Toxicity of benthic dinoflagellates on grazing, behavior and survival of the brine shrimp Artemia salina

Raquel A. F. Neves; Tainá Fernandes; Luciano Neves dos Santos; Silvia M. Nascimento

doi:10.1371/journal.pone.0175168

Abstract

Harmful algae may differently affect their primary grazers, causing sub-lethal effects and/or leading to their death. The present study aim to compare the effects of three toxic benthic dinoflagellates on clearance and grazing rates, behavioral changes, and survival of Artemia salina. Feeding assays consisted in 1-h incubations of brine shrimps with the toxic Prorocentrum lima, Gambierdiscus excentricus and Ostreopsis cf. ovata and the non-toxic Tetraselmis sp. Brine shrimps fed unselectively on all toxic and non-toxic algal preys, without significant differences in clearance and ingestion rates. Acute toxicity assays were performed with dinoflagellate cells in two growth phases during 7-h to assess differences in cell toxicity to A. salina. Additionally, exposure to cell-free medium was performed to evaluate its effects on A. salina survival. The behavior of brine shrimps significantly changed during exposure to the toxic dinoflagellates, becoming immobile at the bottom by the end of the trials. Dinoflagellates significantly affected A. salina survival with 100% mortality after 7-h exposure to cells in exponential phase (all treatments) and to P. lima in stationary phase. Mortality rates of brine shrimps exposed to O. cf. ovata and G. excentricus in stationary phase were 91% and 75%, respectively. However, incubations of the brine shrimps with cell-free medium did not affect A. salina survivorship. Significant differences in toxic effects between cell growth phases were only found in the survival rates of A. salina exposed to G. excentricus. Acute exposure to benthic toxic dinoflagellates induced harmful effects on behavior and survival of A. salina. Negative effects related to the toxicity of benthic dinoflagellates are thus expected on their primary grazers making them more vulnerable to predation and vectors of toxins through the marine food webs.

Citation: Neves RAF, Fernandes T, Santos LNd, Nascimento SM (2017) Toxicity of benthic dinoflagellates on grazing, behavior and survival of the brine shrimp Artemia salina. PLoS ONE 12(4): e0175168. https://doi.org/10.1371/journal.pone.0175168

Editor: Elena Gorokhova, Stockholm University, SWEDEN

Received: October 3, 2016; Accepted: March 21, 2017; Published: April 7, 2017

Copyright: © 2017 Neves et al. This is an open access article distributed under the terms of the Creative Commons Attribution License, which permits unrestricted use, distribution, and reproduction in any medium, provided the original author and source are credited.

Data Availability: All relevant data are included within the paper and its Supporting Information files.

Funding: Partial funding was provided by the Graduate Course in Neotropical Biodiversity (PPGBIO-UNIRIO) and Fundação Carlos Chagas Filho de Amparo à Pesquisa do Estado do Rio de Janeiro – FAPERJ (grant nº E-26/203.493/2015). The funders had no role in study design, data collection and analysis, decision to publish, or preparation of the manuscript.

Competing interests: The authors have declared that no competing interests exist.

Introduction

Marine benthic dinoflagellates are important primary producers and most of their representatives are potentially toxic [1]. Recently, this group of organisms has received significant scientific attention since the occurrence of benthic harmful algal blooms (HAB) has increased worldwide [1]. Among benthic dinoflagellates, blooms of Ostreopsis cf. ovata have been recorded with increasing frequency, intensity and distribution, particularly in the Mediterranean Sea [2–3], with adverse consequences on benthic communities and human intoxication, mainly through the inhalation of marine aerosols [4]. In Brazil, blooms of O. cf. ovata became a recurrent event since 1998 in Rio de Janeiro state [5] with high mortalities of the sea urchin Echinometra lucunter reported during these HAB events [6]. Similar ecological effects were observed in New Zealand, where blooms of O. siamensis caused decline in the numbers of the sea urchin Evechinus chloroticus [7]. Ostreopsis cf. ovata produces ovatoxins, and other PLTX analogues, one of the most toxic molecules occurring in nature that cause intoxication in humans [8]. There is evidence of PLTX and its analogues presence in crustaceans, molluscs and fish that, when contaminated, can cause the clupeotoxism disease by the consumption of sardines and anchovies (clupeoid fish) [4]. Moreover, a cytotoxic non-palytoxin derivative recently isolated from O. cf. ovata, ostreol A, was shown to have in vitro cytotoxicity against the brine shrimp Artemia salina [9].

Prorocentrum lima is a cosmopolitan species and often constitutes a significant part of benthic dinoflagellate communities worldwide [10]. This dinoflagellate produces okadaic acid (OA) and dinophysistoxins (DTXs), the main toxins responsible for diarrheic shellfish poisoning [11]. Evidence shows that every culture of P. lima tested to date has been found to produce OA and its analogues in varying quantities [12]. In Rio de Janeiro, P. lima is found all year round growing epiphytically on macroalgae and locally isolated strains demonstrated the production of high OA concentrations [13]. There is some evidence that P. lima can act as a vector for DSP toxins in shellfish and suspected cases of intoxication have been recorded in Argentina [14], Canada [15], the United Kingdom [16] and Japan [17].

Gambierdiscus excentricus produces maitotoxins (MTXs) and ciguatoxins (CTX) responsible for ciguatera fish poisoning [18], a disease caused by the consumption of herbivorous and carnivorous fish that have accumulated CTX through the food web. It is estimated that 50.000 to 500.000 people are affected by CTX every year, and ciguatera is the most frequently reported non-bacterial illness associated with seafood consumption worldwide [19]. Moreover, reef disturbance by hurricanes, military and tourist developments, as well as coral bleaching and the rise in water temperatures are increasing the risk of ciguatera by freeing up space for the growth of macroalgae in which Gambierdiscus colonize upon [20].

Despite some grazers may avoid certain toxic algae [21–22], most benthic species feed indiscriminately on toxic cells. Effects of algal toxins on their primary grazers are variable [23]. Zooplankton species may be affected by toxins showing sub-lethal symptoms such as changes in pulsation frequency and immobility [24], grazing and motility suppression [25], and even lethal responses [26–27]. Moreover, HAB events may lead to shifts in community composition to more resistant species, generating complex cascading effects through the pelagic and benthic food webs [28]. Contaminated individuals may also transfer phycotoxins through the marine trophic web by the direct predation [29–30], by the elimination of toxic cells in biodeposits and feces and through the release of toxins after their death making toxins available for detritivorous species [31–33].

Most information on interactions between toxic algae and their grazers comprises phytoplankton and zooplankton grazers, particularly copepods [23, 34–35]. Differential survival rates of toxin-exposed individuals may indicate some degree of toxin resistance by individuals selected in natural populations in the long-term [36, 37]. From an ecological point of view, toxin-resistant zooplankton fed harmful alga are more hazardous than sensitive individuals by its higher capacity for toxin accumulation which favors toxin transfer to higher consumers and dispersion of toxins through marine environments. Thus, the interactions between benthic harmful algae and their grazers must be better investigated in order to assess the potential ecological impacts of phycotoxins on grazers and the toxins distribution through marine food webs.

The brine shrimp Artemia salina is widely used for toxicity tests due to its widespread distribution, short life cycle, non-selective grazing, and sensitivity to toxic substances [38]. Therefore, A. salina seems to be a suitable model species to assess the toxicity of marine benthic dinoflagellates. The purpose of the present study was to evaluate the effects of the benthic dinoflagellates P. lima, G. excentricus, and O. cf. ovata on clearance and grazing rates, behavioral changes, and survival of A. salina during acute exposure. Acute toxicity assays were performed with cells in different growth phases of a batch culture (exponential and stationary) to assess differences in cell toxicity to brine shrimps.

Material and methods

Algae cultures

Clonal cultures of Prorocentrum lima (strain UNR-01), Ostreopsis cf. ovata (strain UNR-05) and Gambierdiscus excentricus (strain UNR-08) used in this study were isolated from Armação dos Búzios (22°45ʹ18ʹʹ S, 41°54ʹ07ʹʹ W), Rio de Janeiro state as described in [5, 11, 39]. A non-toxic strain of the chlorophyte Tetraselmis sp. was isolated from Guanabara Bay, Rio de Janeiro state (22°15’-23°05’ S, 43°30’-42°30’ W). Scientific research and collecting permit authorizing field studies were obtained from Instituto Chico Mendes de Conservação da Biodiversidade (ICMBio), Brazilian Ministry of the Environment (permit number: 35192–3). No protected or endangered species was caught through field studies or used in experimental trials.

Benthic dinoflagellates were maintained in filtered seawater (glass-fiber filter, Millipore AP-40, Millipore Brazil) supplemented with L2 enrichment medium [40], modified by omitting silicate, nickel, vanadium and chromium; except for O. cf. ovata which was grown in L2/2 medium. Salinity was adjusted to 34 for all the cultures, except for G. excentricus that was cultivated at salinity 32. All stock cultures were kept in a temperature-controlled cabinet at 24 ± 2°C, with a 12:12 h dark-light cycle and photon flux density of 60 μmol m^-2s^-1 provided by cool-white fluorescent tubes. Photosynthetically active radiation was measured with a QSL-100 quantum sensor (Biospherical Instruments, San Diego, CA, USA).

The strain of Prorocentrum lima (UNR-01) used in the current study synthesizes mostly OA and small amounts of DTX-1 [13]. The Gambierdiscus excentricus strain (UNR-08) produces at least one MTX analog, while the evaluation of CTX production by this strain has not yet been completed (P Hess personal communication). The strain of Ostreopsis cf. ovata (UNR-05) has not been analyzed for toxin production. However, other strains of O. cf. ovata isolated from the same sampling location produce PLTX analogues (ovatoxins) [5].

Lugol preserved cells from the cultured strains were observed using light microscopy (Primovert, Zeiss, Germany) to measure the cells for the determination of cellular biovolume. Images were collected using an Axiocam Icc1digital camera (Zeiss, Germany), and cells (n = 40) were measured using the Axiovision software (Zeiss, Germany). Cellular biovolume was estimated using geometric shapes and mathematical equations suggested for each genus [41]: prolate spheroid for the chlorophyte and ellipsoid for the dinoflagellates. Cellular carbon content was calculated according to equations for dinoflagellates and chlorophytes [42].

