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Dinoflagellates are marine organisms that undergo seasonal proliferation events known as algal blooms. Vegetative cell proliferation is a main contributing factor in these events. However, mechanistical understanding of mitosis and cytokinesis in dinoflagellates remains rudimentary. Using an optimized immunofluorescence protocol, we analysed changes in microtubule organization occurring during the mitotic cycle of the toxic dinoflagellate Ostreopsis cf. ovata. We find that the flagella and the cortical microtubule array persist throughout the mitotic cycle. Two cytoplasmic microtubule bundles originate from the ventral area, where the basal bodies are located - a cortical bundle and a cytoplasmic bundle. The latter associates with the nucleus in the cell centre before mitosis and with the acentrosomal extranuclear spindle during mitosis. Analysis of tubulin post-translational modifications identifies two populations of spindle microtubules - polar acetylated microtubules, whose length is constant, and central tyrosinated microtubules, which elongate during chromosome segregation. During cell division a microtubule-rich structure forms along the dorsal-ventral axis, associated with the site of cytokinesis, consistent with a cytokinetic mechanism that is independent of the actomyosin ring typical of animal and yeast cells., Competing Interests: Competing interests The authors declare no competing or financial interests., (© 2024. Published by The Company of Biologists Ltd.)

Dinophyte evolution is essentially inferred from the pattern of thecal plates, and two different labelling systems are used for the important subgroups Gonyaulacales and Peridiniales. The partiform hypotheca of cladopyxidoid dinophytes fits into the morphological concepts of neither group, although they are assigned to the Gonyaulacales. Here, we describe the thecate dinophyte Fensomea setacea, gen. & sp. nov., which has a cladopyxidoid tabulation. The cells displayed a Kofoidean plate formula APC, 3', 4a, 7″, 7C, 6S, 6''', 2'''', and slender processes were randomly distributed over the echinate or baculate surface. In addition, we obtained rRNA sequences of F. setacea, gen. & sp. nov., but dinophytes that exhibit a partiform hypotheca did not show a close relationship to Gonyaulacales. Character evolution of thecate dinophytes may have progressed from the ancestral state of six postcingular plates, and two more or less symmetrically arranged antapical plates, towards patterns of only five postcingular plates (Peridiniales) or more asymmetrical configurations (Gonyaulacales). Based on our phylogenetic reconsiderations the contact between the posterior sulcal plate and the first postcingular plate, as well as the contact between an antapical plate and the distalmost postcingular plate, do not represent a rare, specialized gonyaulacoid plate configuration (i.e., the partiform hypotheca of cladopyxidoid dinophytes). Instead, these contacts correspond to the common and regular configuration of peridinioid (and other) dinophytes.

Many corals establish symbiosis with Symbiodiniaceae cells from surrounding environments, but very few Symbiodiniaceae cells exist in the water column. Given that the N -acetyl-d-glucosamine-binding lectin ActL attracts Symbiodiniaceae cells, we hypothesized that corals must attract Symbiodiniaceae cells using ActL to acquire them. Anti-ActL antibody inhibited acquisition of Symbiodiniaceae cells, and rearing seawater for juvenile Acropora tenuis contained ActL, suggesting that juvenile A. tenuis discharge ActL to attract these cells. Among eight Symbiodiniaceae cultured strains, ActL attracted NBRC102920 ( Symbiodinium tridacnidorum ) most strongly followed by CS-161 ( Symbiodinium tridacnidorum ), CCMP2556 ( Durusdinium trenchii ), and CCMP1633 ( Breviolum sp.); however, it did not attract GTP-A6-Sy ( Symbiodinium natans ), CCMP421 ( Effrenium voratum ), FKM0207 ( Fugacium sp.), and CS-156 ( Fugacium sp.). Juvenile polyps of A. tenuis acquired limited Symbiodiniaceae cell strains, and the number of acquired Symbiodiniaceae cells in a polyp also differed from each other. The number of Symbiodiniaceae cells acquired by juvenile polyps of A. tenuis was correlated with the ActL chemotactic activity. Thus, ActL could be used to attract select Symbiodiniaceae cells and help Symbiodiniaceae cell acquisition in juvenile polyps of A. tenuis , facilitating establishment of symbiosis between A. tenuis and Symbiodiniaceae cells.

Background: Dinoflagellates are aquatic protists particularly widespread in the oceans worldwide. Some are responsible for toxic blooms while others live in symbiotic relationships, either as mutualistic symbionts in corals or as parasites infecting other protists and animals. Dinoflagellates harbor atypically large genomes (~ 3 to 250 Gb), with gene organization and gene expression patterns very different from closely related apicomplexan parasites. Here we sequenced and analyzed the genomes of two early-diverging and co-occurring parasitic dinoflagellate Amoebophrya strains, to shed light on the emergence of such atypical genomic features, dinoflagellate evolution, and host specialization., Results: We sequenced, assembled, and annotated high-quality genomes for two Amoebophrya strains (A25 and A120), using a combination of Illumina paired-end short-read and Oxford Nanopore Technology (ONT) MinION long-read sequencing approaches. We found a small number of transposable elements, along with short introns and intergenic regions, and a limited number of gene families, together contribute to the compactness of the Amoebophrya genomes, a feature potentially linked with parasitism. While the majority of Amoebophrya proteins (63.7% of A25 and 59.3% of A120) had no functional assignment, we found many orthologs shared with Dinophyceae. Our analyses revealed a strong tendency for genes encoded by unidirectional clusters and high levels of synteny conservation between the two genomes despite low interspecific protein sequence similarity, suggesting rapid protein evolution. Most strikingly, we identified a large portion of non-canonical introns, including repeated introns, displaying a broad variability of associated splicing motifs never observed among eukaryotes. Those introner elements appear to have the capacity to spread over their respective genomes in a manner similar to transposable elements. Finally, we confirmed the reduction of organelles observed in Amoebophrya spp., i.e., loss of the plastid, potential loss of a mitochondrial genome and functions., Conclusion: These results expand the range of atypical genome features found in basal dinoflagellates and raise questions regarding speciation and the evolutionary mechanisms at play while parastitism was selected for in this particular unicellular lineage.

Eyes are not unique to animals. As described by Nilsson and Marshall, prominent eyes, complete with retina and lens, have unexpectedly evolved in single cell dinoflagellates., (Crown Copyright © 2020. Published by Elsevier Inc. All rights reserved.)

The cell cycle is a critical component of cellular proliferation, differentiation, and response to stress, yet its role in the regulation of intracellular symbioses is not well understood. To explore host-symbiont cell cycle coordination in a marine symbiosis, we employed a model for coral-dinoflagellate associations: the tropical sea anemone Aiptasia ( Exaiptasia pallida ) and its native microalgal photosymbionts ( Breviolum minutum and Breviolum psygmophilum ). Using fluorescent labeling and spatial point-pattern image analyses to characterize cell population distributions in both partners, we developed protocols that are tailored to the three-dimensional cellular landscape of a symbiotic sea anemone tentacle. Introducing cultured symbiont cells to symbiont-free adult hosts increased overall host cell proliferation rates. The acceleration occurred predominantly in the symbiont-containing gastrodermis near clusters of symbionts but was also observed in symbiont-free epidermal tissue layers, indicating that the presence of symbionts contributes to elevated proliferation rates in the entire host during colonization. Symbiont cell cycle progression differed between cultured algae and those residing within hosts; the endosymbiotic state resulted in increased S-phase but decreased G 2 /M-phase symbiont populations. These phenotypes and the deceleration of cell cycle progression varied with symbiont identity and host nutritional status. These results demonstrate that host and symbiont cells have substantial and species-specific effects on the proliferation rates of their mutualistic partners. This is the first empirical evidence to support species-specific regulation of the symbiont cell cycle within a single cnidarian-dinoflagellate association; similar regulatory mechanisms likely govern interpartner coordination in other coral-algal symbioses and shape their ecophysiological responses to a changing climate. IMPORTANCE Biomass regulation is critical to the overall health of cnidarian-dinoflagellate symbioses. Despite the central role of the cell cycle in the growth and proliferation of cnidarian host cells and dinoflagellate symbionts, there are few studies that have examined the potential for host-symbiont coregulation. This study provides evidence for the acceleration of host cell proliferation when in local proximity to clusters of symbionts within cnidarian tentacles. The findings suggest that symbionts augment the cell cycle of not only their enveloping host cells but also neighboring cells in the epidermis and gastrodermis. This provides a possible mechanism for rapid colonization of cnidarian tissues. In addition, the cell cycles of symbionts differed depending on nutritional regime, symbiotic state, and species identity. The responses of cell cycle profiles to these different factors implicate a role for species-specific regulation of symbiont cell cycles within host cnidarian tissues., (Copyright © 2020 Tivey et al.)

