaica in August 1994. The highest cell concentration was 4.6 × 10⁵ cells l⁻¹, a concentration which far exceeds acceptable concentrations (<10³ cells l⁻¹) of PSP-toxin producing A. minutum in several countries including: Spain and Denmark. No PSP human symptoms were reported during the bloom; however it was accompanied by a large kill of small pelagic fish extending across a third of the bay. Since then, smaller blooms of A. minutum have occurred with the most recent in February and April 2004. Hunts Bay is an important fishing, shrimping and to some extent oyster/mussel collection area and provides an important source of livelihood and food for many fishermen in nearby fishing communities as well as an important source of food for members of other communities. Although there are no known records of human illness due to PSP in Jamaica, the occurrence and blooming in Jamaican waters of this potentially toxic dinoflagellate, is great cause for concern.     

Copepod feeding response to varying Alexandrium spp. cellular toxicity and cell concentration among natural plankton samples

We challenged four species of copepod grazers (Acartia hudsonica, Centropages hamatus, Eurytemora herdmani, Calanus finmarchicus) with natural water samples containing non-toxic algae mixed with one of three clones of Alexandrium spp.—A. tamarense GTCN16 (non-toxic), A. fundyense GTCA28 (moderate toxicity), and A. fundyense BC1 (higher toxicity), each at relatively high (10⁵ cells L⁻¹) and low (10⁴ cells L⁻¹) concentrations. Within any one copepod species, significant differences existed in copepod clearance rates and total food ingested between high and low Alexandrium cell concentrations, and between levels of toxicity, but feeding response did not follow a predictable relationship proportional to toxin levels—rather, the presence or absence of toxin was more important than the level of toxicity. C. finmarchicus behaved differently from the smaller coastal copepods, showing less selectivity and greater concentration dependence. In low Alexandrium concentration treatments, copepod clearance rates on Alexandrium were usually higher, and electivity indices for Alexandrium less negative (indicating less avoidance), compared to high concentration treatments. In high toxicity (BC1) treatments of all copepod species (except C. finmarchicus), total food consumption was consistently less at high Alexandrium concentrations compared to low concentrations, suggesting that high toxicity and concentration suppress overall feeding, while in non-toxic (GTCN 16) treatments total consumption was always higher at high Alexandrium concentrations. Copepod grazers do not follow predictable feeding rules throughout a continuum of conditions, but become more predictable at extremes of concentration and toxicity of prey, consistent with the conclusion that both factors are important. Results support the hypothesis that grazer deterrence imparted by toxicity is only effective at high cell concentrations, but even then will not protect against all grazers.     

Karlotoxin mediates grazing by Oxyrrhis marina on strains of Karlodinium veneficum

Karlodinium veneficum is a common member of temperate, coastal phytoplankton assemblages that occasionally forms blooms associated with fish kills. Here, we tested the hypothesis that the cytotoxic and ichthyotoxic compounds produced by K. veneficum, karlotoxins, can have anti-grazing properties against the heterotrophic dinoflagellate, Oxyrrhis marina. The sterol composition of O. marina (>80% cholesterol) renders it sensitive to karlotoxin, and does not vary substantially when fed different algal diets even for prey that are resistant to karlotoxin. At in situ bloom concentrations (10⁴–10⁵ K. veneficum ml⁻¹), grazing rates (cells ingested per Oxyrrhis h⁻¹) on toxic K. veneficum strain CCMP 2064 were 55% that observed on the non-toxic K. veneficum strain MD5. At lower prey concentrations typical of in situ non-bloom levels (<10³ cells ml⁻¹), grazing rates (cells ingested per Oxyrrhis h⁻¹) on toxic K. veneficum strain CCMP 2064 were 70–80% of rates on non-toxic strain MD5. Growth of O. marina was significantly suppressed when fed the toxic strain of K. veneficum. Experiments with mixed prey cultures, where non-toxic strain MD5 was fluorescently stained, showed that the presence of toxic strain CCMP 2064 inhibited grazing of O. marina on the co-occurring non-toxic strain MD5. Exogenous addition of a sub-lethal dose (100 ng ml⁻¹) of purified karlotoxin inhibited grazing of O. marina by approximately 50% on the non-toxic K. veneficum strain MD5 or the cryptophyte S. major. These results identify karlotoxin as an anti-grazing compound for those grazers with appropriate sterol composition (i.e., desmethyl sterols). This strategy is likely to be an important mechanism whereby growth of K. veneficum is uncoupled from losses due to grazing, allowing it to form ichthyotoxic blooms in situ.     