Experimental design

Feeding assays.

Adult individuals of A. salina (7 mm ± 1.14) were acclimatized in experimental conditions in a temperature-controlled cabinet at 24 ± 2°C for 24 to 48 h and fed ad libitum with Tetraselmis sp. Feeding trials were performed in 6-well plates containing, each well, three individuals in 10 ml of filtered seawater (glass-fiber filter, Millipore AP-40, Millipore Brazil) at salinity 35.

Previous tests of A. salina grazing were carried out at various abundances of the non-toxic prey Tetraselmis sp. (150–720 cells ml^-1) during an incubation interval that ranged from 30 min to 6 h. The results indicated the optimum incubation time (1 h) and Tetraselmis sp. abundance (~ 481 cells ml^-1; 40 ng C ml^-1) for feeding experiments in which the brine shrimps depleted 15–30% of cell abundances. Cell size of microalgae used in this study ranged from 13 μm (Tetraselmis sp.) to 78 μm (G. excentricus). Thus, the same carbon concentration was established in all treatments allowing the comparison of clearance and grazing rates of A. salina among different prey species. Cell abundances of microalgae that corresponded to the carbon concentration of 40 ng C ml^-1, per well, were: Tetraselmis sp. (0.09 ng C cell^-1, ~ 481 cells ml^-1), Prorocentrum lima (2.15 ng C cell^-1, ~ 19 cells ml^-1), Ostreopsis cf. ovata (3.21 ng C cell^-1, ~ 13 cells ml^-1), and Gambierdiscus excentricus (10.95 ng C cell^-1, ~ 4 cells ml^-1). Microalgae species were harvested during exponential phase for feeding trials.

Each treatment containing the brine shrimps and one algal species was performed in triplicate, and the control containing solely algal cells without the brine shrimps was performed in duplicate. Aliquots of each replicate (1 ml) were collected using an automatic pipette at the beginning and after 1-h incubation. Cells were preserved with neutral Lugol's iodine solution for later cell counts using a Sedgewick-rafter chamber and observation in an inverted microscope (Primovert, Zeiss, Germany).

Artemia salina clearance (ml ind^-1h^-1; CR) and ingestion (ng C ind^-1h^-1; IR) rates were calculated by the ratio of natural log (ln) of initial and final carbon concentrations after 1-h incubation, and corrected by the carbon concentration in controls [43]. Clearance and ingestion rates were calculated for each replicate and, posteriorly, mean rates were calculated by treatment.

Acute toxicity assays.

Adult individuals of A. salina (7 ± 1.14 mm) were acclimatized in the same experimental conditions described above for feeding experiments. Intoxication assays were performed in 6-well plates containing, each well, 10 ml of filtered seawater (glass-fiber filter, Millipore AP-40, Millipore Brazil) at salinity 35. Three individuals of Artemia salina were incubated for 7-h with each dinoflagellate species (G. excentricus or O. cf. ovata or P. lima) in abundances of 200 cells ml^-1 [44]. Acute exposure of A. salina individuals to each toxic species was performed twice on independent samples, first with cells at exponential phase and in the second time with cells at stationary phase.

Dinoflagellate growth curves were previously determined [5, 11], and the non-harmful species Tetraselmis sp. (control) was kept and harvested in exponential growth for all trials. Toxic treatments were performed in triplicates and the non-toxic control in duplicate. In total, toxic treatments consisted of six different incubations: 1) P. lima in exponential phase, 2) P. lima in stationary phase, 3) O. cf. ovata in exponential phase, 4) O. cf. ovata in stationary phase, 5) G. excentricus in exponential phase, and 6) G. excentricus in stationary phase. Changes on swimming activity (actively swimming or motionless), position (individuals on the water column or at the bottom) and survival of individuals were verified after the first 30 min and after every hour of incubation.

Additional experiments were carried out to evaluate the effects of cell-free medium on brine shrimps survival. Cell-free medium were obtained by the filtration (glass-fiber filter, Millipore AP-40, Millipore Brazil) of G. excentricus, O. cf. ovata and P. lima cultures in abundances of 200 cells ml^-1. Filtrates were used immediately. Incubations were performed in 6-well plates containing, each well, 10 ml of cell-free medium with six individuals of A. salina during 7-h. Each treatment of cell-free medium was performed in six replicates and the control (only filtered seawater) was performed in triplicate.

Statistical analyses

One-way ANOVA was applied to evaluate the influence of four different preys on the ingestion and clearance rates of brine shrimps—categorical factor (P. lima, O. cf. ovata, G. excentricus, and Tetraselmis sp.). Normality and homogeneity of variances were assessed using Kolmogorov-Smirnov and Levene Test, respectively, and log-transformation was applied when necessary. Analyses were performed using the software Statistica 8.0 (StatSoft).

A redundancy analysis (RDA) was applied to data of A. salina individuals from toxic and non-toxic treatments using survival, position and swimming as explanatory variables, and exposure time (7 h) as covariable. A Monte Carlo permutation was used to test the significance of ordination model. The multivariate analysis was performed in the software CANOCO [45].

Generalized linear models (GLM) were applied to survival, swimming and position data (as dependent variables) of A. salina individuals from toxic treatments (as categorical factor) with exposure time (7-h) as continuous predictor in a full factorial design. Tukey test was applied whenever significant differences were detected by GLMs. A Kruskal-Wallis test was applied to survival data of A. salina individuals exposed to cell-free medium of dinoflagellate cultures. Analyses were performed using the software Statistica 8.0 (StatSoft).

Survival functions were estimated from the continuous survival time for 7-h brine shrimps exposure to the toxic dinoflagellates. The time when 50% of the individuals died (tD₅₀) in each treatment was also estimated. A Kaplan-Meier log rank test (Mantel-Cox) was applied to test significant differences in the survivorship curve of A. salina among toxic treatments using the software GraphPad Prism 5.0 (GraphPad Software, San Diego—California). The Kaplan-Meier test takes into account the censored survivorship data [46].

Results

Feeding assays

The brine shrimps Artemia salina actively fed on all different preys offered to them (Fig 1), including the toxic species.

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Fig 1. Toxic dinoflagellates in the digestive tract of Artemia salina individuals.

Dinoflagellate cells are indicated by arrows (A) Prorocentrum lima, (B) Ostreopsis cf. ovata, and (C) Gambierdiscus excentricus. Scale bars: 200 μm.

https://doi.org/10.1371/journal.pone.0175168.g001

There was no significant difference in clearance (One-way ANOVA, F = 1.27, df = 3, p = 0.32) and ingestion rates (One-way ANOVA, F = 2.09, df = 3, p = 0.14) of the brine shrimps among the different treatments (Table 1). Adults of A. salina did not exhibit any unusual behavior when exposed to the toxic dinoflagellates during feeding assay.

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Table 1. Artemia salina clearance and ingestion rates.

https://doi.org/10.1371/journal.pone.0175168.t001

Acute toxicity assays

Redundancy analysis (RDA) applied to survival and behavioral data of A. salina clearly distinguished toxic treatments (negative coordinates on the first axis) from the non-toxic one (positive coordinates on the first axis; Fig 2). The canonical axes were statistically significant (p = 0.002); the first axis explained 98.5% total variance, while the second axis explained 1% total variance. The positive direction of the vectors swimming activity, water column position and survival confirms its positive correlation with samples from the non-toxic treatment (Tetraselmis sp.).

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Fig 2. Survival and behavioral responses of Artemia salina exposed to toxic and non-toxic treatments.

Biplot of survival and behavioral data of A. salina exposed to the non-toxic chlorophyte Tetraselmis sp. (Tet(C),●) and toxic dinoflagellates: Prorocentrum lima in exponential phase (Pli(E),▲), P. lima in stationary phase (Pli(S),■), Ostreopsis cf. ovata in exponential phase (Oov(E),▲), O. cf. ovata in stationary phase (Oov(S),■), Gambierdiscus excentricus in exponential phase (Gex(E), ▲), and G. excentricus in stationary phase (Gex(S), ■).

https://doi.org/10.1371/journal.pone.0175168.g002

The brine shrimp A. salina exposed to the toxic dinoflagellates P. lima, O. cf. ovata and G. excentricus showed abnormal behavior related to swimming activity and its position on the water column, independently of dinoflagellate growth phase (Fig 3). Artemia salina behavior was significantly affected by the time of exposure to toxic dinoflagellates (GLM, F_{swimming =} 370.6, F_position = 451.2, p<0.001), leading to the immobility of individuals at the bottom. Behavioral changes in the swimming activity and the position of the brine shrimps were noticed in the first 30-min incubations with dinoflagellates in stationary phase. After 1-h exposure to toxic cells, either in exponential or stationary phase, individuals have shown changes in swimming activity and in their position on the water column (Fig 3).

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Fig 3. Behavioral changes in Artemia salina during acute exposure to toxic dinoflagellates.

(A-B) Brine shrimps (%) actively swimming during exposure to dinoflagellate cells in exponential (A) and stationary (B) phase. (C-D) Brine shrimps (%) on the water column (in contrast to bottom position) during exposure to dinoflagellate cells in exponential (C) and stationary (D) phase. Different treatments are indicated by symbols: P. lima (•, full line), G. excentricus (○, dashed line), O. cf. ovata (▪, full line), and the chlorophyte Tetraselmis sp. (□, full line). The non-harmful chlorophyte used as control was kept and harvested in exponential phase for all the assays. Vertical bars: standard error considering four replicates.

https://doi.org/10.1371/journal.pone.0175168.g003

The swimming activity of A. salina was significantly affected by dinoflagellate species (GLM, F = 3.09, p = 0.01) and by the interaction between dinoflagellate species and time of exposure (GLM, F ₌ 4.9, p< 0.001). The A. salina activity was overall more affected by the exposure to P. lima and O. cf. ovata than to G. excentricus (Fig 3). No significant effect of dinoflagellate species (GLM, F = 1.36, p = 0.24) nor interaction between dinoflagellate and exposure time (GLM, F ₌ 0.64, p = 0.67) was detected on A. salina position on the water column.

