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The harmful jellyfish Cyanea nozakii Kishinouye has frequently occurred on Korean coasts, and its blooms have caused serious ecological and economic damages. DNA sequences of the C. nozakii for molecular detection and discrimination are relatively scarce. In this study, we determined the complete sequence of a single unit of tandemly repeated ribosomal DNA (rDNA) of the Korean C. nozakii and characterized the molecular features of the rDNA. The complete rDNA contained 8,003 bp (48.4% GC) with the same gene arrangement (18S, ITS1, 5.8S, ITS2, 28S, and IGS) to the typical eukaryotes. Dot plot analysis showed that the coding regions (18S, 5.8S, and 28S) were highly conserved, while the non-coding regions (ITS1, ITS2, and IGS) were more variable and parsimony-informative. The IGS contained a putative transcription termination signal (poly(T) tract) and four repeats of block minisatellites. Phylogenetic analyses using 18S and 28S rDNA revealed well-resolved relationships of C. nozakii within the order Semaeostomeae, separating it from other Cyanea species. The complete rDNA sequence provides various options for the selection of jellyfish taxonomic markers and may be useful for discriminating between species of C. nozakii and phylogeny reconstruction with close relatives.

Programmed cell death (PCD) in dinoflagellates has been introduced as a new concept that facilitates the demise of harmful algal blooms. Metacaspases (MCAs) play a role in PCD, but their function in dinoflagellates is unclear. Here, we cloned a novel MCA gene (PmMCA) from the harmful dinoflagellate Prorocentrum minimum and examined its molecular characteristics and gene expression during cell death. The gene was encoded in the nuclear genome with two introns. The putative protein contained 288 amino acids and three conserved MCA signature motifs. Phylogenetic analysis showed that PmMCA may have the same ancestor as other dinoflagellates. PmMCA expression and cell apoptosis were significantly induced under copper exposure, considerably affecting cell growth. These results suggest that PmMCA could be involved in PCD triggered by copper stress.

In this study we characterized the genome scale of the transcriptome of the marine dinoflagellate Alexandrium pacificum under various stresses, focusing on the identification of stress-responsive genes. RNA sequencing generated 18.1 Gb nucleotides that were assembled to 297,808 transcripts. Of them, 207 and 134 transcripts were assigned to the antioxidant enzyme system and heat shock protein (HSP) families, respectively. In addition, the carotenoid biosynthesis pathway and other typical stress-responsive genes (e.g., cold shock proteins and mitogen-activated protein kinases) were widely expressed under stresses. Phylogenetic analysis revealed that these genes might be acquired from different origins via separate evolutionary events. The A. pacificum CuZnSOD and HSP70 were evidently responded to algicides and metals. These findings clearly suggest that A. pacificum possesses many different stress-resistant genes from different sources, which are used for defense and/or adaptation in diverse environments. These may allow the organism to survive and develop when introduced to new environments.

Three strains of Nostoc-related morphotypes isolated from Geum River, Korea. They were studied based on a polyphasic approach using morphology, ecological and molecular data. Phylogenetic analysis of 16S rRNA gene revealed that the selected eight genera were clearly separated into each taxonomic group, and in them, Korean strains formed a clade with Aliinostoc spp. The Aliinostoc sensu lato were monophyletic but divided two distinct lineages with high genetic distance between them (4.2% p-distance between clades vs. 1.8% within clades). In accordance with the statistical analysis, the Korean strain-belonging clade was significantly divergent to segregate from other Aliinostoc clades compared to those of the relative genera (2.8% p-distance between sister-genera vs. 0.9% within the same genus). In addition, the ITS rDNA sequence of the Korean strains showed unique characteristics in terms of nucleotide compositions and length in comparison to that of the Aliinostoc, including their secondary structures of D1–D1ʹ and Box-B. These results supported the establishment of a new genus, proposing Pseudoaliinostoc gen. nov. with a description of P. sejongens sp. nov. for the Korean strains. Likewise, three previously described Aliinostoc species were proposed to be transferred to P. soli comb. nov., P. tiwarii comb. nov., and P. constrictum comb. nov.

