Development of microsatellite markers in the noxious red tide‐causing dinoflagellate Heterocapsa circularisquama (Dinophyceae)

We isolated 15 polymorphic microsatellites from one of the most noxious red tide‐causing dinoflagellate species, Heterocapsa circularisquama. These loci provide one class of highly variable genetic markers, as the number of alleles ranged from two to six, and the estimate of gene diversity from 0.205 to 0.684 across the 15 microsatellites. These loci have the potential to reveal genetic structure and gene flow among H. circularisquama populations.     

THIS ARTICLE HAS BEEN RETRACTEDDevelopment of microsatellite markers in the noxious red tide‐causing dinoflagellate Heterocapsa circularisquama (Dinophyceae)

We isolated 15 polymorphic microsatellites from one of the most noxious red tide‐causing dinoflagellate species, Heterocapsa circularisquama. These loci provide one class of highly variable genetic markers, as the number of alleles ranged from two to six, and the estimate of gene diversity was from 0.205 to 0.684 across the 15 microsatellites. These loci have the potential to reveal genetic structure and gene flow among H. circularisquama populations.     

HETEROCAPSA ARCTICA SUBSP. FRIGIDA SUBSP. NOV. (PERIDINIALES,DINOPHYCEAE)—DESCRIPTION OF A NEW DINOFLAGELLATE AND ITS OCCURRENCE IN THE BALTIC SEA1

Characteristics important in identification of Heterocapsa species (i.e., thecal plate pattern, body scale structure, and shape and position of the nucleus and pyrenoid) are practically identical in the dinoflagellate investigated here and in Heterocapsa arctica T. Horig. described from the Canadian Arctic. Analysis of internal transcribed spacer (ITS) sequences confirms that the two dinoflagellates are very closely related; however, there is a clear difference in their size and shape. Our experiments show that the low‐salinity Baltic Sea brackish water does not reduce the size of the marine H. arctica to match that of the Baltic Sea morphotype. On the basis of these dissimilarities in general morphology and its geographic isolation in the Baltic Sea, we consider our material sufficiently differentiated from the typical H. arctica to warrant the status of a new subspecies, H. arctica subsp. frigida subsp. nov. Being of a distinct cell shape, the occurrence of subsp. frigida has been recorded in Algaline phytoplankton monitoring data collected since 1993. Although it has never been responsible for high biomass blooms, it commonly occurs in spring in the Northern Baltic Proper and in the western Gulf of Finland, when the water temperatures are <5°C.     

GEOTAXIS/PHOTOTAXIS AND BIOCHEMICAL PATTERNS IN HETEROCAPSA (=CACHONINA) ILLDEFINA (DINOPHYCEAE) DURING DIEL VERTICAL MIGRATIONS

Two separate experiments with Heterocapsa (= Cachonina ) illdefina Herman et Sweeney, one with and the other without water volume replacement, were performed in a 250-L laboratory mesocosm (45-cm diameter × 150-cm height) to examine how diel vertical migration (DVM) relates to taxis sign and strength and to cellular biochemical state. Although only the cell population grown with water volume replacement maintained a division per day over the course of the experiment, periodic measurements during both experiments demonstrated that cells aggregating at the surface during the light period generally were deficient in all measured biochemical constituents compared to cells obtained from a midcolumn depth. More specifically, H. illdefina cells that aggregated at the surface during the light period in both experiments exhibited weakened positive geotaxis but strengthened positive phototaxis and were very deficient in lipid and free amino acid compared to midcolumn cells. Cells sampled at midcolumn during the light period exhibited similar but weaker taxes changes compared to surface samples, and geotaxis strength was inversely correlated with cell diameter, cellular DNA and protein content, and RNA/DNA ratio. In comparison, published data on Gymnodinium breve Davis, a harmful algal bloom species, showed that cells aggregating at the surface during the light period generally exhibited weakened negative geotaxis and strengthened positive phototaxis and were very deficient in lipid and chl a compared to midcolumn cells. Although the persistent tendency toward negative geotaxis was weaker in midcolumn subpopulations throughout the day, its strength was inversely correlated with cell diameter and cellular lipid content. The combined results for both species support a revised conceptual model of optimized DVM in autotrophic marine dinoflagellates incorporating generalized expressions of taxis and biochemical state of individual cells.     