The survival of A. salina was significantly affected by dinoflagellate species (GLM, F = 3.75, p = 0.003), by the exposure time (GLM, F = 430.74, p< 0.001), and by the interaction between dinoflagellate and exposure time (GLM, F = 4.41, p<0.001). The incubation of the brine shrimps with cell-free medium did not affect A. salina survivorship. There was no significant difference among treatments and control (Kruskal-Wallis, p = 0.23), survival rates ranged from ~92–100%: control (100%, ±0), and cell-free medium of G. excentricus (97.2%, ±6.8), O. cf. ovata (94.4%, ±8.6), and P. lima (91.7%, ±9.1). In contrast, the exposure to all the toxic dinoflagellate cells in exponential phase severely affected A. salina survival, with 100% mortality during assays (Fig 4). Survival rates of A. salina during exposure to G. excentricus and O. cf. ovata in stationary phase were 25% and 9%, respectively, after 7-h incubations. Only the exposure to P. lima, in both growth phases, lead to the death of all individuals within 6-h incubations (100% mortality, Fig 4).

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Fig 4. Survival rate curves of Artemia salina during acute exposure to the toxic benthic dinoflagellates in two growth phases: exponential (•) and stationary (□).

(A) Prorocentrum lima, (B) Ostreopsis cf. ovata, and (C) Gambierdiscus excentricus.

https://doi.org/10.1371/journal.pone.0175168.g004

Significant differences in the survival rates of A. salina were found for two pairwise comparisons using different species of dinoflagellate: (1) P. lima in stationary phase x G. excentricus in exponential phase (Tukey post-hoc test, p< 0.001), (2) P. lima in stationary phase x O. cf. ovata in stationary phase (Tukey post-hoc test, p = 0.016). During exposure to P. lima in stationary phase, less than 25% of A. salina individuals died in the first 2-h and, after a sharp decline in survival, all individuals were dead within 6-h incubation. In contrast, survival rate of A. salina exposed to G. excentricus cells in exponential phase slightly declined with over 30% survival after 6-h exposure. During exposure to O. cf. ovata cells in stationary phase, there was a sharp decline in A. salina survival in the first hours of incubation followed by a trend to stabilization close to 10% survival in the last hour (Fig 4). Significant differences related to the effect of growth phases on A. salina survival have been found only for Gambierdiscus excentricus (Tukey post-hoc test, p = 0.028). The survival of brine shrimps exposed to G. excentricus cells in exponential phase showed a steady decline until the end of incubation, reaching 100% mortality. In contrast, the exposure to G. excentricus in stationary phase led to a sharp reduction in survival within the first 3-h, with a loss of 25% of individuals per hour, followed by a stabilization in survival rate until the end of incubation with ~25% survival (Fig 4).

Two-hour incubation was the shortest time in which 50% of A. salina individuals have died (lower tD₅₀) during exposure to Gambierdiscus excentricus in stationary phase (Table 2). Despite the quick effect of G. excentricus cells in stationary phase on brine shrimp survival, the highest survival rate (25%) of A. salina was found in the same treatment after 7-h exposure (Fig 4). The highest tD₅₀ value was noticed for brine shrimps exposed to G. excentricus in exponential phase (4 h, Table 2). Intermediary and similar tD₅₀ values were found in P. lima and O. cf. ovata treatments (Table 2).

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Table 2. Time (h) when 50% of the Artemia salina died (tD₅₀) during exposure to toxic dinoflagellates in two cell growth phases.

https://doi.org/10.1371/journal.pone.0175168.t002

Discussion

The four microalgae offered as prey have different morphological and toxicological characteristics that would account for differences in clearance and ingestion rates of potential grazers. However, the brine shrimp Artemia salina actively fed on the different preys offered to them: three species of toxic benthic dinoflagellates and one non-harmful chlorophyte. No significant differences in clearance and ingestion rates were noticed among the four treatments. Thus, clearance and ingestion rates of brine shrimps seem to be unaffected by differences in cell size in the range 13–78 μm. In a previous study, clearance rates of A. salina on the dinoflagellate Cochlodinium polykrikoides were three-to-four times faster than clearance rates on the cryptophyte Rhodomonas salina suggesting preferences for prey in the range 2–35 μm and for dinoflagellate as prey [47].

In addition, neither preference nor avoidance of toxic prey has been shown by the brine shrimps. The clearance and ingestion rates of brine shrimps were not affected by the toxicity of the dinoflagellates during the short-term experiment of 1-h incubation. This is probably due to the low cell abundances offered to them ~4–19 cells ml^-1. Similarly, no preference has been shown by the copepods Acartia tonsa and Temora longicornis with similar ingestion rates on diatoms, the toxic Pseudo-nitzchia multiseries and the non-toxic P. pungens [48]. In the same way, A. salina grazed on the paralytic shellfish toxins (PSTs) producer Alexandrium fundyense at rates similar to that displayed for the non-harmful cryptophyte Rhodomonas salina [47]. However, brine shrimp feeding rates on A. fundyense decreased when dinoflagellate abundances increased, while an opposite trend was observed in feeding rates on the non-harmful species [47]. Lower feeding rates of A. salina were observed when grazing on toxic Alexandrium species (A. catenella, A. minutum and A. tamarense) compared to grazing on a mixture of toxic and non-toxic dinoflagellates (A. tamarense + Prorocentrum donghaiense), and solely on the non-toxic P. donghaiense [49].

The threat of natural marine toxins increases considerable when bioaccumulation is considered along the food chains [50]. As the brine shrimps grazed on the harmful dinoflagellates, ingested cells that remained in A. salina tract can be transferred to secondary consumers as demonstrated for Gambierdiscus cells [31]. The toxic compounds may also be incorporated and accumulated into individuals, as previously described for A. salina exposed to PSTs [51] and microcystin producers [52]. Phycotoxins retained into individuals may be transformed by detoxification processes, such as the glutathione S-transferase described in A. salina, responsible for detoxification of various toxic compounds [53]. Crustaceans are key prey for marine species of higher trophic levels and perform daily and seasonal migration; therefore crustaceans that graze on toxic dinoflagellates (as Artemia salina shown in the present study) can act as vectors of dinoflagellate toxins in marine food webs [54].

The current intoxication trials contribute to increase the knowledge about direct toxic effects of benthic dinoflagellates on a marine invertebrate widely used as model organism in bioassays—Artemia salina [55]. The toxic Prorocentrum lima, Ostreopsis cf. ovata and Gambierdiscus excentricus isolated from tropical marine systems caused harmful effects on behavior and affected the survival of A. salina at abundances of 200 cells ml^-1. The harmful effects on behavior and survival of the brine shrimps were demonstrated by multivariate analysis (RDA) in which toxic treatments were clearly distinguished from the non-toxic control that was positively correlated with swimming activity, water column position and survival of A. salina.

Immobility and mortality were used as endpoints to measure the toxicity of the three benthic dinoflagellates to adults of A. salina. Healthy Artemia individuals are active swimmers, thus, alterations in swimming activity are valid as behavioral endpoints to detect stress at sub-lethal concentrations of toxic compounds [56]. The brine shrimps exposed to the toxic dinoflagellates exhibited reduced swimming activity, ultimately immobility. Swimming behavior has a direct impact on zooplankton dispersal, encounter with prey and predators, and vulnerability to predation which determine the propensity of individuals to graze on harmful species and transfer phycotoxins through the food web [36]. Only ~20% of A. salina individuals exposed to G. excentricus cells in stationary phase showed any movement after 7-h incubation.

Besides, brine shrimp exposure to O. cf. ovata and P. lima induced the loss of movement or death of all the individuals until the end of incubation. Similarly, adult females of the copepod Tigriopus japonicus exposed to Fukuyoa sp. (as Gambierdiscus cf. yasumotoi) showed a decrease in activity, loss of motor control, and abnormal swimming [57]. However, female copepods reached 100% immobility only after 12-d exposure to 200 cells ml^-1 of Fukuyoa sp. (as Gambierdiscus cf. yasumotoi) [57], the same abundance used in acute toxicity assays of the present study. Abnormal behavior exhibited by the crustaceans exposed to toxic dinoflagellates increases their vulnerability to predation [58–59]. In addition, a reduction in locomotion activity may affect the vertical migration of nauplii [60].

The present results showed that the toxic dinoflagellates P. lima and O. cf. ovata were able to induce similar effects on behavior, particularly swimming activity, and survival of the brine shrimp A. salina. The exposure to these two dinoflagellates caused a sharp reduction on swimming activity followed by an increase in mortality of A. salina adults. The OA and DTX-1 toxins produced by P. lima probably lead to physiological disturbances in brine shrimps related to loss of body fluids and lack of physiological control of fluid dynamics, as previously observed in affected organisms [44, 61]. Similarly, PLTX analogues potentially produced by O. cf. ovata alter the mechanisms of ion homeostasis [62], with disruption of cell membrane functions and loss of ion regulation [38]. In the present study, the adverse effects on A. salina induced by the exposure to P. lima and O. cf. ovata seem to be thus related to brine shrimp ion regulation.

The toxic benthic dinoflagellates significantly affected the survival rates of A. salina during acute exposure. All the brine shrimps exposed to the three toxic dinoflagellates in exponential phase died, while the mortality of brine shrimps exposed to cells in stationary phase varied according to dinoflagellate species: 100% (P. lima), 91% (O. cf. ovata), and 75% (G. excentricus). However, the cell-free medium did not significantly affect A. salina survival during acute exposure. Cell-free medium from cultures with higher abundances of O. cf. ovata (~4000 cells ml^-1) [38] were harmful to nauplii of A. salina. In the present study, the high survival of the brine shrimps exposed to cell-free medium of G. excentricus, O. cf. ovata and P. lima confirms that the concentration of diluted cell exudates was within A. salina tolerance and did not compromise their survival.