Dinoflagellates are frequently exposed to cold-water temperatures, and some may survive in winter and in polar regions. Cold shock domain proteins (CSPs) seem to function as part of their adaptation to such stressful environments. In this study, we identified two CSP genes (PmCSP1 and PmCSP2) in the marine dinoflagellate Prorocentrum minimum, characterized their structural properties, and investigated their expression patterns in response to cold shock and heavy metal exposure. Putative PmCSP1 (96 aa, 10.1 kDa) and PmCSP2 (91 aa, 9.4 kDa) proteins had two consensus RNA-binding motifs and might be localized in the cytoplasm, due to the absence of a signal peptide. Phylogenetic analyses revealed that PmCSPs clustered with those of other dinoflagellates and had been recycled between genomic DNA and mRNA. Real-time PCR revealed that PmCSPs significantly responded to low temperatures, but not to nickel or cadmium. These genes may allow the organism to adapt to and survive in cold habitats and winter.

Heat shock proteins (HSPs) and HSP70-HSP90 organizing proteins (HOPs) are related, and they function together to maintain cellular homeostasis and respond to stress. In the present study, we reported the first molecular characteristics of HSP70 (designated as CeHSP70) and HOP (designated as CeHOP) genes from the freshwater green algae Closterium ehrenbergii and examined the changes in their expression profiles under heat stress and toxic chemicals treatment. CeHSP70 presented the conserved motif patterns and EEVD domain specific to cytosolic HSP70; CeHOP contained a typical domain of TPR repeats. Real-time PCR analysis showed that thermal stress considerably up-regulated both CeHOP and CeHSP70. In addition, the genes were significantly induced by CuCl(2), CuSO(4), and NiSO(4), but not by K(2)Cr(2)O(7), herbicide, and endocrine disrupting chemicals. These results suggest that CeHOP and CeHSP70 function together and play a role in responses to specific stressors and indicate their possible use as sensitive specific biomarkers in risk assessments. ELECTRONIC SUPPLEMENTARY MATERIAL: The online version of this article (10.1007/s12192-020-01143-8) contains supplementary material, which is available to authorized users.

Two types of Wilmottia (Cyanobacteria) were collected from the tree-bark of a tree on Mt. Gwanggyo in Suwon City, Gyeonggi-do and a rock wall of Haje Port, Geum River in Jeollabuk-do, Republic of Korea. Morphological observations using light microscopy (LM) and transmission electron microscopy (TEM) revealed that one species was Wilmottia murrayi (Coleofasciculaceae, Oscillatoriales) and the other had similar morphology to W. murrayi, with considerable genetic variations, and thus, it was described as a new species named Wilmottia koreana. Phylogenetic analyses of the 16S rRNA revealed that the family Coleofasciculaceae was clearly separated into taxonomic groups of each genus, and in them, Wilmottia forms a monophyletic lineage. W. koreana diverged the earliest, forming a unique clade separate from the others. Statistical analyses showed that the inter-species genetic distances of Wilmottia were significantly different from the intra-species distances (t-test, P < 0.001). Additionally, the secondary structures of D1–D1’, Box-B, and the V3 helices were different among the Wilmottia species. These results demonstrated that the proposed new Wilmottia species was unique in molecular traits. Therefore, we propose this to be a new species belonging to the genus Wilmottia with the name Wilmottia koreana sp. nov.

Alachlor is one of the most widely used herbicides and can remain in agricultural soils and wastewater. The toxicity of alachlor to marine life has been rarely studied; therefore, we evaluated the ...