Toxicities of the marine algae Alexandrium minutum and its excreted gonyautoxins (GTXs) to the marine crustacean Artemia salina were investigated. Mortality was observed for neither larvae nor adult A. salina exposed to A. minutum at a density of 5000 cells/mL or 0.5 μM GTX2/3. After exposure, the full transcriptome of adult A. salina was assembled and functionally annotated. A total of 599,286 transcripts were obtained, which were clustered into 515,196 unigenes. Results of the transcriptional effect level index revealed that direct exposure to the toxic algae A. minutum caused greater alterations in the transcriptome than did exposure to the extracellular product GTX2/3. Mechanisms of effects were different between exposure of A. salina to A. minutum cells or GTX2/3. Exposure to A. minutum modulated formation of the ribonucleoprotein complex and metabolism of amino acids and lipids in A. salina. Exposure to GTX2/3 exposure inhibited expression of genes related to metabolism of chitin, which might result in disruption of molting process or disturbed sheath morphogenesis. Overall, effects on transcription observed in this study represent the first report based on application of next generation sequencing techniques to investigate the transcriptomic response of A. salina exposed to an environmentally realistic level of A. minutum or GTX2/3., (Copyright © 2019 Elsevier Ltd. All rights reserved.)

Dinoflagellates of the genus Symbiodinium form symbiotic relationships with corals, other marine invertebrates, and protists; thus, they are considered as important species in coral reef ecosystems. If Symbiodinium could be successfully cryopreserved, the cell bank generated could prove to be a valuable resource for researchers interested in basic biological research of Symbiodinium -invertebrate symbioses. Herein, successful cryopreservation of clade D Symbiodinium was achieved using a two-step freezing protocol. Symbiodinium cells were exposed to cryoprotectants (CPAs) for 30 minutes before being vapor frozen for 20 minutes in liquid nitrogen (LN 2 ); afterward, cells were immediately immersed in LN 2 for 2 hours or 10 days. The initial experiment was conducted with the following CPAs at 1, 2, and 3 M concentrations: methanol, dimethyl sulfoxide, glycerol, ethylene glycol (EG), and propylene glycol (PG). It was found that infiltration with 2 M EG and PG yielded cells with the highest percentage viability. Upon thawing, culture of these Symbiodinium was carried out for 2 months in a growth chamber, and cells continued to grow and proliferate over this period. This represents successful cryopreservation of a dominant reef coral symbiont, a feat that will ideally aid in future research of this important lineage of dinoflagellate.

Marine unarmored dinoflagellates in the family Kareniaceae are known to possess chloroplasts of haptophyte origin, which contain fucoxanthin and its derivatives as major carotenoids, and lack peridinin. In the present study, the first species with the peridinin-type chloroplast in this family, Gertia stigmatica gen. et sp. nov., is described on the basis of ultrastructure, photosynthetic pigment composition, and molecular phylogeny inferred from nucleus- and chloroplast-encoded genes. Cells of G. stigmatica were small and harboring a chloroplast with an eyespot and two pyrenoids. The apical structure complex was straight, similar to Karenia and Karlodinium. Under transmission electron microscopy, the chloroplast was surrounded by two membranes, and the eyespot was composed of a single layer of osmiophilic globules (eyespot type A); this was never previously reported from the Kareniaceae. High performance liquid chromatography demonstrated the chloroplast contains peridinin, and neither fucoxanthin nor 19'-acyloxyfucoxanthins was identified. A phylogeny based on nucleus-encoded rDNAs suggested a position of G. stigmatica in the Kareniaceae, but not clustered within the previously described genera, i.e., Karenia, Karlodinium and Takayama. A phylogeny of chloroplast-encoded psbA, psbC and psbD indicated the chloroplast is of peridinin-type typical of dinoflagellates, but the most related species remains unclear., (Copyright © 2019 Elsevier GmbH. All rights reserved.)

Pseudothecadinium campbellii, a phototrophic, thecate, marine benthic species, has been found in the Sea of Japan, Russia. The morphological description of the species has been emended, and the thecal tabulation pattern is now APC 4' 2a 4″ 6c 6 s 5‴ 1⁗. Our study indicates that P. campbellii is related to Thecadinium kofoidii and Thecadiniopsis tasmanica, based on a unique morphological feature: incomplete precingular plate series. Previously, molecular data was not available for the taxa, and thus the phylogenetic position of P. campbellii within the Dinophyceae remained obscure. In this study, analyses of the rRNA gene sequences (partial 18S and 28S) revealed that unexpectedly, P. campbellii is most closely related to Halostylodinium arenarium. It formed a common clade with the Thecadinium sensu stricto clade comprising T. kofoidii and T. pseudokofoidii. This clade was placed within the order Gonyaulacales. However, almost no similarity in morphology was found between the two genetically closest species. In addition, they have different lifestyles: unlike P. campbellii, the nonmotile stage is dominant in the life cycle of H. arenarium. It has been shown that other genetically similar species (T. kofoidii, T. pseudokofoidii, T. yashimaense) exhibited some morphological features that unite them with gonyaulacoids., (Copyright © 2019 Elsevier GmbH. All rights reserved.)

Approximately 70 species of Prorocentrum are known, of which around 30 species are associated with benthic habitats. Some produce okadaic acid (OA), dinophysistoxin (DTX) and their derivatives, which are involved in diarrhetic shellfish poisoning. In this study, we isolated and characterized Prorocentrum concavum and P. malayense from Broome in north Western Australia using light and scanning electron microscopy as well as molecular sequences of large subunit regions of ribosomal DNA, marking the first record of these species from Australian waters. The morphology of the motile cells of P. malayense was similar to P. concavum in the light microscopy, but differed by the smooth thecal surface, the pore pattern and the production of mucous stalk-like structures and a hyaline sheath around the non-motile cells. P. malayense could also be differentiated from other closely related species, P. leve and P. foraminosum , despite the similarity in thecal surface and pore pattern, by its platelet formula and morphologies. We tested the production of OA and DTXs from both species, but found that they did not produce detectable levels of these toxins in the given culturing conditions. This study aids in establishing more effective monitoring of potential harmful algal taxa in Australian waters for aquaculture and recreational purposes.

Dinoflagellates from the Symbiodiniaceae family and corals have an ecologically important endosymbiotic relationship. Scleractinian corals cannot survive for long periods without their symbionts. These algae, also known as zooxanthellae, on the other hand, thrives outside the coral cells. The free-living populations of zooxanthellae are essential for the resilience of the coral to environmental stressors such as temperature anomalies and ocean acidification. Yet, little is known about how ocean acidification may affect the free-living zooxanthellae. In this study we aimed to test morphological, physiological and biochemical responses of zooxanthellae from the Symbiodinium genus isolated from the coral Mussismilia braziliensis, endemic to the Brazilian coast, to acidification led by increased atmospheric CO2. We tested whether photosynthetic yield, cell ultrastructure, cell density and lipid profile would change after up to 16 days of exposure to pH 7.5 in an atmospheric pCO2 of 1633 μatm. Photosynthetic yield and cell density were negatively affected and chloroplasts showed vesiculated thylakoids, indicating morphological damage. Moreover, Symbiodinium fatty acid profile drastically changed in acidified condition, showing lower polyunsaturated fatty acids and higher saturated fatty acids contents, when compared to the control, non-acidified condition. These results show that seawater acidification as an only stressor causes significant changes in the physiology, biochemistry and ultrastructure of free-living Symbiodinium., Competing Interests: The authors have declared that no competing interests exist.

Changes in salinity are known to alter the morphology of protists, and we hypothesized that these changes subsequently alter also the predatory behavior of the dinoflagellate Oxyrrhis marina. Oxyrrhis was grown in media of 33, 25, 20, and 10% of the regular salinity of f/2 medium (31-32‰). In all cases, the cells discharged trichocysts and swelled. Cell surfaces and volumes increased with decreasing salinity, such that cell surface area at least doubled at 10% and the cell volume increased approximately fourfold. After 1 h, the cells started to regain their regular shape, which was almost completed after 24 h. Oxyrrhis immediately regained its regular shape when culture medium was added 5-10 min after the osmotic stress. When incubated with Pyramimonas grossii as prey, those short-term stressed cells showed no significant different prey uptake in comparison to non-stressed cells. In contrast, 24 h after the addition of prey, short-term stressed Oxyrrhis cells had, with weak statistical significance, more Pyramimonas cells engulfed than non-stressed cells. These results indicated that (1) trichocysts were most likely not involved in prey capture and (2) salinity-stressed Oxyrrhis either enhanced its capability to capture more prey, or its digestion apparatus was hampered., (Copyright © 2019 Elsevier GmbH. All rights reserved.)