Artemia salina exhibited similar sensitivity to Prorocentrum lima and Ostreopsis cf. ovata acute exposure; the same pattern seen for behavioral responses. Adults of A. salina showed high sensitivity to P. lima either in exponential or stationary growth phases (respectively, tD₅₀ = 3.1 and 3.5h) and to O. cf. ovata, particularly in exponential growth phase (tD₅₀ = 3h), with 100% mortality after 6-h exposure. Conversely, minimal mortalities of A. salina adults have been reported after exposure to P. lima and P. concavum in abundances up to 1000 cells individual^-1[31]. Previous studies on A. salina larvae have demonstrated high sensitivity of nauplii to both P. lima (tD₅₀ = 1.7h) [44] and O. cf. ovata cells [38, 63]. In addition, A. salina nauplii were reported to be extremely sensitive to O. cf. ovata cells with half maximum effective concentration (EC₅₀) ranging from 6 to 24 cells ml^-1 [63]. Exposure of different crustaceans to O. cf. ovata cells revealed that A. salina larvae have the highest sensitivity among them, with a half lethal concentration (LC₅₀) lower than 4 cells ml^-1 [38]. Artemia salina has also showed high sensitivity to other Ostreopsis species, the PTX-like producer O. siamensis [64]. The sensitivity exhibited by A. salina in the two stages of its life cycle (nauplius and adult) to these toxic dinoflagellates confirms this crustacean as a suitable model species to assess the toxicity of marine harmful algae.

The dinoflagellate Gambierdiscus excentricus rapidly affected the swimming activity, water column position and survival of exposed-individuals of A. salina in the present assays. A short stabilization of effects was observed after 3-h exposure, followed by a reduction in the percentage of individuals swimming on the water column as well as in the survival of the brine shrimps between 5 to 7-h exposure. Maitotoxins (MTX) alter ion transport systems causing an increase in free intracellular Ca²⁺ [65] while ciguatoxins (CTXs) are neurotoxins that bind to the voltage-sensitive sodium channels on cell membranes [66–67]. The toxic effects of ciguatoxins seem to be similar to saxitoxins effects, the neurotoxins produced by the planktonic dinoflagellate Alexandrium spp. that act on the voltage-gated Na⁺ channel of nerve cells, leading to loss of motor control and abnormal swimming on copepod [68]. Exposure to Gambierdiscus sp. (as Gambierdiscus toxicus) and Fukuyoa sp. (as Gambierdiscus cf. yasumotoi) was lethal to the adult brine shrimp Artemia spp. [31] and the copepod Tigriopus japonicus [57]. In the present study, adults of A. salina showed high sensitivity to G. excentricus in both growth phases (respectively, tD₅₀ = 2h and 4h) at 200 cells ml^-1, with 100% mortality after 7-h exposure to cells in exponential phase. Similarly, strains of Gambierdiscus sp. (as G. toxicus) showed high toxicity to Artemia with median lethal dose (LD₅₀) from 2.8 to 104.5 cells individual^-1 [31]. Lower effects on survival, with less than 60% of individuals dying in 6-d exposure were found in copepods exposed to Fukuyoa sp. (as Gambierdiscus cf. yasumotoi) at 200 cells ml^-1 [57], the same abundance used in the present study. However, a suppression in gene expression related to stress or detoxification has suggested a deficiency in detoxification of ciguatoxins by these copepods [57]. Differential responses in zooplankton sensitivity may be related to the toxicity of the species Fukuyoa sp. [57] and G. excentricus (current study), as well as the feeding selectivity against harmful prey or sensitivity of the tested species (T. japonicus and A. salina, respectively).

The toxin content or cell quota may vary along the growth phases of a dinoflagellate species in a batch culture and may be lower during the exponential phase relative to the stationary and the declining phase, as observed for O. cf. ovata [69–70]. In eight species of Gambierdiscus, MTX-related hemolytic activity increased from log phase (exponential) to late log-early stationary phase, but then declined in mid-stationary phase, although no significant differences in toxicity were observed among growth phases [66]. Further study is needed to determine if the amount of toxin per cell and toxin profile varies along the growth phases of Gambierdiscus species. In the current study, significant differences between the toxic effects of dinoflagellate growth phases was solely found in survival rates of A. salina exposed to G. excentricus. Exposure to cells in stationary phase induced an initial quick response on A. salina survival, with high mortality of 50% within the first 3-h incubation, and 75% mortality at the end of 7-h incubation. In contrast, G. excentricus cells in exponential phase induced a steady decrease in survival rates and caused 100% mortality of brine shrimps at the end of incubation. The difference in A. salina survival rates may be related to an effect of the production of different toxin congeners with varying toxicities for the brine shrimps in each growth phase. Moreover, G. excentricus is a large thecate (with cellulose plates) dinoflagellate and it is likely that aged cells (in stationary phase) are thicker than actively growing (in exponential phase) cells. As cells enter the stationary phase, the growth rate diminishes, but cells may continue to expand in size. In many armored species, the edges of each thecal plate overlap, sliding apart as the cells increase in size, likely producing a more robust theca. The thicker and more robust aged cell is possibly more resistant to the digestion by A. salina, which will affect toxin assimilation rate and ultimately the survival of potential grazers.

Finally, acute exposure to the toxic benthic dinoflagellates Prorocentrum lima, Ostreopsis cf. ovata and Gambierdiscus excentricus isolated from tropical marine systems induced severe effects on behavior and survival of adult individuals of Artemia salina. Further studies are necessary to identify the different modes of action of diverse toxic compounds produced by benthic dinoflagellates on their primary grazers. Negative effects related to the toxicity of benthic dinoflagellates are expected on diverse grazers making them more vulnerable to predation and vectors of toxins through the marine food webs.

Supporting information

S1 File.

https://doi.org/10.1371/journal.pone.0175168.s001

(XLS)

Author Contributions

Conceptualization: RN TF LS SN.
Data curation: RN TF.
Formal analysis: RN LS.
Funding acquisition: LS SN.
Investigation: RN TF LS SN.
Methodology: RN TF LS SN.
Resources: LS SN.
Validation: RN TF LS SN.
Visualization: RN LS SN.
Writing – original draft: RN TF LS SN.
Writing – review & editing: RN LS SN.