Occurrence and toxicity of Amphidinium carterae Hulburt in the North Arabian Sea

The occurrence and toxicity of Amphidinium carterae Hulburt is hereby reported for the first time from the North Arabian Sea on the coast of Pakistan. The concentrations of 1.2 × 10⁴ cells ml⁻¹ were found in intertidal pools that were also inhabited by the brown macroalga Sargassum wightii. Both wild and cultured A. carterae cells were tested for ciguatera toxicity through exposure to brine shrimp nauplii (Artemia salina) and albino mice. Although the brine shrimp did not appear to be affected mortalities in mice ranged between 13 and 16% at doses of 7.2 × 10⁴ and 2.5 × 10⁵ cells ml⁻¹, respectively. When mice were affected pharmacological effects such as muscle contraction in lower back area, increased respiration, immobility and paralysis in hind limbs were observed for 2 h. These effects appeared to be reversible and gradually disappeared within 24 h.     

Toxic Alexandrium minutum (Dinophyceae) from Vietnam with new gonyautoxin analogue

Clonal cultures of Alexandrium species collected from a shrimp pond on the northern coast of Vietnam were established and morphologically identified as Alexandrium minutum. Nucleotide sequences of domains 1 and 2 of the large subunit ribosomal (LSU) rRNA gene showed high sequence similarity to A. minutum isolates from Malaysia. Paralytic shellfish toxin profile of the clones was characterized by the dominance of GTX4, GTX1, and NEO. GTX3, GTX2, and dcSTX were also present in trace amount. Toxin content varied among the strains and growth stages, ranged from 3.0 to 12.5 fmol cell⁻¹. In addition to these known toxin components, a new gonyautoxin derivative was detected by HPLC, eluting between GTX4 and GTX1. The peak of this compound disappeared under non-oxidizing HPLC condition but unchanged either after treated with 0.05 M ammonium phosphate/10% mercaptoethanol or 0.1N HCl hydrolysis. LCMS ion scanning showed a parental ion of [M + H]⁺ at m/z 396, [M − SO₃]⁺ at m/z 316, and [M − SO₄]⁺ at m/z 298. Based on these results, the derivative was identified as deoxy-GTX4-12ol, and this represents the first report of this toxin analogue.     

Toxic mucus traps: A novel mechanism that mediates prey uptake in the mixotrophic dinoflagellate Alexandrium pseudogonyaulax

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 (500 cells ml⁻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.     

Small-scale turbulence affects the division rate and morphology of two red-tide dinoflagellates

The effects of small-scale turbulence on two species of dinoflagellates were examined in cultures where the turbulent forces came randomly from all directions and were intermittent both spatially and temporally; much like small-scale turbulence in the ocean. With Lingulodinium polyedrum (Stein) Dodge (syn. Gonyaulax polyedra), division rate increased linearly (from 0.35 to 0.5 per day) and the mean cross-sectional area (CSA) decreased linearly (from 1100 to 750 μm²) as a function of the logarithmic increase in turbulence energy dissipation rate (). These effects were noted when values increased between 10⁻⁸ and 10⁻⁴ m² s⁻³. However, when increased to 10⁻³ m² s⁻³, division rate sharply decreased and mean CSA increased. Over the same range of , Alexandrium catenella (Wheedon and Kofoid) Balech had its division rate decrease linearly (from 0.6 to 0.45 per day) and its CSA increase linearly (from 560 to 650 μm²) as a function of the logarithmic increase in . Even at the highest examined (10⁻³ m² s⁻³), which may be unrealistically high for their ambits, both L. polyedra and A. catenella still had fairly high division rates, 0.2 and 0.45 per day, respectively. Turbulence strongly affected chain formation in A. catenella. In non-turbulent cultures, the mode was single cells (80–90% of the population), but at of 10⁻⁵ to 10⁻⁴ m² s⁻³, the mode was 8 cells per chain. At the highest (10⁻³ m² s⁻³), the mode decreased to 4 cells per chain. The vertical distributions of A. catenella populations in relation to hydrographic flow fields were studied in the summers of 1997 and 1998 in East Sound, Washington, USA (latitude 48°39′N, 122°53′W). In both summers, high concentrations of A. catenella were found as a subsurface bloom in a narrow depth interval (2 m), where both current shear and turbulence intensity were at a minimum. Other researchers have shown that A. catenella orients its swimming in shear flows, and that swimming speed increases with chain length. These responses, when combined with our observations, support a hypothesis that A. catenella actively concentrates at depths with low turbulence and shear.     