IDENTIFICATION AND TOXICITY OF ALEXANDRIUM TAMARENSE (DINOPHYCEAE) IN SCOTTISH WATERS1

Contamination of shellfish with paralytic shellfish poisoning (PSP) toxins produced by Alexandrium species poses a potential threat to the sustainability of the Scottish aquaculture industry. Routine LM analysis of water samples from around the Scottish coast has previously identified Alexandrium (Dinophyceae) as a regular part of the spring and summer phytoplankton communities in Scottish coastal waters. In this study, Alexandrium tamarense (M. Lebour) Balech isolated from sediment and water samples was established in laboratory culture. Species identification of these isolates was confirmed using thecal plate dissections and by molecular characterization based on their LSU and, in some cases, ITS rDNA sequence. Molecular characterization and phylogenetic analysis showed the presence of two ribotypes of A. tamarense: Group I (North American ribotype) and Group III (Western European ribotype). Assessment of PSP toxin production using hydrophilic interaction liquid chromatography–tandem mass spectrometry (HILIC–MS/MS) showed that A. tamarense Group I produced a complex array of toxins (～2,000 fg STX equivalents · cell^?1) with the major toxins being C2, neosaxitoxin (NEO), saxitoxin (STX), gonyautoxin‐4 (GTX‐4), and GTX‐3, while A. tamarense Group III did not produce toxins. Historically, it was considered that all Alexandrium species occurring in Scottish waters produce potent PSP toxins. This study has highlighted the presence of both PSP toxin‐producing and benign species of A. tamarense and questions the ecological significance of this finding.     

Tn5 MUTAGENESIS OF PSEUDOMONAS STUTZERI SF/PS, A BACTERIUM ASSOCIATED WITH ALEXANDRIUM LUSITANICUM (DINOPHYCEAE) AND PARALYTIC SHELLFISH POISONING

It is becoming increasingly clear that bacteria can play an important role in the toxin and population dynamics of harmful algal bloom (HAB) events. In this paper, we document protocols and strategies that can be used to identify bacterial genes involved in either the production of toxic compounds and/or the establishment and maintenance of relationships between bacteria and algae. The protocols we tested involved transposon mutagenesis and complementation with broad-host-range plasmids. We tested six bacterial strains thought to be involved, either directly or indirectly, in the production of toxins associated with paralytic shellfish poisoning (PSP). Five strains were resistant to transformation under the growth conditions used. However, a single strain, Pseudomonas stutzeri SF/PS, was readily transformed when grown under appropriate conditions. This bacterium has been shown to accumulate PSP toxins and to increase toxin production when added to axenic cultures of a toxic dinoflagellate, Alexandrium lusitanicum . We conclude that a transposon mutagenesis strategy can be used to identify genes involved in HAB events.     

STEROL DISTRIBUTION IN THE GENUS HETEROCAPSA (PYRRHOPHYTA)1

The sterol composition of seven strains of marine peridinioid dinoflagellates comprising the four known species of Heterocapsa Stein was examined by gas chromatography-mass spectrometry to determine the utility of these compounds in systematics. Cholest-5-en-3β-ol (cholesterol), 24-methyl-cholest-5-en-3β-ol (24-methylcholesterol), 4α,24(S)-dimethyl-5α-cholestan-3β-ol (4,24-dimethylcholestanol), 4α,23,24(R)-trimethyl-5α-cholest-22-en-3β-ol (dinosterol), 4α,23ξ,24ξ-trimethyl-5α-cholestan-3β-ol (dihydrodinosterol), and an unknown sterol were detected. Sterol composition does not vary significantly from species to species within the genus Heterocapsa and thus cannot be used for species differentiation. Sterols may, however, have value in defining the properties of dinoflagellate taxa above the family level. Over the course of the growth curve for Heterocapsa niei (Loeblich) Morrill & Loeblich 4,24-dimethylcholestanol and dinosterol covaried, suggesting that 4,24-dimethylcholestanol is converted into dinosterol by a previously proposed bioalkylation scheme.     

Comparative Study on the Toxic Effects of Red Tide Flagellates Heterocapsa circularisquama and Chattonella marina on the Short-Necked Clam (Ruditapes philippinarum)

Heterocapsa circularisquama showed much higher toxic effects on short-necked clams than Chattonella marina. Clams exposed to H. circularisquama exhibited morphological changes concomitant with an accumulation of mucus-like substances in the gills, a profound reduction in filtration activity, and lysosomal destabilization in hemocytes. Chattonella marina was less effective than H. circularisquama, and Heterocapsa triquetra was almost harmless in all these criteria. These results suggest that H. circularisquama exerted its lethal effect on short-necked clams through gill tissue damage and subsequent induction of physiological stress.     