References

1. Berdalet E, Tester P, Zingone A. Global Ecology and Oceanography of Harmful Algal Blooms—GEOHAB Core Research Project: HABs in Benthic Systems. Paris and Newark: IOC of UNESCO and SCOR; 2012.
2. Mangialajo L, Bertolotto R, Cattaneo-Vietti R, Chiantore M, Grillo C, Lemee R, et al. The toxic benthic dinoflagellate Ostreopsis ovata: Quantification of proliferation along the coastline of Genoa, Italy. Mar Pollut Bull. 2008;56:1209–14. pmid:18381216
- View Article
- PubMed/NCBI
- Google Scholar
3. Accoroni S, Totti C. The toxic benthic dinoflagellates of the genus Ostreopsis in temperate areas: a review. Adv Oceanogr Limnol. 2016;7:1–15.
- View Article
- Google Scholar
4. Aligizaki K, Katikou P, Milandri A, Diogène J. Occurrence of palytoxin-group toxins in seafood and future strategies to complement the present state of the art. Toxicon. 2011;57:390–9. pmid:21126531
- View Article
- PubMed/NCBI
- Google Scholar
5. Nascimento SM, Corrêa EV, Menezes M, Varela D, Paredes J, Morris S. Growth and toxin profile of Ostreopsis cf. ovata (Dinophyta) from Rio de Janeiro, Brazil. Harmful Algae. 2012;13:1–9.
- View Article
- Google Scholar
6. Granéli E, Ferreira CEL, Yasumoto T, Rodrigues E, Maria H, Neves B. Sea urchins poisoning by the benthic dinoflagellate Ostreopsis ovata on the Brazilian coast. In: Steidinger KA, Landsberg JH, Tomas CR, Vargo GA, editors. Proceedings of the 10th International Conference on Harmful Algae; 2002 Oct 21–25; Florida, USA. Florida: Florida Fish and Wildlife Conservation Commission, Florida Institute of Oceanography and Intergovernmental Oceanographic Commission of UNESCO; 2002. p. 113.
7. Shears NT, Ross PM. Blooms of benthic dinoflagellates of the genus Ostreopsis; an increasing and ecologically important phenomenon on temperate reefs in New Zealand and worldwide. Harmful Algae. 2009;8:916–25.
- View Article
- Google Scholar
8. Tubaro A, Durando P, Del Favero G, Ansaldi F, Icardi G, Deeds JR, et al. Case definitions for human poisoning postulated to palytoxins exposure. Toxicon. 2011;57:478–95. pmid:21255599
- View Article
- PubMed/NCBI
- Google Scholar
9. Hwang BS, Yoon EY, Kim HS, Yih W, Park JY, Jeong HJ, et al. Ostreol A: a new cytotoxic compound isolated from the epiphytic dinoflagellate Ostreopsis cf. ovata from the coastal waters of Jeju Island, Korea. Bioorg Med Chem Lett. 2013;23:3023–7. pmid:23562061
- View Article
- PubMed/NCBI
- Google Scholar
10. Aligizaki K, Katikou P, Nikolaidis G. Toxic benthic dinoflagellates spreading and potential risk in the Mediterranean Sea. In: Lassus P, editor. Proceedings of the 7th International Conference in Molluscan Shellfish Safety; 2009 Jun 14–19; Nantes, France. Nantes: Ifremer; 2009. p. 1–6.
11. Murata M, Shimatani M, Sugitani H, Oshima Y, Yasumoto T. Isolation and structural elucidation of the causative toxin of the diarrhetic shellfish poisoning. Nippon Suisan Gakki. 1982;48:549–52.
- View Article
- Google Scholar
12. Hoppenrath M, Chomérat N, Horiguchi T, Schweikert M, Nagahama Y, Murray S. Taxonomy and phylogeny of the benthic Prorocentrum species (Dinophyceae)–a proposal and review. Harmful Algae. 2013;27:1–28.
- View Article
- Google Scholar
13. Nascimento SM, Salgueiro F, Menezes M, Oliveira FA, Magalhães VCP, De Paula JC, et al. Prorocentrum lima from the South Atlantic: Morphological, molecular and toxicological characterization. Harmful Algae. 2016;57:39–48.
- View Article
- Google Scholar
14. Gayoso AM, Dover S., Morton SL, Busman M, Moeller PDR, Maranda L. Possibility of diarrhetic shellfish poisoning associated with Prorocentrum lima (Dinophyceae) in Patagonian Gulfs (Argentina). J Shellfish Res. 2002;21:461–3.
- View Article
- Google Scholar
15. Lawrence JE, Bauder AG, Quilliam MA, Cembella AD. Prorocentrum lima: a putative link to diarrhetic shellfish poisoning in Nova Scotia, Canada. In: Reguera B, Blanco J, Luisa Fernandez M, Wyatt T, editors. Harmful Algae. Paris: International Oceanographic Commission of UNESCO; 1998. p. 78–79.
16. Nascimento SM, Purdie DA, Morris S. Morphology, toxin composition and pigment content of Prorocentrum lima strains isolated from a coastal lagoon in southern UK. Toxicon. 2005; 45: 633–49. pmid:15777960
- View Article
- PubMed/NCBI
- Google Scholar
17. Koike K, Sato S, Yamaji M, Nagahama Y, Kotaki Y, Ogata T, et al. Occurrence of okadaic acid producing Prorocentrum lima on the Sanriku coast, northern Japan. Toxicon. 1998;36:2039–42. pmid:9839688
- View Article
- PubMed/NCBI
- Google Scholar
18. Fraga S, Rodríguez F, Caillaud A, Diogène J, Raho N, Zapata M. Gambierdiscus excentricus sp. nov. (Dinophyceae), a benthic toxic dinoflagellate from the Canary Islands (NE Atlantic Ocean). Harmful Algae. 2011;11:10–22.
- View Article
- Google Scholar
19. Fleming LE, Baden DG, Bean JA, Weisman R, Blythe DG. Seafood toxin diseases: issues in epidemiology and community outreach. In: Reguera B, Blanco J, Fernandez ML, Wyatt T, editors. Proceedings of the 8th International Conference on Harmful Algae; 1997 June 25–29; Vigo, Spain. Paris: Xunta de Galicia and Intergovernmental Oceanographic Commission of UNESCO, 1998. p. 245–8.
20. Hallegraeff GM. Ocean climate change, phytoplankton community responses, and harmful algal blooms: a formidable predictive challenge. J Phycol. 2010;46:220–35.
- View Article
- Google Scholar
21. Guisande C, Frangópulos M, Carotenuto Y, Maneiro I, Riveiro I, Vergara AR. Fate of paralytic shellfish poisoning toxins ingested by the copepod Acartia clausi. Mar Ecol Prog Ser. 2002;240:105–15.
- View Article
- Google Scholar
22. Basti L, Nagai K, Shimasaki Y, Oshima Y, Honjo T, Segawa S. Effects of the toxic dinoflagellate Heterocapsa circularisquama on the valve movement behaviour of the Manila clam Ruditapes philippinarum. Aquaculture. 2009;291:41–7.
- View Article
- Google Scholar
23. Turner JT, Borkman DG. Impact of zooplankton grazing on Alexandrium blooms in the offshore Gulf of Maine. Deep Sea Res Part II: Top Stud Oceanogr. 2006;52:2801–16.
- View Article
- Google Scholar
24. Giussani V, Costa E, Pecorino D, Berdalet E, De Giampaulis G, Gentile M, et al. Effects of the harmful dinoflagellate Ostreopsis cf. ovata on different life cycle stages of the common jellyfish Aurelia sp. Harmful Algae. 2016;57:49–58.
- View Article
- Google Scholar
25. Sopanen S, Setälä O, Piiparinen J, Erler K, Kremp A. The toxic dinoflagellate Alexandrium ostenfeldii promotes incapacitation of the calanoid copepods Eurytemora affinis and Acartia bifilosa from the northern Baltic Sea. J Plankton Res. 2011;33:1564–73.
- View Article
- Google Scholar
26. Landsberg JH. The effects of harmful algal blooms on aquatic organisms. Rev Fish Sci. 2002;10:113–390.
- View Article
- Google Scholar
27. Shumway SE, Burkholder JM, Springer J. Effects of the estuarine dinoflagellate Pfiesteria shumwayae (Dinophyceae) on survival and grazing activity of several shellfish species. Harmful Algae. 2006;5:442–58.
- View Article
- Google Scholar
28. Silva NJ, Tang KW, Lopes RM. Effects of microalgal exudates and intact cells on subtropical marine zooplankton. J Plankton Res. 2013;35:855–65.
- View Article
- Google Scholar
29. Shumway SE, Allen SM, Dee Boersma P. Marine birds and harmful algal blooms: sporadic victims or under-reported events? Harmful Algae. 2003;2:1–17.
- View Article
- Google Scholar
30. Lopes VM, Baptista M, Repolho T, Rosa R, Costa PR. Uptake, transfer and elimination kinetics of paralytic shellfish toxins in common octopus (Octopus vulgaris). Aquat Toxicol. 2014;146:205–11. pmid:24316438
- View Article
- PubMed/NCBI
- Google Scholar
31. Kelly AM, Kohler CC, Tindall DR. Are crustaceans linked to the ciguatera food chain? Environ Biol Fishes. 1992;33:275–86.
- View Article
- Google Scholar
32. Hégaret H, Wikfors GH, Shumway SE. Diverse feeding responses of five species of bivalve mollusc when exposed to three species of harmful algae. J Shellfish Res. 2007;26:549–59.
- View Article
- Google Scholar
33. Neves RAF, Figueiredo GM, Valentin JL, Silva Scardua PM, Hégaret H. Immunological and physiological responses of the periwinkle Littorina littorea during and after exposure to the toxic dinoflagellate Alexandrium minutum. Aquat Toxicol. 2015;160:96–105. pmid:25621399
- View Article
- PubMed/NCBI
- Google Scholar
34. Turner JT, Tester PA. Toxic marine phytoplankton, zooplankton grazers, and pelagic food webs. Limnol Oceanogr. 1997;42:1203–14.
- View Article
- Google Scholar
35. Turner JT, Hopcroft RR, Lincoln JA, Huestis CS, Tester PA, Roff JC. Zooplankton feeding ecology: grazing by marine copepods and cladocerans upon phytoplankton and cyanobacteria from Kingston Harbour, Jamaica. PSZNI: Mar Ecol. 1998;19:195–208.
- View Article
- Google Scholar
36. Lasley-Rasher RS, Nagel K, Angra A, Yen J. Intoxicated copepods: ingesting toxic phytoplankton leads to risky behaviour. Proc R Soc B. 2016;283:20160176.
- View Article
- Google Scholar
37. Bricelj VM, Connell L, Konoki K, MacQuarrie SP, Scheuer T, Catterall WA, et al. Sodium channel mutation leading to saxitoxin resistance in clams increases risk of PSP. Nature. 2005;434:763–7. pmid:15815630
- View Article
- PubMed/NCBI
- Google Scholar
38. Faimali M, Giussani V, Piazza V, Garaventa F, Corrà C, Asnaghi V, et al. Toxic effects of harmful benthic dinoflagellate Ostreopsis ovata on invertebrate and vertebrate marine organisms. Mar Environ Res. 2012;76:97–107. pmid:22000703
- View Article
- PubMed/NCBI
- Google Scholar
39. Nascimento SM, Melo G, Salgueiro F, Diniz BDS, Fraga S. Morphology of Gambierdiscus excentricus (Dinophyceae) with emphasis on sulcal plates. Phycologia. 2016;54:628–39.
- View Article
- Google Scholar
40. Guillard RRJ. Culture Methods. In: Hallegraeff GM, Anderson DM, Cembella AD, editors. Manual on Harmful Marine Microalgae—IOC Manual and Guides No. 33. France: UNESCO; 1995. p. 45–56.
41. Hillebrand H, Dürselen CD, Kirschtel D, Pollinger U, Zohary T. Biovolume calculation for pelagic and benthic microalgae. J Phycol. 1999;35:403–24.
- View Article
- Google Scholar
42. Menden-Deuer S, Lessard EJ. Carbon to volume relationships for dinoflagellates, diatoms, and other protist plankton. Limnol Oceanogr. 2000;45:569–79.
- View Article
- Google Scholar
43. Ohman MD, Theilacker GH, Kaupp SE. Immunochemical Detection of Predation on Ciliate Protists by Larvae of the Northern Anchovy (Engraulis mordax). Biol Bull. 1991;181:500–4.
- View Article
- Google Scholar
44. Ajuzie CC. Palatability and fatality of the dinoflagellate Prorocentrum lima to Artemia salina. J Appl Phycol. 2007;19:513–9.
- View Article
- Google Scholar
45. Leps J, Smilauer P. Multivariate analysis of ecological data using CANOCO. New York: Cambridge; 2003.
46. Hosmer DW, Lemeshow S. Applied survival analysis: regression modelling of time to event data. New York: Wiley; 1999.
47. Marcoval MA, Pan JN, Tang Y, Gobler CJ. The ability of the branchiopod, Artemia salina, to graze upon harmful algal blooms caused by Alexandrium fundyense, Aureococcus anophagefferens, and Cochlodinium polykrikoides. Estuar Coast Shelf Sci. 2013;131:235–44.
- View Article
- Google Scholar
48. Lincoln JA, Turner JT, Bates SS, Léger C, Gauthier DA. Feeding, egg production, and egg hatching success of the copepods Acartia tonsa and Temora longicornis on diets of the toxic diatom Pseudo-nitzschia multiseries and the non-toxic diatom Pseudo-nitzschia pungens. Hydrobiologia. 2001;453:107–20.
- View Article
- Google Scholar
49. Zhenxing W, Yinglin Z, Mingyuan Z, Zongling W, Dan W. Effects of toxic Alexandrium species on the survival and feeding rates of brine shrimp, Artemia salina. Acta Ecologica Sinica. 2006;26:3942–7.
- View Article
- Google Scholar
50. Ramos V, Vasconcelos V. Palytoxin and Analogs: Biological and Ecological Effects. Mar Drugs. 2010; 8: 2021–37. pmid:20714422
- View Article
- PubMed/NCBI
- Google Scholar
51. Tan ZJ, Yan T, Yu RC, Zhou MJ. Transfer of paralytic shellfish toxins via marine food chains: a simulated experiment. Biomed Environ Sci. 2007;20:235–41. pmid:17672215
- View Article
- PubMed/NCBI
- Google Scholar
52. El Ghazali I, Saqrane S, Carvalho AP, Ouahid Y, Del Campo F, Oudra B, et al. Effect of different microcystin profiles on toxic boaccumulation in common carp (Cyprinus carpio) larvae via Artemia nauplii. Ecotoxicol Environ Saf. 2010;73:762–70. pmid:20045191
- View Article
- PubMed/NCBI
- Google Scholar
53. Beattie KA, Ressler J, Wiegand C, Krause E, Codd GA, Steinberg CEW, et al. Comparative effects and metabolism of two microcystins and nodularin in the brine shrimp Artemia salina. Aquat Toxicol. 2003;62:219–26. pmid:12560170
- View Article
- PubMed/NCBI
- Google Scholar
54. Furlan M, Antonioli M, Zingone A, Sardo A, Blason C, Pallavicini A, et al. Molecular identification of Ostreopsis cf. ovata in filter feeders and putative predators. Harmful Algae. 2013;21–22:20–9.
- View Article
- Google Scholar
55. Nunes BS, Carvalho FlD, Guilhermino LCM, Van Stappen G. Use of the genus Artemia in ecotoxicity testing. Environ Pollut. 2006;144:453–62. pmid:16677747
- View Article
- PubMed/NCBI
- Google Scholar
56. Garaventa F, Gambardella C, Di Fino A, Pittore M, Faimali M. Swimming speed alteration of Artemia sp. and Brachionus plicatilis as a sub-lethal behavioural end-point for ecotoxicological surveys. Ecotoxicology. 2010; 19: 512–19. pmid:20099027
- View Article
- PubMed/NCBI
- Google Scholar
57. Lee KW, Kang JH, Baek SH, Choi YU, Lee DW, Park HS. Toxicity of the dinoflagellate Gambierdiscus sp. toward the marine copepod Tigriopus japonicus. Harmful Algae. 2014;37:62–7.
- View Article
- Google Scholar
58. Engström-Öst J, Lehtiniemi M, Green S, Kozlowsky-Suzuki B, Viitasalo M. Does cyanobacterial toxin accumulate in mysid shrimps and fish via copepods? J Exp Mar Biol Ecol. 2002;276:95–107.
- View Article
- Google Scholar
59. Samson JC, Shumway SE, Weis JS. Effects of the toxic dinoflagellate, Alexandrium fundyense on three species of larval fish: a food-chain approach. J Fish Biol. 2008;72:168–88.
- View Article
- Google Scholar
60. Forward RB Jr, Rittschof D. Activation of photoresponses of brine shrimp nauplii involved in diel vertical migration by chemical cues from fish. J Plankton Res. 1993; 15: 693–701.
- View Article
- Google Scholar
61. Demaret A, Sohet K, Houvenaghel G. Effects of toxic dinoflagellates on the feeding and mortality of Artemia franciscana larvae. In: Lassus P, Arzul G, Erard-Le Denn E, Gentien P, Marcaillou-Le Baut C, editors. Proceedings of the 6th International Conference on Toxic Marine Phytoplankton; 1993 October 18–22; Nantes, France. Paris: Lavoisier, 1995. p. 427–32.
62. Rossini GP, Bigiani A. Palytoxin action on the Na⁺, K⁺-ATPase and the disruption of ion equilibria in biological systems. Toxicon. 2011;57:429–39. pmid:20932855
- View Article
- PubMed/NCBI
- Google Scholar
63. Pezzolesi L, Guerrini F, Ciminiello P, Dell’Aversano C, Dello Iacovo E, Fattorusso E, et al. Influence of temperature and salinity on Ostreopsis cf. ovata growth and evaluation of toxin content through HR LC-MS and biological assays. Water Res. 2012;46:82–92. pmid:22078255
- View Article
- PubMed/NCBI
- Google Scholar
64. Rhodes L, Munday R, Heasman K, Briggs L, Holland P, Adamson J. Uptake of palytoxin-related compounds by shellfish and paddle crabs. In: Busby P, editor. Proceedings of the 6th International Conference of Molluscan Shellfish Safety; 2007 March18-23; Blenheim, New Zealand. Wellington: Royal Society of New Zealand, 2008. p. 168–76.
65. Holland WC, Litaker RW, Tomas CR, Kibler SR, Place AR, Davenport ED, et al. Differences in the toxicity of six Gambierdiscus (Dinophyceae) species measured using an in vitro human erytrocyte lysis assay. Toxicon. 2013;65:15–33. pmid:23313447
- View Article
- PubMed/NCBI
- Google Scholar
66. Lombet A, Bidard JN, Lazdunski M. Ciguatoxin and brevetoxins share a common receptor site on the neuronal voltage-dependent Na+ channel. FEBS Lett. 1987;219:355–9. pmid:2440718
- View Article
- PubMed/NCBI
- Google Scholar
67. Vasconcelos VT, Azevedo J, Silva M, Ramos VT. Effects of marine toxins on the reproduction and early stages development of aquatic organisms. Mar Drugs. 2010;8:59. pmid:20161971
- View Article
- PubMed/NCBI
- Google Scholar
68. Bagoien E, Miranda A, Reguera B, Franco JM. Effects of two paralytic shellfish toxin producing dinoflagellates on the pelagic harpacticoid copepod Euterpina acutifrons. Mar Biol. 1996;126:361–9.
- View Article
- Google Scholar
69. Guerrini F, Pezzolesi L, Feller A, Riccardi M, Ciminiello P, Dell’ Aversano C, et al. Comparative growth and toxin profile of cultured Ostreopsis ovata from the Tyrrhenian and Adriatic Seas. Toxicon. 2010;55:211–20. pmid:19638281
- View Article
- PubMed/NCBI
- Google Scholar
70. Granéli E, Vidyarathna NK, Funari E, Cumaranatunga PRT, Scenati R. Can increases in temperature stimulate blooms of the toxic benthic dinoflagellate Ostreopsis ovata? Harmful Algae. 2011;10:165–72.
- View Article
- Google Scholar