Apicomplexa is a group of obligate intracellular parasites that includes the causative agents of human diseases such as malaria and toxoplasmosis. Apicomplexans evolved from free-living phototrophic ancestors, but how this transition to parasitism occurred remains unknown. One potential clue lies in coral reefs, of which environmental DNA surveys have uncovered several lineages of uncharacterized basally branching apicomplexans 1,2 . Reef-building corals have a well-studied symbiotic relationship with photosynthetic Symbiodiniaceae dinoflagellates (for example, Symbiodinium 3 ), but the identification of other key microbial symbionts of corals has proven to be challenging 4,5 . Here we use community surveys, genomics and microscopy analyses to identify an apicomplexan lineage-which we informally name 'corallicolids'-that was found at a high prevalence (over 80% of samples, 70% of genera) across all major groups of corals. Corallicolids were the second most abundant coral-associated microeukaryotes after the Symbiodiniaceae, and are therefore core members of the coral microbiome. In situ fluorescence and electron microscopy confirmed that corallicolids live intracellularly within the tissues of the coral gastric cavity, and that they possess apicomplexan ultrastructural features. We sequenced the genome of the corallicolid plastid, which lacked all genes for photosystem proteins; this indicates that corallicolids probably contain a non-photosynthetic plastid (an apicoplast 6 ). However, the corallicolid plastid differs from all other known apicoplasts because it retains the four ancestral genes that are involved in chlorophyll biosynthesis. Corallicolids thus share characteristics with both their parasitic and their free-living relatives, which suggests that they are evolutionary intermediates and implies the existence of a unique biochemistry during the transition from phototrophy to parasitism.

Perkinsozoa is an exclusively parasitic group within the alveolates and infections have been reported from various organisms, including marine shellfish, marine dinoflagellates, freshwater cryptophytes, and tadpoles. Despite its high abundance and great genetic diversity revealed by recent environmental rDNA sequencing studies, Perkinsozoa biodiversity remains poorly understood. During the intensive samplings in Korean coastal waters during June 2017, a new parasitoid of dinoflagellates was detected and was successfully established in culture. The new parasitoid was most characterized by the presence of two to four dome-shaped, short germ tubes in the sporangium. The opened germ tubes were biconvex lens-shaped in the top view and were characterized by numerous wrinkles around their openings. Phylogenetic analyses based on the concatenated SSU and LSU rDNA sequences revealed that the new parasitoid was included in the family Parviluciferaceae, in which all members were comprised of two separate clades, one containing Parvilucifera species (P. infectans, P. corolla, and P. rostrata), and the other containing Dinovorax pyriformis, Snorkelia spp., and the new parasitoid from this study. Based on morphological, ultrastructural, and molecular data, we propose to erect a new genus and species, Tuberlatum coatsi gen. n., sp. n., from the new parasitoid found in this study. Further, we examined and discussed the validity of some diagnostic characteristics reported for parasitoids in the family Parviluciferaceae at both the genus and species levels., (Copyright © 2018 Elsevier GmbH. All rights reserved.)

The planktonic phototrophic dinoflagellate Gonyaulax whaseongensis sp. nov., isolated from coastal waters of western Korea, was described from living and fixed cells under light and scanning electron microscopy, and its rDNA was sequenced. Gonyaulax whaseongensis had a plate formula of 2pr, 4', 6'', 6c, 6''', 1p, and 1'''' with S-type ventral organization like the other species in the genus. However, this dinoflagellate had a narrow cingulum (ca. 2.6 μm), small displacement of the cingulum, slight overhang and steep angle between the ends of the cingulum, quadrangular sixth precingular plate, reticulated cell surface without longitudinal lines or ridges, and two unequal antapical spines, together which distinguish this from all other reported Gonyaulax species. In addition, the SSU and LSU rDNA sequences were 8%-12% and 11%-24%, respectively, different from those of Gonyaulax polygramma, Gonyaulax spinifera, Gonyaulax fragilis, Gonyaulax membranacea, and Gonyaulax digitale, the putatively closest related species in the phylogenetic analysis., (© 2018 Phycological Society of America.)

Causative species of Harmful Algal Bloom (HAB) and toxins in commercially exploited molluscan shellfish species are monitored weekly from four classified shellfish production areas in Perú (three in the north and one in the south). Okadaic acid (OA) and pectenotoxins (PTXs) were detected in hand-picked cells of Dinophysis ( D. acuminata -complex and D. caudata ) and in scallops ( Argopecten purpuratus ), the most important commercial bivalve species in Perú. LC-MS analyses revealed two different toxin profiles associated with species of the D. acuminata -complex: (a) one with OA (0.3⁻8.0 pg cell -1 ) and PTX2 (1.5⁻11.1 pg cell -1 ) and (b) another with only PTX2 which included populations with different toxin cell quota (9.3⁻9.6 pg cell -1 and 5.8⁻9.2 pg cell -1 ). Toxin results suggest the likely presence of two morphotypes of the D. acuminata -complex in the north, and only one of them in the south. Likewise, shellfish toxin analyses revealed the presence of PTX2 in all samples (10.3⁻34.8 µg kg -1 ), but OA (7.7⁻15.2 µg kg -1 ) only in the northern samples. Toxin levels were below the regulatory limits established for diarrhetic shellfish poisoning (DSP) and PTXs (160 µg OA kg -1 ) in Perú, in all samples analyzed. This is the first report confirming the presence of OA and PTX in Dinophysis cells and in shellfish from Peruvian coastal waters.

A study of modern sediment from the Western Arctic has revealed the presence of a distinctive brown-colored cyst with a spherical central body bearing unbranched processes that are usually solid with a small basal pericoel. Distinctive barbs project from some processes, and process tips are usually minutely expanded into conjoined barbs. The archeopyle is apical and saphopylic. This cyst corresponds to Islandinium? cezare morphotype 2 of Head et al. (2001, J. Quat. Sci., 16:621). Phylogenetic analyses based on the small and large subunit rRNA genes infer close relationship with Islandinium minutum, the type of which is that of the genus. Re-examination of specimens of I. minutum reveals the presence of minute barbs on its processes, but differences with Islandinium? cezare morphotype 2 remain based on size, process distribution, and barb development. Furthermore, the internal transcribed spacer shows I. minutum to be distinct from this morphotype. On the basis of these small but discrete differences, we propose the new subspecies Islandinium minutum subsp. barbatum subsp. nov. Molecular sequencing of other cysts encountered, namely Echinidinium karaense, an unidentified flattened cyst, and "Polykrikos quadratus", places them in the Monovela clade, the latter showing greater morphological variability than previously thought., (© 2018 The Author(s) Journal of Eukaryotic Microbiology © 2018 International Society of Protistologists.)

The genus Gymnodinium includes many morphologically similar species, but molecular phylogenies show that it is polyphyletic. Eight strains of Gymnodinium impudicum, Gymnodinium dorsalisulcum and a novel Gymnodinium-like species from Chinese and Malaysian waters and the Mediterranean Sea were established. All of these strains were examined with light microscopy, scanning electron microscopy and transmission electron microscopy. SSU, LSU and internal transcribed spacers rDNA sequences were obtained. A new genus, Wangodinium, was erected to incorporate strains with a loop-shaped apical structure complex (ASC) comprising two rows of amphiesmal vesicles, here referred to as a new type of ASC. The chloroplasts of Wangodinium sinense are enveloped by two membranes. Pigment analysis shows that peridinin is the main accessory pigment in W. sinense. Wangodinium differs from other genera mainly in its unique ASC, and additionally differs from Gymnodinium in the absence of nuclear chambers, and from Lepidodinium in the absence of Chl b and nuclear chambers. New morphological information was provided for G. dorsalisulcum and G. impudicum, e.g., a short sulcal intrusion in G. dorsalisulcum; nuclear chambers in G. impudicum and G. dorsalisulcum; and a chloroplast enveloped by two membranes in G. impudicum. Molecular phylogeny was inferred using maximum likelihood and Bayesian inference with independent SSU and LSU rDNA sequences. Our results support the classification of Wangodinium within the Gymnodiniales sensu stricto clade and it is close to Lepidodinium. Our results also support the close relationship among G. dorsalisulcum, G. impudicum, and Barrufeta. Further research is needed to assign these Gymnodinium species to Barrufeta or to erect new genera., (© 2018 Phycological Society of America.)

Three strains of the toxic benthic dinoflagellate Prorocentrum hoffmannianum were isolated in the Canary Islands (north-east Atlantic Ocean, Spain). The identity of the strains was determined by phylogenetic analyses of partial LSU rDNA (D1-D2 regions) but their morphology based on SEM images corresponded to P. maculosum. Their toxin profiles were analyzed by liquid chromatography and high resolution mass spectrometry analysis (LC-HRMS) on cell extracts and culture media. Okadaic acid and three analogs were detected in all strains. Rather, in culture media the detected compounds were variable among strains, two of them being okadaic acid analogs not found on cell extracts. As a result, the taxonomy of the species was revised and P. maculosum is proposed as a junior synonym of P. hoffmannianum whose description is emended., (Copyright © 2018. Published by Elsevier B.V.)