Microalgae, the key components of aquatic environments, are executive natural biotic models for exploring genotoxic effect of heavy metals, irradiation and other external stimuli and the toxicant elimination. In the current scenario, removal of metal contaminants is a significant challenge. Recently, the effective removal of heavy metals from the aquatic environment using microalgae has gained immense intention. However, the concurrent toxicity has limited their use as bio-accumulants to reduce the concentration of heavy metals and promote heavy metal tolerance. Few studies opined that low dose priming with non-ionizing radiations, such as gamma radiation, increased heavy metal tolerance in plant as well as aquatic photosynthetic microalgae. In the present study, we have hypothesized the growth inhibitory physio-chemical properties by cadmium (Cd) in the green algae Chlamydomonas reinhardtii, and analysed the protective role of low-dose gamma radiations priming against Cd induced growth inhibition. In addition, we have specially emphasized on the mechanism of cell survival in the experimental species with effective notation to antioxidant defence system during Cd induced toxicity. Experimentally, we primed the Chlamydomonas reinhardtii with low-dose gamma radiation prior to Cd treatment. On the other hand, gamma-radiated and Cd-treated organisms were considered as positive controls. We calculated the rate of cell death, the deployment of antioxidant enzymes such as catalase (CAT), ascorbate peroxidase (APX), glutathione peroxidase (GPX), glutathione reductase (GSR), and superoxide dismutase (SOD). Furthermore, the role of oxidative stress related genes was analysed computationally to delineate their involvement in cell death/survival, suggesting that the low-dose priming of gamma radiation enhances the Cd tolerance by alterating cell/death pathways and other biochemical responses.

Salinity is an important factor for regulating metabolic processes in aquatic organisms, however, its effects on toxicity and STX biosynthesis gene responses in dinoflagellates require further elucidation. Herein, we evaluated the physiological responses, toxin production, and expression levels of two STX synthesis core genes, sxtA4 and sxtG, in the dinoflagellate Alexandrium pacificum Alex05 under different salinities (20, 25, 30, 35, and 40 psu). Optimal growth was observed at 30 psu (0.12 cell division/d), but cell growth significantly decreased at 20 psu and was irregular at 25 and 40 psu. The cell size increased at lower salinities, with the highest size of 31.5 µm at 20 psu. STXs eq was highest (35.8 fmol/cell) in the exponential phase at 30 psu. GTX4 and C2 were predominant at that time but were replaced by GTX1 and NeoSTX in the stationary phase. However, sxtA4 and sxtG mRNAs were induced, and their patterns were similar in all tested conditions. PCA showed that gene transcriptional levels were not correlated with toxin contents and salinity. These results suggest that A. pacificum may produce the highest amount of toxins at optimal salinity, but sxtA4 and sxtG may be only minimally affected by salinity, even under high salinity stress.

The molecular mechanisms through which dinoflagellates adapt to nitrate fluctuations in aquatic environments remain poorly understood. Here, we sequenced the full-length cDNA of a nitrate transporter (NRT) gene from the harmful marine dinoflagellate Prorocentrum minimum Schiller. The cDNA length was 2431 bp. It encoded a 529-amino acid protein, which was phylogenetically clustered with proteins from other dinoflagellates. Nitrate supply promoted cell growth up to a certain concentration (∼1.76 mM) but inhibited it at higher concentrations. Interestingly, at the inhibitory concentrations, nitrite levels in the medium were considerably increased. Nitrate concentration affected the expression of PmNRT, nitrite transporter (PmNiRT), nitrate reductase (PmNR), and nitrite reductase (PmNiR). Specifically, PmNRT was upregulated after 24 h, with ∼6-fold change compared with the control level, in both nitrate-depleted and nitrate-repleted cultures. In addition, PmNR transcript levels increased to the maximum of 4-fold at 48 h but decreased thereafter. In contrast, PmNiR levels remained unchanged in both nitrate-repleted and nitrate-depleted cultures. Therefore, P. minimum likely copes with nitrate fluctuations in its environment by regulating a set of genes responsible for nitrate uptake.

The collection of Pinocchia daecheonga sp. nov. was performed at the Daecheong Lake in Korea, five strains of which with four clones were investigated through light microscopy, transmission electron microscopy and molecular data comprising the 16S rRNA to 23S rRNA gene. P. daecheonga differed from type species P. polymorpha by absence of the polar aerotopes. Investigated stains and four clones of P. daecheonga turned out to be a sister clade to the P. polymorpha according to the phylogenetic analysis of 16S rRNA gene. In addition, the Pinocchia was clustered with the family Leptolyngbyaceae members, genera Leptothoe and Leptoelongatus. 16S–23S rRNA intergenic transcribed spacer (ITS) region of P. daecheonga was found to be substantially distinct to P. polymorpha in terms of the secondary structure and nucleotide sequence composition, which concludes that the Pinocchia daecheonga isolated from the Daecheong Lake is a unique species due to differences in morphology and genetic traits compared to the relative P. polymorpha.