[ref1] 1. Berdalet E, Tester P, Zingone A. Global Ecology and Oceanography of Harmful Algal Blooms—GEOHAB Core Research Project: HABs in Benthic Systems. Paris and Newark: IOC of UNESCO and SCOR; 2012.

[ref2] 2. Mangialajo L, Bertolotto R, Cattaneo-Vietti R, Chiantore M, Grillo C, Lemee R, et al. The toxic benthic dinoflagellate Ostreopsis ovata: Quantification of proliferation along the coastline of Genoa, Italy. Mar Pollut Bull. 2008;56:1209–14. pmid:18381216
View Article
PubMed/NCBI
Google Scholar

[3] View Article

[4] PubMed/NCBI

[5] Google Scholar

[ref3] 3. Accoroni S, Totti C. The toxic benthic dinoflagellates of the genus Ostreopsis in temperate areas: a review. Adv Oceanogr Limnol. 2016;7:1–15.
View Article
Google Scholar

[7] View Article

[8] Google Scholar

[ref4] 4. Aligizaki K, Katikou P, Milandri A, Diogène J. Occurrence of palytoxin-group toxins in seafood and future strategies to complement the present state of the art. Toxicon. 2011;57:390–9. pmid:21126531
View Article
PubMed/NCBI
Google Scholar

[10] View Article

[11] PubMed/NCBI

[12] Google Scholar

[ref5] 5. Nascimento SM, Corrêa EV, Menezes M, Varela D, Paredes J, Morris S. Growth and toxin profile of Ostreopsis cf. ovata (Dinophyta) from Rio de Janeiro, Brazil. Harmful Algae. 2012;13:1–9.
View Article
Google Scholar

[14] View Article

[15] Google Scholar

[ref6] 6. Granéli E, Ferreira CEL, Yasumoto T, Rodrigues E, Maria H, Neves B. Sea urchins poisoning by the benthic dinoflagellate Ostreopsis ovata on the Brazilian coast. In: Steidinger KA, Landsberg JH, Tomas CR, Vargo GA, editors. Proceedings of the 10th International Conference on Harmful Algae; 2002 Oct 21–25; Florida, USA. Florida: Florida Fish and Wildlife Conservation Commission, Florida Institute of Oceanography and Intergovernmental Oceanographic Commission of UNESCO; 2002. p. 113.

[ref7] 7. Shears NT, Ross PM. Blooms of benthic dinoflagellates of the genus Ostreopsis; an increasing and ecologically important phenomenon on temperate reefs in New Zealand and worldwide. Harmful Algae. 2009;8:916–25.
View Article
Google Scholar

[18] View Article

[19] Google Scholar

[ref8] 8. Tubaro A, Durando P, Del Favero G, Ansaldi F, Icardi G, Deeds JR, et al. Case definitions for human poisoning postulated to palytoxins exposure. Toxicon. 2011;57:478–95. pmid:21255599
View Article
PubMed/NCBI
Google Scholar

[21] View Article

[22] PubMed/NCBI

[23] Google Scholar

[ref9] 9. Hwang BS, Yoon EY, Kim HS, Yih W, Park JY, Jeong HJ, et al. Ostreol A: a new cytotoxic compound isolated from the epiphytic dinoflagellate Ostreopsis cf. ovata from the coastal waters of Jeju Island, Korea. Bioorg Med Chem Lett. 2013;23:3023–7. pmid:23562061
View Article
PubMed/NCBI
Google Scholar

[25] View Article

[26] PubMed/NCBI

[27] Google Scholar

[ref10] 10. Aligizaki K, Katikou P, Nikolaidis G. Toxic benthic dinoflagellates spreading and potential risk in the Mediterranean Sea. In: Lassus P, editor. Proceedings of the 7th International Conference in Molluscan Shellfish Safety; 2009 Jun 14–19; Nantes, France. Nantes: Ifremer; 2009. p. 1–6.