The mixotroph Yihiella yeosuensis is a small- and fast-swimming dinoflagellate. To investigate its protistan predators, interactions between Y. yeosuensis and 11 heterotrophic protists were explored. No potential predators were able to feed on actively swimming Y. yeosuensis cells, which escaped via rapid jumps, whereas Aduncodinium glandula, Oxyrrhis marina, and Strombidinopsis sp. (approximately 150 μm in cell length) were able to feed on weakly swimming cells that could not jump. Furthermore, Gyrodinium dominans, Luciella masanensis, and Pfiesteria piscicida were able to feed on heat-killed Yihiella cells, whereas Gyrodinium moestrupii, Noctiluca scintillans, Oblea rotunda, Polykrikos kofoidii, and Strombidium sp. (20 μm) did not feed on them. Thus, the jumping behavior of Y. yeosuensis might be primarily responsible for the observed lack of predation. With increasing Yihiella concentration, the growth rate of O. marina decreased, whereas that of Strombidinopsis did not change. However, with increasing Yihiella concentration (up to 530 ng C/ml), the ingestion rate of Strombidinopsis on Yihiella increased linearly. The highest ingestion rate was 24.1 ng C per predator per d. The low daily carbon acquisition from Yihiella relative to the body carbon content of Strombidinopsis might be responsible for its negligible growth. Thus, Y. yeosuensis might have an advantage over its competitors due to its low mortality rate., (© 2018 The Author(s) Journal of Eukaryotic Microbiology © 2018 International Society of Protistologists.)

The formation, propagation, and maintenance of harmful algal blooms are of interest due to their negative effects on marine life and human health. Some bloom-forming algae utilize allelopathy, the release of compounds that inhibit competitors, to exclude other species dependent on a common pool of limiting resources. Allelopathy is hypothesized to affect bloom dynamics and is well established in the red tide dinoflagellate Karenia brevis. K. brevis typically suppresses competitor growth rather than being acutely toxic to other algae. When we investigated the effects of allelopathy on two competitors, Asterionellopsis glacialis and Thalassiosira pseudonana, using nuclear magnetic resonance (NMR) spectroscopy and mass spectrometry (MS)-based metabolomics, we found that the lipidomes of both species were significantly altered. However, A. glacialis maintained a more robust metabolism in response to K. brevis allelopathy whereas T. pseudonana exhibited significant alterations in lipid synthesis, cell membrane integrity, and photosynthesis. Membrane-associated lipids were significantly suppressed for T. pseudonana exposed to allelopathy such that membranes of living cells became permeable. K. brevis allelopathy appears to target lipid biosynthesis affecting multiple physiological pathways suggesting that exuded compounds have the ability to significantly alter competitor physiology, giving K. brevis an edge over sensitive species.

The occurrence of the ciguatera fish poisoning (CFP) causative Gambierdiscus spp. was confirmed in the Sea of Japan for the first time in 2009. This paper reports seasonal distribution of Gambierdiscus spp. and epiphytic diatoms in the Sea of Japan. Monitoring results suggested an antagonistic interaction in abundances between epiphytic diatoms and the dinoflagellate Gambierdiscus spp. Allelopathic effects of diatoms were considered to be involved in the competitive phenomenon. Therefore it is hypothesized that cell densities of epiphytic pennate diatoms on macroalgae are a novel determinant affecting the abundance of Gambierdiscus spp. other than sea water temperature, salinity and nutrients. Monitorings of the abundance of epiphytic diatoms would lead us to predict the occurrences of Gambierdiscus spp. blooms in the CFP area, and thereby the CFP risk assessments would be developed. Phylogenetic analyses indicated that Gambierdiscus spp. in the Sea of Japan belonged to Gambierdiscus sp. type 2 which was reported to be non-toxic. Nevertheless, based on morphological characteristics, at least two types of Gambierdiscus spp. were found in the Sea of Japan. It is needed to test the toxicity of the both types of Gambierdiscus recognized in the present study for evaluation of the probability of CFP outbreak risks in the Sea of Japan in the future., (Copyright © 2018 Elsevier B.V. All rights reserved.)

Oxygen supply is an important factor during Crypthecodinium cohnii fermentation for docosahexaenoic acid (DHA) production. However, few studies about the intrinsic correlation between dissolved oxygen (DO) and cellular metabolism have been reported. In this study, the responses of C. cohnii to different DO levels were evaluated. The results showed the growth and glucose consumption rates of C. cohnii were much higher under high oxygen supply condition. Furthermore, GC-MS based comparative metabolomic analysis was employed to discriminate the responsive metabolites associated with varying DO levels. The results showed the intermediates involved in glycolytic pathway and TCA cycle were up-regulated under high DO levels at exponential phase. At stationary phase, under high DO levels, metabolites involved in triacylglycerol metabolism were up-regulated, while the OPP pathway intermediate product ribose 5-phosphate was down-regulated. Together, these results provide useful insights into the functional metabolic relationship between DO levels and DHA production in C. cohnii., (Copyright © 2018 Elsevier Inc. All rights reserved.)

The total dinoflagellate cyst community and the cysts of Cochlodinium polykrikoides in the surface sediments of South Sea (Tongyeong coast), South Korea, were analysed using next-generation sequencing (NGS) and morphological approaches. Dinoflagellate cysts can be highly abundant (111-4,087 cysts g -1 dry weight) and have diverse species composition. A total of 35 taxa of dinoflagellate cysts representing 16 genera, 21 species (including four unconfirmed species), and 14 complex species were identified by NGS analysis. Cysts of Scrippsiella spp (mostly Scrippsiella trochoidea) were the most dominant and Polykrikos schwartzii, Pentapharsodinium dalei, Ensiculifera carinata, and Alexandrium catenella/tamarense were common. Thus, a combination of NGS and morphological analysis is effective for studying the cyst communities present in a given environment. Although C. polykrikoides developed massive blooms during 2013-2014, microscopy revealed low density of their cysts, whereas no cysts were detected by NGS. However, the vegetative C. polykrikoides not appeared during 2015-2017 in spite of the observation of C. polykrikoides cysts. This suggests that the C. polykrikoides blooms were not due to development of their cysts but to other factors such as currents transporting them to a marine environment suitable for their growth.

Short-period variability in plankton communities is poorly documented, especially for variations occurring in specific groups in the assemblage because traditional analysis is laborious and time-consuming. Moreover, it does not allow the high sampling frequency required for decision making. To overcome this limitation, we tested the submersible CytoSub flow cytometer. This device was anchored at a distance of approximately 10 metres from the low tide line at a depth of 1.5 metres for 12 hours to monitor the plankton at a site in the biological reserve of Barra da Tijuca beach, Rio de Janeiro. Data analysis was performed with two-dimensional scatter plots, individual pulse shapes and micro images acquisition. High-frequency monitoring results of two interesting groups are shown. The abundance and carbon biomass of ciliates were relatively stable, whereas those from dinoflagellates were highly variable along the day. The linear regression of biovolume measures between classical microscopy and in situ flow cytometry demonstrate high degree of adjustment. Despite the success of the trial and the promising results obtained, the large volume of images generated by the method also creates a need to develop pattern recognition models for automatic classification of in situ cytometric images.

Species of Fukuyoa, recently revised from the globular Gambierdiscus, are toxic benthic dinoflagellates associated with ciguatera. In this study, a total of ten strains of Fukuyoa collected from Hong Kong waters were characterized using morphological and phylogenetic analyses. Results from both analyses showed that one of the strains is a putative new species, namely Fukuyoa sp. HK Type 1 (plate formula Po, 3', 7″, 6c, 7s, 5‴, 1p and 2'‴ with a distinctive small and narrow cell shape, narrow Po plate, high Po pore density, large and broad Plate 1' but small and round Po pore size, small and narrow Plate 2', long and narrow Plates 2'‴ and 1p), and the others were F. ruetzleri. This is the first report of these two species of Fukuyoa in the South China Sea and Asia-Pacific region. Phylogenies on 18S, 28S D1/D3 and D8/D10 ribosomal DNA sequences strongly support that Fukuyoa sp. HK Type 1 is currently the most divergent species in the genus Fukuyoa. The diagrammatic plots on the p-distance matrices of 18S, 28S D1/D3 and D8/D10 and ITS regions resolved that the species of Fukuyoa were separated into three main groups, i.e., Fukuyoa sp. HK Type 1, F. paulensis and a group consisting of F. ruetzleri, F. yasumotoi and F. cf. yasumotoi, while Fukuyoa sp. HK Type 1 was always the most distant from the other two groups. Additionally, the pairwise p-distance values calculated based on the ITS region have always been the highest for pairs between Fukuyoa sp. HK Type 1 and other Fukuyoa species, ranging from 0.142 to 0.150. Our molecular results suggested that Fukuyoa sp. HK Type 1 is a putative new species. Both morphological and molecular data of more strains from different localities should be, however, collected to address its intraspecific variability and further evaluate its taxonomic status. A bioassay analysis demonstrated that algal lysates extracted from F. ruetzleri and Fukuyoa sp. HK Type 1 were lethal to brine shrimp larvae, indicating that both species were toxic. Bulk cultures were tested for Pacific ciguatoxins (P-CTXs) and maitotoxins (MTXs) by liquid chromatography-tandem mass spectrometry (LC-MS/MS). All isolates of Fukuyoa produced neither P-CTXs nor MTX-1, but isolates of F. ruetzleri produced a compound putatively assigned as MTX-3. This study has updated the current biodiversity and distribution of the toxic benthic dinoflagellates Fukuyoa, and thus contributes to the understanding of their emerging threats to the sub-tropical reef systems locally and regionally., (Copyright © 2018 Elsevier B.V. All rights reserved.)