THE MECHANISMS OF FLUORIDE TOXICITY AND FLUORIDE RESISTANCE IN SYNECHOCOCCUS LEOPOLIENSIS (CYANOPHYCEAE)1

Fluoride was supplied as dissolved NaF at concentrations ranging from 0.26 to 7.9 mM (5–150 ppm) to three freshwater microalgae: Synechococcus leopoliensis (Racib.) Komarek (Cyanophyta), Oscillatoria limnetica Lemmermann (Cyanophyta) and Chlorella pyrenoidosa Chick (Chlorophyta). Growth of C. pyrenoidosa was unaffected by fluoride, and uptake of fluoride by this organism was not detectable. Growth of the cyanophytes was temporarily inhibited by NaF. The duration of this growth lag increased markedly as the pH was lowered at constant external fluoride concentration. In S. leopoliensis, fluoride uptake and inhibition of photosynthesis by NaF increased in the same way as did the growth lag in response to pH. Growth-inhibitory NaF treatments decreased the ATP level in cells of S. leopoliensis by 75% and also abolished phosphate uptake. Cells of S. leopoliensis in which fluoride-resistance was induced by prior growth in non-growth-inhibitory levels of NaF accumulated much less fluoride than did normal (“sensitive”) cells, and also did not respond to fluride by reduction of the ATP pool. It is suggested (1) that fluoride enters sensitive cells of S. leopoliensis principally as undissociated HF; (2) that its major inhibitory effect in these cells is the reduction in cellular ATP; (3)that fluoride-resistant cells accumulate less fluoride by developing incresed permeability to the fluoride anion.     

CHROMATIC REGULATION OF QUANTUM YIELDS FOR PHOTOSYSTEM II CHARGE SEPARATION,OXYGEN EVOLUTION,AND CARBON FIXATION IN HETEROCAPSA PYGMAEA (PYRROPHYTA)1

Operational and maximum quantum yields for system (PSII) charge separation, oxygen evolution, and carbon fixation were quantified and compared for Heterocapsa pygmaea Loeblich, Schmidt et Sherley populations chromatically adapted to white, green, blue, and red light. Significant variability in quantum yields was induced by chromatic adaptation alone or when chromatically adapted cells were suddenly exposed to biased light fields (i.e. white light). Results indicated a close coupling between the variability in quantum yields for PSII charge separation and oxygen evolution, but not between quantum yields for oxygen evolution and carbon fixation. But not between quantum yields for oxygen evolution and carbon fixation. The ability to regrlate and optimize light energy distribution between PSII and photosystem I (PSI) appears to be the mechanism underlying chromatic adaptation for PSII charge separation and oxygen evolution. Conceptually, the resulting impacts on PSI cyclic electron transport rates could account for observed variability in quantum yields for oxygen evolution and some variability in quantum yields for carbon fixation. Similarly, enzymatic processes associated with organic carbon synthesis appeared to be variably dependent on spectral growth irradiances and contributing to the observed variability in quantum yields for carbon fixation. The relevance of these findings to the in situ primary production is discussed.     

EFFECTS OF CADMIUM TOXICITY ON GROWTH AND ELEMENTAL COMPOSITION OF MARINE PHYTOPLANKTON

Some classes of marine phytoplankton are believed to be more tolerant of high concentrations of trace metals than others, but the results of experimental tests of this hypothesis are ambiguous. Eleven species of phytoplankton representing five classes were grown in Aquil medium containing Cd concentrations between 10⁻⁸ and 10⁻⁵ M ([Cd²⁺]= 10^−9.85 to 10^−6.84 M), and growth rates and intracellular concentrations of Cd, C, N, and S were measured. The mean Cd²⁺ concentration (pCd⁵⁰) that reduced the growth rate of each species to 50% of its maximum varied by 2.5 orders of magnitude, from 10^−6.23 for Emiliania huxleyi to 10^−8.79 for Synechococcus sp. Taxonomic trends in Cd resistance were not apparent in these data. Cadmium quotas (mol Cd·L⁻¹ cell volume) were lowest in species of Bacillariophyceae (ANOVA, P < 0.001), suggesting that they might regulate Cd transport differently than other taxa. Cellular S:C molar ratios increased in four of seven phytoplankton grown at high pCd (7.37–6.84) compared to low Cd ion concentrations (no added Cd), a result of increases in S·L⁻¹ cell volume. Nitrogen:carbon molar ratios were also higher in Cd-exposed phytoplankton, as changes in N and S were highly correlated ( r = 0.98, P < 0.0001). In two species that were examined, S:C ratios increased as a linear function of increasing Cd concentration. The results demonstrate large variability in Cd resistance among phytoplankton that is primarily a function of interspecific differences in Cd detoxification.     