[ref11] 11. Murata M, Shimatani M, Sugitani H, Oshima Y, Yasumoto T. Isolation and structural elucidation of the causative toxin of the diarrhetic shellfish poisoning. Nippon Suisan Gakki. 1982;48:549–52.
View Article
Google Scholar

[30] View Article

[31] Google Scholar

[ref12] 12. Hoppenrath M, Chomérat N, Horiguchi T, Schweikert M, Nagahama Y, Murray S. Taxonomy and phylogeny of the benthic Prorocentrum species (Dinophyceae)–a proposal and review. Harmful Algae. 2013;27:1–28.
View Article
Google Scholar

[33] View Article

[34] Google Scholar

[ref13] 13. Nascimento SM, Salgueiro F, Menezes M, Oliveira FA, Magalhães VCP, De Paula JC, et al. Prorocentrum lima from the South Atlantic: Morphological, molecular and toxicological characterization. Harmful Algae. 2016;57:39–48.
View Article
Google Scholar

[36] View Article

[37] Google Scholar

[ref14] 14. Gayoso AM, Dover S., Morton SL, Busman M, Moeller PDR, Maranda L. Possibility of diarrhetic shellfish poisoning associated with Prorocentrum lima (Dinophyceae) in Patagonian Gulfs (Argentina). J Shellfish Res. 2002;21:461–3.
View Article
Google Scholar

[39] View Article

[40] Google Scholar

[ref15] 15. Lawrence JE, Bauder AG, Quilliam MA, Cembella AD. Prorocentrum lima: a putative link to diarrhetic shellfish poisoning in Nova Scotia, Canada. In: Reguera B, Blanco J, Luisa Fernandez M, Wyatt T, editors. Harmful Algae. Paris: International Oceanographic Commission of UNESCO; 1998. p. 78–79.

[ref16] 16. Nascimento SM, Purdie DA, Morris S. Morphology, toxin composition and pigment content of Prorocentrum lima strains isolated from a coastal lagoon in southern UK. Toxicon. 2005; 45: 633–49. pmid:15777960
View Article
PubMed/NCBI
Google Scholar

[43] View Article

[44] PubMed/NCBI

[45] Google Scholar

[ref17] 17. Koike K, Sato S, Yamaji M, Nagahama Y, Kotaki Y, Ogata T, et al. Occurrence of okadaic acid producing Prorocentrum lima on the Sanriku coast, northern Japan. Toxicon. 1998;36:2039–42. pmid:9839688
View Article
PubMed/NCBI
Google Scholar

[47] View Article

[48] PubMed/NCBI

[49] Google Scholar

[ref18] 18. Fraga S, Rodríguez F, Caillaud A, Diogène J, Raho N, Zapata M. Gambierdiscus excentricus sp. nov. (Dinophyceae), a benthic toxic dinoflagellate from the Canary Islands (NE Atlantic Ocean). Harmful Algae. 2011;11:10–22.
View Article
Google Scholar

[51] View Article

[52] Google Scholar

[ref19] 19. Fleming LE, Baden DG, Bean JA, Weisman R, Blythe DG. Seafood toxin diseases: issues in epidemiology and community outreach. In: Reguera B, Blanco J, Fernandez ML, Wyatt T, editors. Proceedings of the 8th International Conference on Harmful Algae; 1997 June 25–29; Vigo, Spain. Paris: Xunta de Galicia and Intergovernmental Oceanographic Commission of UNESCO, 1998. p. 245–8.

[ref20] 20. Hallegraeff GM. Ocean climate change, phytoplankton community responses, and harmful algal blooms: a formidable predictive challenge. J Phycol. 2010;46:220–35.
View Article
Google Scholar

[55] View Article

[56] Google Scholar

[ref21] 21. Guisande C, Frangópulos M, Carotenuto Y, Maneiro I, Riveiro I, Vergara AR. Fate of paralytic shellfish poisoning toxins ingested by the copepod Acartia clausi. Mar Ecol Prog Ser. 2002;240:105–15.
View Article
Google Scholar

[58] View Article

[59] Google Scholar

[ref22] 22. Basti L, Nagai K, Shimasaki Y, Oshima Y, Honjo T, Segawa S. Effects of the toxic dinoflagellate Heterocapsa circularisquama on the valve movement behaviour of the Manila clam Ruditapes philippinarum. Aquaculture. 2009;291:41–7.
View Article
Google Scholar

[61] View Article

[62] Google Scholar

[ref23] 23. Turner JT, Borkman DG. Impact of zooplankton grazing on Alexandrium blooms in the offshore Gulf of Maine. Deep Sea Res Part II: Top Stud Oceanogr. 2006;52:2801–16.
View Article
Google Scholar

[64] View Article

[65] Google Scholar

[ref24] 24. Giussani V, Costa E, Pecorino D, Berdalet E, De Giampaulis G, Gentile M, et al. Effects of the harmful dinoflagellate Ostreopsis cf. ovata on different life cycle stages of the common jellyfish Aurelia sp. Harmful Algae. 2016;57:49–58.
View Article
Google Scholar

[67] View Article

[68] Google Scholar

[ref25] 25. Sopanen S, Setälä O, Piiparinen J, Erler K, Kremp A. The toxic dinoflagellate Alexandrium ostenfeldii promotes incapacitation of the calanoid copepods Eurytemora affinis and Acartia bifilosa from the northern Baltic Sea. J Plankton Res. 2011;33:1564–73.
View Article
Google Scholar

[70] View Article

[71] Google Scholar

[ref26] 26. Landsberg JH. The effects of harmful algal blooms on aquatic organisms. Rev Fish Sci. 2002;10:113–390.
View Article
Google Scholar

[73] View Article

[74] Google Scholar

[ref27] 27. Shumway SE, Burkholder JM, Springer J. Effects of the estuarine dinoflagellate Pfiesteria shumwayae (Dinophyceae) on survival and grazing activity of several shellfish species. Harmful Algae. 2006;5:442–58.
View Article
Google Scholar

[76] View Article

[77] Google Scholar

[ref28] 28. Silva NJ, Tang KW, Lopes RM. Effects of microalgal exudates and intact cells on subtropical marine zooplankton. J Plankton Res. 2013;35:855–65.
View Article
Google Scholar

[79] View Article

[80] Google Scholar

[ref29] 29. Shumway SE, Allen SM, Dee Boersma P. Marine birds and harmful algal blooms: sporadic victims or under-reported events? Harmful Algae. 2003;2:1–17.
View Article
Google Scholar

[82] View Article

[83] Google Scholar

[ref30] 30. Lopes VM, Baptista M, Repolho T, Rosa R, Costa PR. Uptake, transfer and elimination kinetics of paralytic shellfish toxins in common octopus (Octopus vulgaris). Aquat Toxicol. 2014;146:205–11. pmid:24316438
View Article
PubMed/NCBI
Google Scholar

[85] View Article

[86] PubMed/NCBI

[87] Google Scholar

[ref31] 31. Kelly AM, Kohler CC, Tindall DR. Are crustaceans linked to the ciguatera food chain? Environ Biol Fishes. 1992;33:275–86.
View Article
Google Scholar

[89] View Article

[90] Google Scholar

[ref32] 32. Hégaret H, Wikfors GH, Shumway SE. Diverse feeding responses of five species of bivalve mollusc when exposed to three species of harmful algae. J Shellfish Res. 2007;26:549–59.
View Article
Google Scholar

[92] View Article

[93] Google Scholar

[ref33] 33. Neves RAF, Figueiredo GM, Valentin JL, Silva Scardua PM, Hégaret H. Immunological and physiological responses of the periwinkle Littorina littorea during and after exposure to the toxic dinoflagellate Alexandrium minutum. Aquat Toxicol. 2015;160:96–105. pmid:25621399
View Article
PubMed/NCBI
Google Scholar

[95] View Article

[96] PubMed/NCBI

[97] Google Scholar

[ref34] 34. Turner JT, Tester PA. Toxic marine phytoplankton, zooplankton grazers, and pelagic food webs. Limnol Oceanogr. 1997;42:1203–14.
View Article
Google Scholar

[99] View Article

[100] Google Scholar

[ref35] 35. Turner JT, Hopcroft RR, Lincoln JA, Huestis CS, Tester PA, Roff JC. Zooplankton feeding ecology: grazing by marine copepods and cladocerans upon phytoplankton and cyanobacteria from Kingston Harbour, Jamaica. PSZNI: Mar Ecol. 1998;19:195–208.
View Article
Google Scholar

[102] View Article

[103] Google Scholar

[ref36] 36. Lasley-Rasher RS, Nagel K, Angra A, Yen J. Intoxicated copepods: ingesting toxic phytoplankton leads to risky behaviour. Proc R Soc B. 2016;283:20160176.
View Article
Google Scholar

[105] View Article

[106] Google Scholar

[ref37] 37. Bricelj VM, Connell L, Konoki K, MacQuarrie SP, Scheuer T, Catterall WA, et al. Sodium channel mutation leading to saxitoxin resistance in clams increases risk of PSP. Nature. 2005;434:763–7. pmid:15815630
View Article
PubMed/NCBI
Google Scholar

[108] View Article

[109] PubMed/NCBI

[110] Google Scholar

[ref38] 38. Faimali M, Giussani V, Piazza V, Garaventa F, Corrà C, Asnaghi V, et al. Toxic effects of harmful benthic dinoflagellate Ostreopsis ovata on invertebrate and vertebrate marine organisms. Mar Environ Res. 2012;76:97–107. pmid:22000703
View Article
PubMed/NCBI
Google Scholar

[112] View Article

[113] PubMed/NCBI

[114] Google Scholar

[ref39] 39. Nascimento SM, Melo G, Salgueiro F, Diniz BDS, Fraga S. Morphology of Gambierdiscus excentricus (Dinophyceae) with emphasis on sulcal plates. Phycologia. 2016;54:628–39.
View Article
Google Scholar

[116] View Article

[117] Google Scholar

[ref40] 40. Guillard RRJ. Culture Methods. In: Hallegraeff GM, Anderson DM, Cembella AD, editors. Manual on Harmful Marine Microalgae—IOC Manual and Guides No. 33. France: UNESCO; 1995. p. 45–56.