The family Ceratocoryaceae includes the genera Ceratocorys, Protoceratium, and Schuettiella, whose phylogenetic relationships are poorly known. Here, the new non-yessotoxin-producing species of the genus Ceratocorys, Ceratocorys mariaovidiorum sp. nov., previously reported as the toxic Protoceratium reticulatum, is described from examinations by light and scanning electron microscopy, molecular phylogeny, and toxin analyses. The species description is made from culture samples of strains CCMP1740 and CCMP404 from USA waters. Ceratocorys mariaovidiorum is globular and has thick and strongly reticulated plates with one pore within each reticule, just like P. reticulatum, but the key difference between the two species is the presence of five precingular plates in C. mariaovidiorum instead of six as in P. reticulatum. The thecal plate formula is Po, 4', 0a, 5″, 6c, ~7s, 5‴, 0p, 2''''. The apical pore plate is oval with a λ-shaped pore. The first apical plate is narrow with a ventral pore on the right anterior side; it contacts the apical pore plate and its contact with the anterior sulcal plate is slight or absent. The fourth precingular plate of other Gonyaulacales is absent. Ceratocorys mariaovidiorum may have small spines on the second antapical plate. A phylogenetic study based on internal transcribed spacer/5.8SrDNA supports the morphological classification of C. mariaovidiorum as a new species of Ceratocorys and in a different clade from P. reticulatum., (© 2017 The Authors. Journal of Phycology published by Wiley Periodicals, Inc. on behalf of Phycological Society of America.)

A planktonic-benthic relationship has been described for many dinoflagellate species as part of their ecological strategy to overcome highly variable aquatic environments. Here, the phylogenetically and morphologically related marine dinoflagellates Protoceratium reticulatum and Ceratocorys mariaovidiorum were studied in relation to an unknown benthic life form. In vivo and fixed samples from cultures were analyzed in detail by light and scanning electron microscopy. In both species, a cell type with a morphology different from that of vegetative cells was observed in cultures grown until stationary phase. This cell type was always benthic, swimming sporadically only when it was disturbed. Its main feature included a strong dorsoventral compression. These cells originated from vegetative cells whose protoplasm underwent a progressive flattening, resulting in a gradual detachment of the reticulate and thick thecal plates and the formation of very thin non-reticulated new plates with pores. When returned to fresh full-strength medium, the cells recovered their spherical vegetative-like morphology, including new reticulated thick plates and subsequent cell divisions. The kinetics of flattened cell formation showed that in both species, this cell type increased exponentially until the onset of the culture stationary phase and then decreased. The results of this study are discussed in the context of the planktonic-benthic coupling in dinoflagellate life cycles, including those newly appreciated to be well adapted to the benthic environment., (© 2017 Phycological Society of America.)

The cytoskeleton is crucial to cell mechanics and sensing the extracellular physical environment. The objective of this study was to examine the role of the cortical cytoskeleton in mechanosensitivity in a unicellular protist, the marine dinoflagellate Lingulodinium polyedra, using its intrinsic bioluminescence as a rapid reporter of mechanotransduction. Pharmacological treatments resolved effects due to immediate cytoskeleton disruption from those due to cytoskeletal remodeling during the light to dark phase transition. The cytoskeleton was visualized by confocal laser scanning microscopy of immunohistochemically labeled microtubules and phalloidin labeled F-actin, and mechanosensitivity assessed based on the bioluminescence response to mechanical stimulation measured during the dark phase. Latrunculin B treatment after the transition from the light to dark phase resulted in some disruption of cortical F-actin, no observed effect on the cortical microtubules, and partial inhibition of the bioluminescence response. Treatment with oryzalin, which depolarizes microtubules, completely disrupted the microtubule network and cortical F-actin, and partially inhibited bioluminescence. These results demonstrate that cells retain some mechanosensitivity despite a disrupted cytoskeleton; link mechanosensitivity to intact F-actin; show a close connection between F-actin and microtubules comprising the cortical cytoskeleton; confirm a strong contribution of the actin cytoskeleton to the translocation of scintillons, vesicles containing the luminescent chemistry; and support the role of the actin cytoskeleton in the association of scintillons with the vacuole membrane., (© 2017 Wiley Periodicals, Inc.)

A new marine woloszynskioid dinoflagellate Dactylodinium pterobelotum gen. et sp. nov., collected from a southern Vietnamese estuary, was described on the basis of LM, SEM, and TEM, and molecular phylogeny inferred from rDNA sequences. This species had the smallest number of amphiesmal vesicles (5 latitudinal series) in woloszynskioid dinoflagellates assigned to the Suessiaceae and Borghiellaceae. The eyespot was of type B, composed of osmiophilic globules and brick-like material, located in- and outside of the chloroplast respectively. An apical structure comprised a pair of elongate anterior vesicles (PEV). A large peduncle was conspicuous, located in the sulcal extension in the epicone, and supported by a microtubular strand of ~140 microtubules. Ultrastructural features of trichocysts represent a novel type in the Dinophyceae, bearing lateral hairs besides anterior fibers. The molecular phylogeny based on partial LSU rDNA showed the species in a basal position in the family Suessiaceae; this indicates the eyespot type B and PEV of the Borghiellaceae are ancestral states of the eyespot comprising brick-like material (type E) and an elongate apical vesicle of the Suessiaceae., (© 2017 Phycological Society of America.)

This paper reports a toxic strain of Protoceratium reticulatum, its morphology, phylogeny, yessotoxins (YTXs) production and abundance in northern Yellow Sea of China from 2011 to 2015 was investigated. YTXs in hepatopancreas and edible parts of bottom sowing cultured Japanese scallop Patinopecten yessoensis in this sea area were determined weekly for 5 years. Other potential producers of YTXs, Gonyaulax spinifera and Lingulodinium polyedrum, were also investigated. Results revealed that Protoceratium reticulatum strain from the northern Yellow Sea belongs to a geographically widely distributed species. Motile cells of Protoceratium reticulatum contribute to YTXs in Japanese scallop, and G. spinifera may also be a potential contributor. Resting cysts of Protoceratium reticulatum, G. spinifera, and L. polyedrum in sediments were possibly important origins of YTXs in scallop cultured at sea bottom. YTXs in scallop decreased from 2011 to 2015, most toxins were concentrated in hepatopancreas, while a small portion in edible parts which was safe for consumption the whole year around., (Copyright © 2017 Elsevier Ltd. All rights reserved.)

One of the most common marine dinophytes is a species known as Heterocapsa triquetra. When Stein introduced the taxon Heterocapsa, he formally based the type species H. triquetra on the basionym Glenodinium triquetrum. The latter was described by Ehrenberg and is most likely a species of Kryptoperidinium. In addition to that currently unresolved nomenclatural situation, the thecal plate composition of H. triquetra sensu Stein (1883) was controversial in the past. To clarify the debate, we collected material and established the strain UTKG7 from the Baltic Sea off Kiel (Germany, the same locality as Stein had studied), which was investigated using light and electron microscopy, and whose systematic position was inferred using molecular phylogenetics. The small motile cells (18-26 μm in length) had a biconical through fusiform shape and typically were characterized by a short asymmetrically shaped, horn-like protuberance at the antapex. A large spherical nucleus was located in the episome, whereas a single pyrenoid laid in the lower cingular plane. The predominant plate pattern was identified as apical pore complex (Po, cp?, X), 4', 2a, 6'', 6c, 5s, 5''', 2''''. The triradiate body scales were 254-306 nm in diameter, had 6 ridges radiating from a central spine, 9 peripheral and 3 radiating spines, and 12 peripheral bars as well as a central depression in the basal plate. Our work provides a clarification of morphological characters and a new, validly published name for this important but yet formally undescribed species of Heterocapsa: H. steinii sp. nov., (© 2017 Phycological Society of America.)