Superoxide dismutase (SOD) acts as the first line of defense against reactive oxygen species (ROS) within cells. In the present study, we determined two novel CuZnSOD genes (designated as CeCSD1 and CeCSD2) from the toxicity-testing freshwater algae Closterium ehrenbergii and examined their structural features, phylogenetic relationships, and gene expression under exposure to different metals. Putative CeCSD1 (204 aa, 20.6 kDa) and CeCSD2 (155 aa, 15.3 kDa) proteins had conserved CuZnSOD family motifs and metal (Cu, Zn) binding sites, but different N-terminus structures, that is, CeCSD1 has a signal peptide to chloroplasts. Phylogenetic analysis of each protein revealed that C. ehrenbergii was well clustered with other green algae and plants. Real-time PCR results showed that the gene expression obviously increased with heavy metal exposure. In addition, excess copper considerably increased the SOD activity and ROS generation but decreased the photosynthetic efficiency in treated cells. These results suggest that CeCSDs are involved in the antioxidant defense system and can be regarded as potential biomarkers for monitoring metal contaminants in aquatic environments.

In cold water coastal regions, the marine dinoflagellate Alexandrium catenella (Group I) is associated with paralytic shellfish poisoning. Here, we characterized the transcriptome of A. catenella isolated from Korean coastal waters during spring. RNA sequencing generated 15.5 Gb nucleotides that were assembled into 103,362 unique transcripts (unigenes). Upon comparison with known proteins, 53,547 (51.81%) unigenes were successfully annotated in public databases, among which, 28,463 and 35,447 unigenes were assigned to gene ontology categories and evolutionary genealogy of genes with enhanced non-supervised orthologous groups, respectively. Notably, we identified numerous toxin- and cold response-related genes in this species. The conserved dinoflagellate spliced leader (dinoSL) was detected in many unigenes, thereby indicating the occurrence of trans-splicing mRNA processing. Moreover, the identification of second or third dinoSL relicts in a few genes indicate that they have been recycled in the past between the genome and mRNA and are potential sources of genome expansion in this species.

Toxic dinoflagellate Alexandrium spp. produce saxitoxins (STXs), whose biosynthesis pathway is affected by temperature. However, the link between the regulation of the relevant genes and STXs’ accumulation and temperature is insufficiently understood. In the present study, we evaluated the effects of temperature on cellular STXs and the expression of two core STX biosynthesis genes (sxtA4 and sxtG) in the toxic dinoflagellate Alexandrium catenella Alex03 isolated from Korean waters. We analyzed the growth rate, toxin profiles, and gene responses in cells exposed to different temperatures, including long-term adaptation (12, 16, and 20 °C) and cold and heat stresses. Temperature significantly affected the growth of A. catenella, with optimal growth (0.49 division/day) at 16 °C and the largest cell size (30.5 µm) at 12 °C. High concentration of STXs eq were detected in cells cultured at 16 °C (86.3 fmol/cell) and exposed to cold stress at 20→12 °C (96.6 fmol/cell) compared to those at 20 °C and exposed to heat stress. Quantitative real-time PCR (qRT-PCR) revealed significant gene expression changes of sxtA4 in cells cultured at 16 °C (1.8-fold) and cold shock at 20→16 °C (9.9-fold). In addition, sxtG was significantly induced in cells exposed to cold shocks (20→16 °C; 19.5-fold) and heat stress (12→20 °C; 25.6-fold). Principal component analysis (PCA) revealed that low temperature (12 and 16 °C) and cold stress were positively related with STXs’ production and gene expression levels. These results suggest that temperature may affect the toxicity and regulation of STX biosynthesis genes in dinoflagellates.