[ref41] 41. Hillebrand H, Dürselen CD, Kirschtel D, Pollinger U, Zohary T. Biovolume calculation for pelagic and benthic microalgae. J Phycol. 1999;35:403–24.
View Article
Google Scholar

[120] View Article

[121] Google Scholar

[ref42] 42. Menden-Deuer S, Lessard EJ. Carbon to volume relationships for dinoflagellates, diatoms, and other protist plankton. Limnol Oceanogr. 2000;45:569–79.
View Article
Google Scholar

[123] View Article

[124] Google Scholar

[ref43] 43. Ohman MD, Theilacker GH, Kaupp SE. Immunochemical Detection of Predation on Ciliate Protists by Larvae of the Northern Anchovy (Engraulis mordax). Biol Bull. 1991;181:500–4.
View Article
Google Scholar

[126] View Article

[127] Google Scholar

[ref44] 44. Ajuzie CC. Palatability and fatality of the dinoflagellate Prorocentrum lima to Artemia salina. J Appl Phycol. 2007;19:513–9.
View Article
Google Scholar

[129] View Article

[130] Google Scholar

[ref45] 45. Leps J, Smilauer P. Multivariate analysis of ecological data using CANOCO. New York: Cambridge; 2003.

[ref46] 46. Hosmer DW, Lemeshow S. Applied survival analysis: regression modelling of time to event data. New York: Wiley; 1999.

[ref47] 47. Marcoval MA, Pan JN, Tang Y, Gobler CJ. The ability of the branchiopod, Artemia salina, to graze upon harmful algal blooms caused by Alexandrium fundyense, Aureococcus anophagefferens, and Cochlodinium polykrikoides. Estuar Coast Shelf Sci. 2013;131:235–44.
View Article
Google Scholar

[134] View Article

[135] Google Scholar

[ref48] 48. Lincoln JA, Turner JT, Bates SS, Léger C, Gauthier DA. Feeding, egg production, and egg hatching success of the copepods Acartia tonsa and Temora longicornis on diets of the toxic diatom Pseudo-nitzschia multiseries and the non-toxic diatom Pseudo-nitzschia pungens. Hydrobiologia. 2001;453:107–20.
View Article
Google Scholar

[137] View Article

[138] Google Scholar

[ref49] 49. Zhenxing W, Yinglin Z, Mingyuan Z, Zongling W, Dan W. Effects of toxic Alexandrium species on the survival and feeding rates of brine shrimp, Artemia salina. Acta Ecologica Sinica. 2006;26:3942–7.
View Article
Google Scholar

[140] View Article

[141] Google Scholar

[ref50] 50. Ramos V, Vasconcelos V. Palytoxin and Analogs: Biological and Ecological Effects. Mar Drugs. 2010; 8: 2021–37. pmid:20714422
View Article
PubMed/NCBI
Google Scholar

[143] View Article

[144] PubMed/NCBI

[145] Google Scholar

[ref51] 51. Tan ZJ, Yan T, Yu RC, Zhou MJ. Transfer of paralytic shellfish toxins via marine food chains: a simulated experiment. Biomed Environ Sci. 2007;20:235–41. pmid:17672215
View Article
PubMed/NCBI
Google Scholar

[147] View Article

[148] PubMed/NCBI

[149] Google Scholar

[ref52] 52. El Ghazali I, Saqrane S, Carvalho AP, Ouahid Y, Del Campo F, Oudra B, et al. Effect of different microcystin profiles on toxic boaccumulation in common carp (Cyprinus carpio) larvae via Artemia nauplii. Ecotoxicol Environ Saf. 2010;73:762–70. pmid:20045191
View Article
PubMed/NCBI
Google Scholar

[151] View Article

[152] PubMed/NCBI

[153] Google Scholar

[ref53] 53. Beattie KA, Ressler J, Wiegand C, Krause E, Codd GA, Steinberg CEW, et al. Comparative effects and metabolism of two microcystins and nodularin in the brine shrimp Artemia salina. Aquat Toxicol. 2003;62:219–26. pmid:12560170
View Article
PubMed/NCBI
Google Scholar

[155] View Article

[156] PubMed/NCBI

[157] Google Scholar

[ref54] 54. Furlan M, Antonioli M, Zingone A, Sardo A, Blason C, Pallavicini A, et al. Molecular identification of Ostreopsis cf. ovata in filter feeders and putative predators. Harmful Algae. 2013;21–22:20–9.
View Article
Google Scholar

[159] View Article

[160] Google Scholar

[ref55] 55. Nunes BS, Carvalho FlD, Guilhermino LCM, Van Stappen G. Use of the genus Artemia in ecotoxicity testing. Environ Pollut. 2006;144:453–62. pmid:16677747
View Article
PubMed/NCBI
Google Scholar

[162] View Article

[163] PubMed/NCBI

[164] Google Scholar

[ref56] 56. Garaventa F, Gambardella C, Di Fino A, Pittore M, Faimali M. Swimming speed alteration of Artemia sp. and Brachionus plicatilis as a sub-lethal behavioural end-point for ecotoxicological surveys. Ecotoxicology. 2010; 19: 512–19. pmid:20099027
View Article
PubMed/NCBI
Google Scholar

[166] View Article

[167] PubMed/NCBI

[168] Google Scholar

[ref57] 57. Lee KW, Kang JH, Baek SH, Choi YU, Lee DW, Park HS. Toxicity of the dinoflagellate Gambierdiscus sp. toward the marine copepod Tigriopus japonicus. Harmful Algae. 2014;37:62–7.
View Article
Google Scholar

[170] View Article

[171] Google Scholar

[ref58] 58. Engström-Öst J, Lehtiniemi M, Green S, Kozlowsky-Suzuki B, Viitasalo M. Does cyanobacterial toxin accumulate in mysid shrimps and fish via copepods? J Exp Mar Biol Ecol. 2002;276:95–107.
View Article
Google Scholar

[173] View Article

[174] Google Scholar

[ref59] 59. Samson JC, Shumway SE, Weis JS. Effects of the toxic dinoflagellate, Alexandrium fundyense on three species of larval fish: a food-chain approach. J Fish Biol. 2008;72:168–88.
View Article
Google Scholar

[176] View Article

[177] Google Scholar

[ref60] 60. Forward RB Jr, Rittschof D. Activation of photoresponses of brine shrimp nauplii involved in diel vertical migration by chemical cues from fish. J Plankton Res. 1993; 15: 693–701.
View Article
Google Scholar

[179] View Article

[180] Google Scholar

[ref61] 61. Demaret A, Sohet K, Houvenaghel G. Effects of toxic dinoflagellates on the feeding and mortality of Artemia franciscana larvae. In: Lassus P, Arzul G, Erard-Le Denn E, Gentien P, Marcaillou-Le Baut C, editors. Proceedings of the 6th International Conference on Toxic Marine Phytoplankton; 1993 October 18–22; Nantes, France. Paris: Lavoisier, 1995. p. 427–32.

[ref62] 62. Rossini GP, Bigiani A. Palytoxin action on the Na⁺, K⁺-ATPase and the disruption of ion equilibria in biological systems. Toxicon. 2011;57:429–39. pmid:20932855
View Article
PubMed/NCBI
Google Scholar

[183] View Article

[184] PubMed/NCBI

[185] Google Scholar

[ref63] 63. Pezzolesi L, Guerrini F, Ciminiello P, Dell’Aversano C, Dello Iacovo E, Fattorusso E, et al. Influence of temperature and salinity on Ostreopsis cf. ovata growth and evaluation of toxin content through HR LC-MS and biological assays. Water Res. 2012;46:82–92. pmid:22078255
View Article
PubMed/NCBI
Google Scholar

[187] View Article

[188] PubMed/NCBI

[189] Google Scholar

[ref64] 64. Rhodes L, Munday R, Heasman K, Briggs L, Holland P, Adamson J. Uptake of palytoxin-related compounds by shellfish and paddle crabs. In: Busby P, editor. Proceedings of the 6th International Conference of Molluscan Shellfish Safety; 2007 March18-23; Blenheim, New Zealand. Wellington: Royal Society of New Zealand, 2008. p. 168–76.

[ref65] 65. Holland WC, Litaker RW, Tomas CR, Kibler SR, Place AR, Davenport ED, et al. Differences in the toxicity of six Gambierdiscus (Dinophyceae) species measured using an in vitro human erytrocyte lysis assay. Toxicon. 2013;65:15–33. pmid:23313447
View Article
PubMed/NCBI
Google Scholar

[192] View Article

[193] PubMed/NCBI

[194] Google Scholar

[ref66] 66. Lombet A, Bidard JN, Lazdunski M. Ciguatoxin and brevetoxins share a common receptor site on the neuronal voltage-dependent Na+ channel. FEBS Lett. 1987;219:355–9. pmid:2440718
View Article
PubMed/NCBI
Google Scholar

[196] View Article

[197] PubMed/NCBI

[198] Google Scholar

[ref67] 67. Vasconcelos VT, Azevedo J, Silva M, Ramos VT. Effects of marine toxins on the reproduction and early stages development of aquatic organisms. Mar Drugs. 2010;8:59. pmid:20161971
View Article
PubMed/NCBI
Google Scholar

[200] View Article

[201] PubMed/NCBI

[202] Google Scholar

[ref68] 68. Bagoien E, Miranda A, Reguera B, Franco JM. Effects of two paralytic shellfish toxin producing dinoflagellates on the pelagic harpacticoid copepod Euterpina acutifrons. Mar Biol. 1996;126:361–9.
View Article
Google Scholar

[204] View Article

[205] Google Scholar

[ref69] 69. Guerrini F, Pezzolesi L, Feller A, Riccardi M, Ciminiello P, Dell’ Aversano C, et al. Comparative growth and toxin profile of cultured Ostreopsis ovata from the Tyrrhenian and Adriatic Seas. Toxicon. 2010;55:211–20. pmid:19638281
View Article
PubMed/NCBI
Google Scholar

[207] View Article

[208] PubMed/NCBI

[209] Google Scholar

[ref70] 70. Granéli E, Vidyarathna NK, Funari E, Cumaranatunga PRT, Scenati R. Can increases in temperature stimulate blooms of the toxic benthic dinoflagellate Ostreopsis ovata? Harmful Algae. 2011;10:165–72.
View Article
Google Scholar

[211] View Article

[212] Google Scholar

Figures

Abstract

Introduction

Material and methods

Algae cultures

Experimental design

Feeding assays.

Acute toxicity assays.

Statistical analyses

Results

Feeding assays

Acute toxicity assays

Discussion

Supporting information

S1 File.

Author Contributions

References