A gymnodinioid photosynthetic dinoflagellate was isolated from Argentina and examined by light and electron microscopy and analysis of nuclear-encoded LSU rDNA. Kirithra asteri gen. et sp. nov. was proposed as morphology and molecular phylogeny separated this dinoflagellate from others within the family Ceratoperidiniaceae. Cells were surrounded by a hyaline amphiesma comprising polygonal vesicles. Each vesicle contained a honeycomb and a trilaminar structure. An anterior sulcal extension ending in a complete circle formed the apical structure complex (ASC), which characterizes Ceratoperidiniaceae. The ASC comprised three rows of vesicles. The nucleus was located in the hypocone, and several large, irregularly shaped vesicles were present in the epi- and hypocone. Chloroplasts were surrounded by three membranes, and grana-like arrangements of thylakoids were observed in one strain used for ultrastructural study. The cell centre contained 1-3 multiple-stalked pyrenoids and membrane-bound vesicles containing tile-like structures surrounded each pyrenoid. Two pusules with collecting chambers and associated vesicles branched off each of the flagellar canals. The flagellar apparatus featured a ventral connective between the amphiesma and the R1 root, and almost opposite basal bodies, rarely seen in dinoflagellates. This was the first ultrastructural study of a species within Ceratoperidiniaceae., (Copyright © 2017 Elsevier GmbH. All rights reserved.)

Asexual and sexual life cycle events were studied in cultures of the toxic marine dinoflagellate Protoceratium reticulatum. Asexual division by desmoschisis was characterized morphologically and changes in DNA content were analyzed by flow cytometry. The results indicated that haploid cells with a C DNA content occurred only during the light period whereas a shift from a C to a 2C DNA content (indicative of S phase) took place only during darkness. The sexual life cycle was documented by examining the mating type as well as the morphology of the sexual stages and nuclei. Gamete fusion resulted in a planozygote with two longitudinal flagella, but longitudinally biflagellated cells arising from planozygote division were also observed, so one of the daughter cells retained two longitudinal flagella while the other daughter cell lacked them. Presumed planozygotes (identified by their longitudinally biflagellated form) followed two life-cycle routes: division and encystment (resting cyst formation). Both the division of longitudinally biflagellated cells and resting cyst formation are morphologically described herein. Resting cyst formation through sexual reproduction was observed in 6.1% of crosses and followed a complex heterothallic pattern. Clonal strains underwent sexuality (homothallism for planozygote formation and division) but without the production of resting cysts. Ornamental processes of resting cysts formed from the cyst wall under an outer balloon-shaped membrane and were fully developed in <1h. Obligatory dormancy period was of ∼4 months. Excystment resulted in a large, rounded, pigmented, longitudinally biflagellated but motionless, thecate germling that divided by desmoschisis. Like the planozygote, the first division of the germling yielded one longitudinally biflagellated daughter cell and another without longitudinal flagella. The longitudinal biflagellation state of both sexual stages and of the first division products of these cells is discussed., (Copyright © 2017 Elsevier B.V. All rights reserved.)

The genus Amphidinium is an important group of athecated dinoflagellates because of its high abundance in marine habitats, its member's ability to live in a variety of environmental conditions and ability to produce toxins. Furthermore, the genus is of particular interest in the biotechnology field for its potential in the pharmaceutical arena. Taxonomically the there is a history of complication and confusion over the proper identities and placements of Amphidinium species due to high genetic variability coupled with high morphological conservation. Thirteen years has passed since the most recent review of the group, and while many issues were resolved, some remain. The present study used microscopy, phylogenetics of the 28S region of rDNA, secondary structure of the ITS2 region of rDNA, compensatory base change data, and cytotoxicity data from Amphidinium strains collected world-wide to elucidate remaining confusion. This holistic approach using multiple lines of evidence resulted in a more comprehensive understanding of the morphological, ecological, and genetic characteristics that are attributed to organisms belonging to Amphidinium, including six novel species: A. fijiensis, A. magnum, A. paucianulatum, A. pseudomassartii, A. theodori, and A. tomasii., (Copyright © 2017 Elsevier B.V. All rights reserved.)

Photo-damage to photosystem II (PSII) from ultraviolet radiation (UVR) was determined using chlorophyll fluorescence in relation to sunscreen factors on the dinoflagellate Scrippsiella sweeneyae based on the cellular mycosporine-like amino acid contents (C MAAs, cell ) and cell diameter (=light path, d). Three different C MAAs, cell were prepared by acclimating cells to three levels (30.8, 15.2, and 7.7Wm -2 ) of photosynthetically active radiation (PAR). PAR-acclimated cells were exposed to PAR (0.64Wm -2 )+UVR (3.94W m -2 =2.51Wm -2 UVB+1.43Wm -2 UVA) for 12min. High PAR (HL) and medium PAR (NDF1) treatments acclimated cells to induce shinorine and porphyra-334 (longer λ max at 333 and 334nm); whereas, the low PAR (NDF2) treatment acclimated cells to induce mycosporine-glycine and palythine (shorter λ max at 310 and 320nm). Absorption spectra for the individual MAAs were reconstructed using the λ max and C MAAs, cell and were summed to reconstruct the absorption of the total C MAAs, cell (m 2 cell -1 ) to estimate the sunscreen factor (S [λ] ) at λ max . The highest S (λmax) was obtained for cells that acclimated to the highest PAR (highest C MAAs, cell and longest d); whereas, the lowest S (λmax) was obtained for cells acclimated to the lowest PAR (the lowest C MAAs, cell and the shortest d). C MAAs, cell contributed approximately 94%, whereas d contributed <6%, of the sunscreen factor (S λmax ). UVR-induced damage was indexed with a temporal decrease in the optimum quantum yield (F v /F m ) in the Photosystem II. The highest damage was observed for cells acclimated to the lowest S λmax (lowest C MAAs, cell and shortest d); whereas, the lowest damage was observed for cells acclimated to the highest S λmax (highest C MAAs, cell and longest d). The C MAAs, cell mitigated most of the UVR-induced damage in photosystem II of the dinoflagellate S. sweeneyae., (Copyright © 2017 Elsevier B.V. All rights reserved.)

Heterotrophic nanoflagellates are ubiquitous and known to be major predators of bacteria. The feeding of free-living heterotrophic nanoflagellates on phytoplankton is poorly understood, although these two components usually co-exist. To investigate the feeding and ecological roles of major heterotrophic nanoflagellates Katablepharis spp., the feeding ability of Katablepharis japonica on bacteria and phytoplankton species and the type of the prey that K. japonica can feed on were explored. Furthermore, the growth and ingestion rates of K. japonica on the dinoflagellate Akashiwo sanguinea-a suitable algal prey item-heterotrophic bacteria, and the cyanobacteria Synechococcus sp., as a function of prey concentration were determined. Among the prey tested, K. japonica ingested heterotrophic bacteria, Synechococcus sp., the prasinophyte Pyramimonas sp., the cryptophytes Rhodomonas salina and Teleaulax sp., the raphidophytes Heterosigma akashiwo and Chattonella ovata, the dinoflagellates Heterocapsa rotundata, Amphidinium carterae, Prorocentrum donghaiense, Alexandrium minutum, Cochlodinium polykrikoides, Gymnodinium catenatum, A. sanguinea, Coolia malayensis, and the ciliate Mesodinium rubrum, however, it did not feed on the dinoflagellates Alexandrium catenella, Gambierdiscus caribaeus, Heterocapsa triquetra, Lingulodinium polyedra, Prorocentrum cordatum, P. micans, and Scrippsiella acuminata and the diatom Skeletonema costatum. Many K. japonica cells attacked and ingested a prey cell together after pecking and rupturing the surface of the prey cell and then uptaking the materials that emerged from the ruptured cell surface. Cells of A. sanguinea supported positive growth of K. japonica, but neither heterotrophic bacteria nor Synechococcus sp. supported growth. The maximum specific growth rate of K. japonica on A. sanguinea was 1.01 d -1 . In addition, the maximum ingestion rate of K. japonica for A. sanguinea was 0.13ngC predator -1 d -1 (0.06 cells predator -1 d -1 ). The maximum ingestion rate of K. japonica for heterotrophic bacteria was 0.019ngC predator -1 d -1 (266 bacteria predator -1 d -1 ), and the highest ingestion rate of K. japonica for Synechococcus sp. at the given prey concentrations of up to ca. 10 7 cells ml -1 was 0.01ngC predator -1 d -1 (48 Synechococcus predator -1 d -1 ). The maximum daily carbon acquisition from A. sanguinea, heterotrophic bacteria, and Synechococcus sp. were 307, 43, and 22%, respectively, of the body carbon of the predator. Thus, low ingestion rates of K. japonica on heterotrophic bacteria and Synechococcus sp. may be responsible for the lack of growth. The results of the present study clearly show that K. japonica is a predator of diverse phytoplankton, including toxic or harmful algae, and may also affect the dynamics of red tides caused by these prey species., (Copyright © 2017 Elsevier B.V. All rights reserved.)