Toxin profiles of natural populations and cultures ofAlexandrium minutum Halim from Galician (Spain) coastal waters

The toxin profiles of three isolates and natural populations of the PSP agentAlexandrium minutum from several Galician rías (NW Spain) was obtained by HPLC. The toxin content of cultures ofA. minutum is dominated by GTX4 (80–90%) and GTX4 (10–15%) with small amounts of GTX3 and GTX2 (less than 3% of each); similar results were obtained for natural populations ofAlexandrium from three different Galician rías, where a mixture ofA. lusitanicum Balech andA. minutum can occur. Important quantitative differences were found between the three isolates, one being highly and two weakly toxic. The results obtained from these isolates and natural populations ofAlexandrium were very similar to those obtained from HPLC analyses of mussels intoxicated during a PSP outbreak in Ría de Ares (Rías Altas) in 1984, confirming thatA. minutum (previously identified asGonyaulax tamarensis Lebour andAlexandrium lusitanicum) was the PSP agent during the toxic outbreak in May 1984. Toxin profiles obtained from natural populations during different PSP outbreaks in different rías and from cultures are fairly consistent and suggest that at least from the toxin point of view,A. lusitanicum andA. minutum are identical, and that the toxin profile ofA. minutum from Galicia can be used as a biochemical marker.     

Toxicity of Dinophysis spp. in relation to population density and environmental conditions on the Swedish west coast

The aim of this study in the field was to investigate whether there are differences between the outer archipelago (Gullmar Fjord) and a semi-enclosed fjord system (Koljö Fjord) in occurrences of D. acuta and D. acuminata as well as in their content of diarrheic shellfish toxin (DST) per cell. When all data pairs of cell toxicity of D. acuminata and the corresponding number of cells l⁻¹ from the two sites were tested in a regression analysis, a statistically significant negative correlation became evident and was apparent as a straight line on a log–log plot (p < 0.0001). Obviously, there was an overall inverse relationship between the population density of D. acuminata and the toxin content per cell. Plotted on a linear scale, all data-pairs of cell toxicity and cell number made up a parabolic curve. On this curve the data-pairs could be separated into three groups: (i) D. acuminata occurring in numbers of fewer than approximately 100 cells l⁻¹, and with a toxin content per cell above 5 ρg cell⁻¹; (ii) cell numbers between 100 and approximately 250 cells l⁻¹ with a cell toxin content from 5 to 2 ρg cell⁻¹; (iii) when the population became greater than 250 cells l⁻¹, the toxicity, with few exceptions, was less than 2 ρg cell⁻¹. By applying this subdivision, some clear patterns of the distribution of the differently toxic D. acuminata became evident. When comparing the cell toxicity of the two sites, it was obvious that the D. acuminata cells from all depths from the Gullmar Fjord as a mean were significantly more toxic compared to the Koljö Fjord samples. The results have demonstrated that approximately 100 high-toxicity cells in a low-density population at surface may lead to the same accumulation of DST in a mussel as the ingestion of 1500 low-toxicity cells from a high-density pycnocline population.