NITROGEN LIMITATION INCREASES BREVETOXINS IN KARENIA BREVIS (DINOPHYCEAE): IMPLICATIONS FOR BLOOM TOXICITY1

Laboratory and field measurements of the toxin content in Karenia brevis cells vary by >4‐fold. These differences have been largely attributed to genotypic variations in toxin production among strains. We hypothesized that nutrient limitation of growth rate is equally or more important in controlling the toxicity of K. brevis, as has been documented for other toxic algae. To test this hypothesis, we measured cellular growth rate, chlorophyll a, cellular carbon and nitrogen, cell volume, and brevetoxins in four strains of K. brevis grown in nutrient‐replete and nitrogen (N)‐limited semi‐continuous cultures. N‐limitation resulted in reductions of chlorophyll a, growth rate, volume per cell and nirtogen:carbon (N:C) ratios as well as a two‐fold increase (1%–4% to 5%–9%) in the percentage of cellular carbon present as brevetoxins. The increase in cellular brevetoxin concentrations was consistent among genetically distinct strains. Normalizing brevetoxins to cellular volume instead of per cell eliminated much of the commonly reported toxin variability among strains. These results suggest that genetically linked differences in cellular volume may affect the toxin content of K. brevis cells as much or more than innate genotypic differences in cellular toxin content per unit of biomass. Our data suggest at least some of the >4‐fold difference in toxicity per cell reported from field studies can be explained by limitation by nitrogen or other nutrients and by differences in cell size. The observed increase in brevetoxins in nitrogen limited cells is consistent with the carbon:nutrient balance hypothesis for increases in toxins and other plant defenses under nutrient limitation.     

TOXICITY AND GROWTH-REGULATING ACTIVITY OF A NEEM SEED KERNEL EXTRACT (AZAL-S) TO THE LARVAE OF CULEX QUINQUEFASCIATUS

Abstract Toxicity of a neem seed kernel extract (AZAL-S) was assessed under laboratory conditions against larvae of Culex quinquefasciatus . The LC₅₀ value for AZAL-S was 0.78× 10–6. Exposure of 4th instar larvae to 0. 3× 10–6 and 0. 5× 10–6 AZAL-S for 120 hours caused 46. 2% and 67. 5% mortality respectively, with various types of morphogenetic aberration. AZAL-S induced an obvious prologation of the larval period when 1st instar larvae were treated with 0. 4–1. 0× 10–6 AZAL-S for 7 days. The insect growth regulating effect of AZAL-S on mosquito larvae was similar to those reported for certain insect growth regulators.     

INFLUENCE OF PHOSPHORUS LIMITATION ON TOXICITY AND PHOTOSYNTHESIS OF ALEXANDRIUM MINUTUM (DINOPHYCEAE) MONITORED BY IN‐LINE DETECTION OF VARIABLE CHLOROPHYLL FLUORESCENCE1

The effects of phosphorus (P) limitation on growth, toxicity, and variable chl fluorescence of Alexandrium minutum were examined in batch culture experiments. Cell division was greatly impaired in P‐limited cultures, but P spiking of these cultures after 9 days stimulated high levels of cell division equivalent to P‐replete cultures. The cellular concentration of paralytic shellfish toxins was consistent over the growth cycle of control cultures from lag phase into logarithmic growth phase, with toxins repeatedly lost to daughter cells during division. The low level of cell division in P‐limited cultures resulted in a 10‐fold increase of cellular toxin compared with controls, but this dropped upon P spiking due to increased rates of cell division. The history of phosphorus supply had an important effect on toxin concentration, with the P‐limited and the P‐spiked cultures showing values 2‐fold higher than the P‐replete cultures. Toxin profiles of the A. minutum strain used in these experiments were dominated by the N₁‐hydroxy toxins, gonyautoxins (GTX) GTX1 and GTX4, which were approximately 40 times more abundant than their analogues, GTX2 and GTX3, in P‐limited cultures. The dominance of the N₁‐hydroxy toxins increased significantly in control cultures as they advanced through logarithmic growth. In‐line measurements of the variable chl fluorescence of light‐adapted cells indicated consistent photochemical efficiency under P‐replete conditions. P limitation induced a drop in fluorescence‐based photochemical efficiency that was reversible by P spiking. There was an inverse linear relationship between in‐line fluorescence and cell toxin quota (r = ?0.88). Monitoring fluorescence in‐line may be valuable in managing efficient biotechnological production of toxins.     