This study confirms the presence of the toxigenic benthic dinoflagellates Gambierdiscus belizeanus and Ostreopsis spp. in the central Red Sea. To our knowledge, this is also the first report of these taxa in coastal waters of Saudi Arabia, indicating the potential occurrence of ciguatera fish poisoning (CFP) in that region. During field investigations carried out in 2012 and 2013, a total of 100 Turbinaria and Halimeda macroalgae samples were collected from coral reefs off the Saudi Arabian coast and examined for the presence of Gambierdiscus and Ostreopsis, two toxigenic dinoflagellate genera commonly observed in coral reef communities around the world. Both Gambierdiscus and Ostreopsis spp. were observed at low densities (<200 cells g -1 wet weight algae). Cell densities of Ostreopsis spp. were significantly higher than Gambierdiscus spp. at most of the sampling sites, and abundances of both genera were negatively correlated with seawater salinity. To assess the potential for ciguatoxicity in this region, several Gambierdiscus isolates were established in culture and examined for species identity and toxicity. All isolates were morphologically and molecularly identified as Gambierdiscus belizeanus. Toxicity analysis of two isolates using the mouse neuroblastoma cell-based assay for ciguatoxins (CTX) confirmed G. belizeanus as a CTX producer, with a maximum toxin content of 6.50±1.14×10 -5 pg P-CTX-1 eq. cell -1 . Compared to Gambierdiscus isolates from other locations, these were low toxicity strains. The low Gambierdiscus densities observed along with their comparatively low toxin contents may explain why CFP is unidentified and unreported in this region. Nevertheless, the presence of these potentially toxigenic dinoflagellate species at multiple sites in the central Red Sea warrants future study on their possible effects on marine food webs and human health in this region., (Copyright © 2017 Elsevier B.V. All rights reserved.)

Bysmatrum subsalsum is a cosmopolitan dinoflagellate species that inhabits marine and transitional habitats. Despite its wide distribution, information on the morphological variability, phylogeny and ecology of B. subsalsum is scarce. In this study, we provide morphological and molecular data on B. subsalsum strains and wild cells from different locations in the Mediterranean Basin. The dynamics of cell abundances and the associated environmental conditions during a field bloom are also described. Genetic sequences of B. subsalsum obtained in this study showed large intraspecific differences, clustering in two well-differentiated clades. Despite a certain degree of variation with respect to cell size, apical pore complex (APC) morphology and size, and cingulum displacement, cells from the two clades showed similar morphological traits. These findings indicated the occurrence of cryptic species. Comparisons of the morphology of our B. subsalsum specimens with the few descriptions available in the literature revealed larger than previously known intraspecific morphological variability. Phylogenetic trees inferred from the concatenated SSU, 5.8S-ITS, and LSU rRNA and the individual 5.8S-ITS regions suggested the inclusion of Bysmatrum in the Peridiniales and a close phylogenetic relationship with Peridinium sensu stricto. However, the low statistical support prevented the assignment of Bysmatrum to a particular family of Peridiniales. Ecological data obtained from a bloom in La Pletera salt marshes (Catalan Coast, Spain) suggested the species reaches high cell abundances at water temperatures >20°C and salinity levels >30. Our results add new information regarding the morphology, phylogeny, and ecology of B. subsalsum., (© 2017 Phycological Society of America.)

Recent molecular phylogenetic studies of Gambierdiscus species flagged several new species and genotypes, thus leading to revitalizing its systematics. The inter-relationships of clades revealed by the primary sequence information of nuclear ribosomal genes (rDNA), however, can sometimes be equivocal, and therefore, in this study, the taxonomic status of a ribotype, Gambierdiscus sp. type 6, was evaluated using specimens collected from the original locality, Marakei Island, Republic of Kiribati; and specimens found in Rawa Island, Peninsular Malaysia, were further used for comparison. Morphologically, the ribotype cells resembled G. scabrosus, G. belizeanus, G. balechii, G. cheloniae and G. lapillus in thecal ornamentation, where the thecal surfaces are reticulate-foveated, but differed from G. scabrosus by its hatchet-shaped Plate 2', and G. belizeanus by the asymmetrical Plate 3'. To identify the phylogenetic relationship of this ribotype, a large dataset of the large subunit (LSU) and small subunit (SSU) rDNAs were compiled, and performed comprehensive analyses, using Bayesian-inference, maximum-parsimony, and maximum-likelihood, for the latter two incorporating the sequence-structure information of the SSU rDNA. Both the LSU and SSU rDNA phylogenetic trees displayed an identical topology and supported the hypothesis that the relationship between Gambierdiscus sp. type 6 and G. balechii was monophyletic. As a result, the taxonomic status of Gambierdiscus sp. type 6 was revised, and assigned as Gambierdiscus balechii. Toxicity analysis using neuroblastoma N2A assay confirmed that the Central Pacific strains were toxic, ranging from 1.1 to 19.9 fg P-CTX-1 eq cell -1 , but no toxicity was detected in a Western Pacific strain. This suggested that the species might be one of the species contributing to the high incidence rate of ciguatera fish poisoning in Marakei Island., (Copyright © 2017 Elsevier B.V. All rights reserved.)

Reef-building corals form symbiotic relationships with dinoflagellates of the genus Symbiodinium. Symbiodinium are genetically and physiologically diverse, and corals may be able to adapt to different environments by altering their dominant Symbiodinium phylotype. Notably, each coral species associates only with specific Symbiodinium phylotypes, and consequently the diversity of symbionts available to the host is limited by the species specificity. Currently, it is widely presumed that species specificity is determined by the combination of cell-surface molecules on the host and symbiont. Here we show experimental evidence supporting a new model to explain at least part of the specificity in coral-Symbiodinium symbiosis. Using the laboratory model Aiptasia-Symbiodinium system, we found that symbiont infectivity is related to cell size; larger Symbiodinium phylotypes are less likely to establish a symbiotic relationship with the host Aiptasia. This size dependency is further supported by experiments where symbionts were replaced by artificial fluorescent microspheres. Finally, experiments using two different coral species demonstrate that our size-dependent-infection model can be expanded to coral-Symbiodinium symbiosis, with the acceptability of large-sized Symbiodinium phylotypes differing between two coral species. Thus the selectivity of the host for symbiont cell size can affect the diversity of symbionts in corals.

The marine dinoflagellate Erythropsidinium possesses an ocelloid, the most elaborate photoreceptor organelle known in a unicellular organism, and a piston, a fast contractile appendage unknown in any other organism. The ocelloid is able to rotate, often before the cell swims. The ocelloid contains lenses that function to concentrate light. The flagellar propulsion is atrophied, and the piston is responsible for locomotion through successive extensions and contractions. During the "locomotion mode", the contraction is ~4 times faster than the extension. The piston attained up to 50 mm · s -1 and the cell jumps backwards at -4 mm · s -1 , while during the piston extension the cell moves forwards. The net speed of ~-1 mm · s -1 is faster than other dinoflagellates. The piston usually moved in the "static mode" without significant cell swimming. This study suggests that the piston is also a tactile organelle that scans the surrounding waters for prey. Erythropsidinium feeds on copepod eggs by engulfing. The end of the piston possesses a "suction cup" able to attach the prey and place it into the posterior cavity for engulfing. The cylindrical shape of Erythropsidinium, and the anterior position of the ocelloid and nucleus, are morphological adaptations that leave space for the large vacuole. Observations are provided on morphological development during cell division. Most of the described species of Erythropsidinium apparently correspond to distinct life stages of known species, and the genus Greuetodinium (=Leucopsis) corresponds to an earlier division stage., (© 2017 Phycological Society of America.)

Blooms of the harmful dinoflagellate Akashiwo sanguinea are responsible for the mass mortality of fish and invertebrates in coastal waters. This cosmopolitan species includes several genetically differentiated clades. Four clonal cultures were established by isolating single cells from Xiamen Harbour (the East China Sea) for morphological and genetic analyses. The cultures displayed identical morphology but were genetically different, thus revealing presence of cryptic diversity in the study area. New details of the apical structure complex of Akashiwo sanguinea were also found. To investigate whether the observed cryptic diversity was related to environmental differentiation, 634 cells were obtained from seasonal water samples collected from 2008 to 2012. These cells were sequenced by single-cell PCR. For comparison with Chinese material, additional large subunit ribosomal DNA sequences were obtained for three established strains from Malaysian and French waters. To examine potential ecological differentiation of the distinct genotypes, growth responses of the studied strains were tested under laboratory conditions at temperatures of 12°C to 33°C. These experiments showed four distinct ribotypes of A. sanguinea globally, with the ribotypes A and B co-occuring in Xiamen Harbour. Ribotype A of A. sanguinea was present year-round in Xiamen Harbour, but it only bloomed in the winter and spring, thus corresponding to the winter type. In contrast, A. sanguinea ribotype B bloomed only in the summer, corresponding to the summer type. This differentiation supports the temperature optimum conditions that were established for these two ribotypes in the laboratory. Ribotype A grew better at lower temperatures compared to ribotype B which preferred higher temperatures. These findings support the idea that various ribotypes of A. sanguinea correspond to distinct ecotypes and allopatric speciation occurred in different climatic regions followed by dispersal., (Copyright © 2017 Elsevier B.V. All rights reserved.)

Species within the marine toxic dinoflagellate genus Dinophysis are phagotrophic organisms that exploit chloroplasts (kleptochloroplasts) from other protists to perform photosynthesis. Dinophysis spp. acquire the kleptochloroplasts from the ciliate Mesodinium rubrum, which in turn acquires the chloroplasts from a unique clade of cryptophytes. Dinophysis spp. digest the prey nuclei and all other cell organelles upon ingestion (except the kleptochloroplasts) and they are therefore believed to constantly acquire new chloroplasts as the populations grow. Previous studies have, however, indicated that Dinophysis can keep the kleptochloroplasts active during long term starvation and are able to produce photosynthetic pigments when exposed to prey starvation. This indicates a considerable control over the kleptochloroplasts and the ability of Dinophysis to replicate its kleptochloroplasts was therefore re-investigated in detail in this study. The kleptochloroplasts of Dinophysis acuta and Dinophysis acuminata were analyzed using confocal microscopy and 3D bioimaging software during long term starvation experiments. The cell concentrations were monitored to confirm cell divisions and samples were withdrawn each time a doubling had occurred. The results show direct evidence of kleptochloroplastidic division and that the decreases in total kleptochloroplast volume, number of kleptochloroplasts and number of kleptochloroplast centers were not caused by dilution due to cell divisions. This is the first report of division of kleptochloroplasts in any protist without the associated prey nuclei. This indicates that Dinophysis spp. may be in a transitional phase towards possessing permanent chloroplasts, which thereby potentially makes it a key organism to understand the evolution of phototrophic protists.

Oxyrrhis marina was subjected to conventional transmission electron microscopy, with emphasis being laid on its extrusomes. Mainly regular trichocysts were obvious in ultrathin sections. They were highly abundant, approximately 2 μm in length and 200 nm in width, and composed of the characteristic features, i.e., an anterior tip and the posterior crystalline body. The tip measures approximately 440 nm in length and is built by an outer less electron-dense concentric layer followed by an inner electron-dense core with a translucent center in the middle. The less electron-dense layer most likely ends up in a bundle of filaments which are concentrically placed around the electron-dense core in the transition zone between the tip and body. Trichocyst bodies which are sectioned along the longitudinal axis are approximately 1.5 μm in length and show a regular striation of electron-dense and electron-translucent lines with a spacing of 9 nm. Cross-sectioned bodies are square-shaped and show a crystalline lattice composed of particles which are 8-9 nm in size. Discharge of regular trichocysts results in long rigid rods. They are square-shaped, 54 nm broad, and with a regular striation of approximately 54 nm along their longitudinal axes. Besides regular trichocysts, an additional type of extrusome was registered. It is not as abundant as regular trichocysts, membrane-enclosed, 2 μm in length and 180 nm in width, and resembles two bullets adjacent to each other with the tips facing in opposite directions. The two parts are slightly of different lengths (anterior part, 740 nm; posterior part, 590 nm) and widths (anterior part, 126 nm; posterior part, 117 nm) and separated from each other by a gap of 30 nm. The anterior part is more electron-dense than the posterior one. A faint electron-dense sheet-like structure was registered between the envelope membrane and these two inner structures. In extrusomes which had been arrested in the process of discharge, the anterior part gives rise to an oozing, amorphous, fibrous blob, while the posterior part consists of twisted filaments which most likely function as the charge of a gun for the release of the anterior part.

A small (7-11 μm long) dinoflagellate with thin amphiesmal plates was isolated into culture from a water sample collected in coastal waters of Yeosu, southern Korea, and examined by LM, SEM, and TEM, and molecular analyses. The hemispheric episome was smaller than the hyposome. The nucleus was oval and situated from the central to the episomal region of the cell. A large yellowish-brown chloroplast was located at the end of the hyposome, and some small chloroplasts extended into the periphery of the episome. The dinoflagellate had a single elongated apical vesicle (EAV) and a type E eyespot, which are key characteristics of the family Suessiaceae. Unlike other genera in this family, it had two long furrow lines, one on the episome and the other on the hyposome, and encircling the dorsal, and lateral sides of the cell body. The pyrenoid lacked starch sheaths, but tubular invaginations into the pyrenoid matrix from the cytoplasm were observed. In the TEM, the dinoflagellate was observed to have cable-like structures (CLSs) near the eyespot but so far not observed in other dinoflagellates. The SSU rDNA sequences examined were 1.2%-5.1% different from those of other genera in the family Suessiaceae, whereas the LSU (D1-D3) rDNA sequences of this dinoflagellate were 15.1%-31.5% different. The dinoflagellate lacked a 51-bp fragment in domain D2 of the LSU rDNA, but it had an ~100-bp fragment in domain D2. This feature has been found previously only in the genera Leiocephalium and Polarella, two other genera of the Suessiaceae. The molecular phylogeny and sequence divergence based on SSU, and LSU rDNA indicate that the Korean dinoflagellate holds a taxonomically distinctive position and we consider it to be a new species in a new genus in the family Suessiaceae, named Yihiella yeosuensis gen. et sp. nov., (© 2016 Phycological Society of America.)

Ciguatera fish poisoning (CFP) has been reported for many years in Rarotonga, Cook Islands, and has had the world's highest reported incidence of this illness for the last 20 years. Following intensive sampling to understand the distribution of the causative organisms of CFP, an undescribed Gambierdiscus species was isolated from the Rarotongan lagoon. Gambierdiscus cheloniae sp. nov. has the common Gambierdiscus Kofoidian plate formula (except for a variability in the number of precingular plates in aberrant cells): Po, 3', 6″ (7″), 6C?, 6 or 7S, 5'″, 1p and 2″″. The 2' plate is hatchet shaped and the dorsal end of 1p is pointed and the relatively narrow 1p plate. Morphologically G. cheloniae is similar to the genetically closely related species G. pacificus, G. toxicus and G. belizeanus, although smaller (depth and length) than G. toxicus. The apical pore plate varies from those of G. belizeanus and G. pacificus, which are shorter and narrower, and from G. toxicus, which is larger. G. cheloniae also differs from G. pacificus in the shape of the 2' plate. The description of this new species is supported by phylogenetic analyses using three different gene regions. G. cheloniae produced the putative maitotoxin-3 analogue, MTX-3, but neither maitotoxin or monitored ciguatoxin. Extracts of G. cheloniae were shown to be highly toxic to mice by intraperitoneal (i.p.) injection, although they were less toxic by gavage. It is possible that this species produces toxins other than putative MTX-3., (Copyright © 2016 Elsevier B.V. All rights reserved.)

Harmful benthic dinoflagellates, usually developing in tropical areas, are expanding to temperate ecosystems facing water warming. Reports on harmful benthic species are particularly scarce in the Southern Mediterranean Sea. For the first time, three thermophilic benthic dinoflagellates ( Ostreopsis cf. ovata , Prorocentrum lima and Coolia monotis ) were isolated from Bizerte Bay (Tunisia, Mediterranean) and monoclonal cultures established. The ribotyping confirmed the morphological identification of the three species. Maximum growth rates were 0.59 ± 0.08 d -1 for O. cf. ovata , 0.35 ± 0.01 d -1 for C. monotis and 0.33 ± 0.04 d -1 for P. lima. Toxin analyses revealed the presence of ovatoxin-a and ovatoxin-b in O. cf. ovata cells. Okadaic acid and dinophysistoxin-1 were detected in P. lima cultures. For C. monotis , a chromatographic peak at 5.6 min with a mass m / z = 1061.768 was observed, but did not correspond to a mono-sulfated analogue of the yessotoxin. A comparison of the toxicity and growth characteristics of these dinoflagellates, distributed worldwide, is proposed., Competing Interests: The authors declare no conflict of interest.

Coral bleaching is an important environmental phenomenon, whose mechanism has not yet been clarified. The involvement of reactive oxygen species (ROS) has been implicated, but direct evidence of what species are involved, their location and their mechanisms of production remains unknown. Histidine-mediated chemical trapping and singlet oxygen sensor green (SOSG) were used to detect intra- and extracellular singlet oxygen ((1) O2 ) in Symbiodinium cultures. Inhibition of the Calvin-Benson cycle by thermal stress or high light promotes intracellular (1) O2 formation. Histidine addition, which decreases the amount of intracellular (1) O2 , provides partial protection against photosystem II photoinactivation and chlorophyll (Chl) bleaching. (1) O2 production also occurs in cell-free medium of Symbiodinium cultures, an effect that is enhanced under heat and light stress and can be attributed to the excretion of (1) O2 -sensitizing metabolites from the cells. Confocal microscopy imaging using SOSG showed most extracellular (1) O2 around the cell surface, but it is also produced across the medium distant from the cells. We demonstrate, for the first time, both intra- and extracellular (1) O2 production in Symbiodinium cultures. Intracellular (1) O2 is associated with photosystem II photodamage and pigment bleaching, whereas extracellular (1) O2 has the potential to mediate the breakdown of symbiotic interaction between zooxanthellae and their animal host during coral bleaching., (© 2016 The Authors. New Phytologist © 2016 New Phytologist Trust.)