印楝种核提取物AZAL-S对致倦库蚊幼虫的毒性和生长调节活性(英文)

在实验室条件下,测试了印楝种核提取物AZAL-S对致倦库蚊幼虫的毒力,其LC_(50)为0.78×10~(-6)。分别用0.3×10~(-6)和0.5×10~(-6)的AZAL-S处理致倦库蚊4龄幼虫120小时,死亡率为46.2％和67.5％,并出现各种畸形。用0.4×10~(-6)—1.0×10~(-6)的AZAL-S处理致倦库蚊1龄幼虫7天,发现其幼虫期显著延长。AZAL-S对蚊幼虫的生长调节作用与某些昆虫生长调节剂对蚊幼虫的作用相似。     

EFFECT OF CULTURE AND BLOOM DEVELOPMENT AND OF SAMPLE STORAGE ON PARALYTIC SHELLFISH POISONS IN THE CYANOBACTERIUM ANABAENA CIRCINALIS

Paralytic shellfish poisons (PSPs) were detected in 24 of 31 bloom samples dominated by the cyanobacterium Anabaena circinalis Rabenhorst, collected from across Austraia. The ability to produce PSPs has been maintained in everal non-axenic strains of A. circinalis kept in culture, whereas strains that were non-toxin-producing when isolated have remained as such. PSPs were detected and quantified by high-performance liquid chromatography (HPLC), and the structures were confirmed by electrospray mass spectroscopy. The concentration of toxins in PSP positive samples ranged from 50 to 3400 μg.g^-1 dry weight. Toxin profiles were always dominated by the N-sulfocarbamoyl-11-hydroxysulfate C toxins, C1 and C2 (44–85 mol%), with the remainder consisting of gonyautoxins-2, −3, and −5, decarbamoylgonyautoxins-2 and −3, saxitoxin, and decarbamoylraxitoxin. N₁-_-hydroxy PSPs, commonly found in marine dinoflagellates, were absent, suggesting that A. circinalis lacks the enzyme responsible for N₁-_-hydroxylation. On a dry weight basis, the amount of toxin in cultured Anabaena circinalis (strain ACMB06)rose significantly (P < 0.05)over time from 570 to 3400 μg.g.^-1 cells in late stationary phase. However, there was no significant trend in cellular toxin quota (toxin per cell) over the life of the culture; this may be explained by variation in cell mass. On average, batch cultures of Anabaena circinalis contained 19% extracellular toxin, which increased slightly over the growth cycle and had a composition similar to that of the intracellular toxins. As cultures aged, the formation of decarbamoyl toxins and increases in theα-/β-epimer ratios of C toxins and gonyautoxins were observed. The variation in these components during stationary phase in culture was sufficient to explain the variation in relative PSP composition observed among natural bloom samples. Because decarbamoylgonyautoxins are much more toxic than C toxins on a molar basis, these transformations also lead to an increase in toxicity of the sample or bloom over time. The transformations of PSPs, which occur during aging and sample storage, render the comparison of PSPs by HPLC unreliable for phenotyping Anabaena circinalis, unless strains are cultured, harvested, and analyzed under standard conditions.     

BIOCHEMICAL COMPOSITION AND METABOLIC ACTIVITY OF SCRIPPSIELLA TROCHOIDEA (DINOPHYCEAE) RESTING CYSTS1

The composition and metabolic activity of cysts of the marine dinoflagellate Scrippsiella trochoidea (Stein) Loeblich were examined during dormancy, quiescence, and germination. On a per cell basis, newly formed cysts contained an order of magnitude more carbohydrate but significantly less protein and chlorophyll a than did exponentially growing vegetative cells. Loss of lipid and carbohydrate from cysts during the initial dormancy period reflected a respiration rate estimated to be 10% of the respiratory activity in vegetative cells. Among older, quiescent cysts the calculated respiration rate decreased further to approximately 1.5% of the vegetative rate and appeared to proceed largely at the expense of carbohydrate reserves. These estimated rates of respiration were in good agreement with direct measurements of cyst oxygen consumption. The transfer of quiescent cysts to conditions permissive for germination resulted in a rapid increase in respiration rate, as evidenced by carbohydrate loss and O₂ consumption. The increased respiratory activity was followed by an increase in protein content and, later, by an increase in chlorophyll a content and photosynthetic capacity. Just prior to germination the P/R ratio became greater than 1, and the estimated chlorophyll-specific photosynthetic activity reached 75% of the rate in vegetative cells. Complete restoration of photosynthetic and respiratory capacity apparently was not achieved until after excystment. These data confirm the common assumption that dinoflagellate cysts represent true “resting” cells, containing extensive energy reserves and displaying greatly reduced metabolic activity.     

KLEPTOPLASTIDY IN THE TOXIC DINOFLAGELLATE PFIESTERIA PISCICIDA (DINOPHYCEAE)

The ichthyotoxic dinoflagellate Pfiesteria piscicida Steidinger et Burkholder has a complex life cycle with several heterotrophic flagellated and amoeboid stages. A prevalent flagellated form, the nontoxic zoospore stage, has a proficient grazing ability, especially on cryptophyte prey. Although P. piscicida zoospores lack the genetic capability to synthesize chloroplasts, they can obtain functional chloroplasts from algal prey (i.e. kleptoplastidy), as demonstrated here with a cryptophyte prey. Zoospores grown with Rhodomonas sp. Karsten CCMP757 (Cryptophyceae) grazed the cryptophyte population to minimal densities. After placing the cultures in near darkness where cryptophyte recovery was restricted and further prey ingestion did not occur, the time-course patterns in growth, prey chloroplast content·zoospore⁻¹, and prey nucleus content·zoospore⁻¹ were followed. Ingested chloroplasts were selectively retained in the dinoflagellate, as indicated by the decline and, ultimately, near absence of cryptophyte nuclei in plastid-containing zoospores. Chloroplasts retained inside P. piscicida cells for at least a week were photosynthetically active, as indicated by starch accumulation and microscope-autoradiographic measurements of bicarbonate uptake. Recognition that P. piscicida can function as a phototroph broadens our perspective of the physiological ecology of the dinoflagellate because it suggests that, at least during part of its life cycle, P. piscicida 's growth and survival might be affected by photoregulation and nutritional control of photosynthesis.     

ENVIRONMENTAL MODULATION OF KARLOTOXIN LEVELS IN STRAINS OF THE COSMOPOLITAN DINOFLAGELLATE,KARLODINIUM VENEFICUM (DINOPHYCEAE)1

We examined the influence of N or P depletion, alternate N‐ or P‐sources, salinity, and temperature on karlotoxin (KmTx) production in strains of Karlodinium veneficum (D. Ballant.) J. Larsen, an ichthyotoxic dinoflagellate that shows a high degree of variability of toxicity in situ. The six strains examined represented KmTx 1 (CCMP 1974, MD 2) and KmTx 2 (CCMP 2064, CCMP 2283, MBM1) producers, and one strain that did not produce detectable karlotoxin under nutrient‐replete growth conditions (MD 5). We hypothesized that growth‐limiting conditions would result in higher cell quotas of karlotoxin. KmTx was present in toxic strains during all growth phases and increased in stationary and senescent phase cultures under low N or P, generally 2‐ to 5‐fold but with some observations in the 10‐ to 15‐fold range. No karlotoxin was observed under low‐N or low‐P conditions in the nontoxic strain MD 5. Nutrient‐quality (NO₃, NH₄, urea, and glycerophosphate) did not affect growth rate, but growth on NH₄ produced 2‐ to 3‐fold higher cellular toxicity and a 50% higher ratio of KmTx 1‐1:KmTx 1‐3 in CCMP 1974. CCMP 1974 showed higher cellular toxicity at low salinity (≤5 ppt) and high temperature (25°C). Our results suggested that given the presence of a toxic strain of K. veneficum in situ, the existence of environmental conditions that favor cellular accumulation of karlotoxin is likely a significant factor underlying K. veneficum–related fish kills that require both high cell densities (10⁴ · mL^?1) and high cellular toxin quotas relative to those generally observed in nutrient‐replete cultures.