Temperature may affect the production of saxitoxin (STX) and its derivatives (STXs); however, this is still controversial. Further, STX-biosynthesis gene regulation and the relation of its toxicity with temperature are not clearly understood. In the present study, we evaluated the effects of different temperatures (12 °C, 16 °C, and 20 °C) on the growth, toxin profiles, and expression of two core STX-biosynthesis genes, sxtA and sxtG, in the toxic dinoflagellate Alexandrium pacificum Alex05, isolated from Korean coasts. We found that temperature significantly affected cell growth, with maximum growth recorded at 16 °C, followed by 20 °C and 12 °C. HPLC analysis revealed mostly 12 of STXs from the tested cultures. Interestingly, the contents of STXs increased in the cells cultured at 16 °C and exposed to cold stress, compared to the 20 °C culture and heat stress; however, toxin components were much more diverse under heat stress. These toxin profiles generally matched with the sxtA and sxtG expression levels. Incubation at lower temperatures (12 °C and 16 °C) and exposure to cold stress increased sxtA and sxtG expressions in the cells, whereas heat stress showed little change or downregulated the transcription of both genes. Principal component analysis (PCA) showed low correlation between STXs eq and expressional levels of sxtA and sxtG in heat-stressed cells. These results suggest that temperature might be a crucial factor affecting the level and biosynthesis of STXs in marine toxic dinoflagellates.

The chloroacetanilides are among the most commonly used herbicides worldwide, which contaminate aquatic environments and affect aquatic phototrophs. Their sub-lethal toxicity has been evaluated using freshwater algae; however, the modes of cellular toxicity and levels of toxicity to marine organisms are not fully understood. In the present study, we assessed the cellular and molecular effects of chloroacetanilides on marine phototrophs using the dinoflagellate Prorocentrum minimum and the herbicide metazachlor (MZC). The MZC treatment led to a considerable reduction in cell number and pigment, and the EC50 of MZC was calculated to be 0.647 mg/L. The photosynthetic parameters, Fv/Fm and chlorophyll fluorescence significantly decreased with MZC exposure time in a dose-dependent manner. In addition, MZC significantly induced photosynthesis genes, including PmpsbA, PmpsaA, and PmatpB, and the antioxidant PmGST, but not PmKatG. These findings were well matched to reactive oxygen species (ROS) production in MZC-treated cells. Interestingly, we observed inflated vacuoles, undivided chloroplasts, and breakdown of thylakoid membranes in MZC-treated cells. These results support the hypothesis that MZC severely damages chloroplasts, resulting in dysfunction of the dinoflagellate photosynthesis and possibly marine phototrophs in the environment.

Biomonitoring of phytoplankton communities in freshwater ecosystems is imperative for efficient water quality management. In the present study, we present the seasonal diversity of phytoplankton from the non-reservoir area of the Han River (Korea), assessed using the 18S rRNA amplicon sequencing. Our results uncovered a considerably high eukaryotic diversity, which was predominantly represented by phytoplankton in all the seasons (38–63%). Of these, the diatoms, Cyclostephanos tholiformis, Stephanodiscus hantzschii, and Stephanodiscus sp., were frequently detected in spring and winter. Interestingly, for the first time in the Han River, we detected a large number of operational taxonomic unit (OTU) reads belonging to the naked dinoflagellate Gymnodinium sp., which dominated in autumn (15.8%) and was observed only in that season. Molecular cloning and quantitative real-time polymerase chain reaction (PCR) confirmed the presence of Gymnodinium sp. in the samples collected in 2012 and 2019. Moreover, a comparison of the present data with our previous data from a reservoir area (Paldang Dam) revealed similar patterns of phytoplankton communities. This molecular approach revealed a prospective toxic species that was not detected through microscopy. Collectively, resolving phytoplankton communities at a level relevant for water quality management will provide a valuable reference for future studies on phytoplankton for environmental monitoring.

Microalgae have been attracting attention as feedstock for biorefinery because they have various advantages, such as carbon fixation, high growth rate and high energy yield. The bioactive compounds and lutein contained in microalgae are known to be beneficial for human health, especially eye and brain health. In this study, in order to improve the recovery of bioactive extracts including lutein from Tetraselmis suecica with higher efficiency, an effective solvent was selected, and the extraction parameters such as temperature, time and solid loading were optimized by response surface methodology. The most effective solvent for lutein recovery was identified as 100% methanol, and the optimum condition was determined (42.4 &deg, C, 4.0 h and 125 g/L biomass loading) by calculation of the multiple regression model. The maximum content of recovered lutein was found to be 2.79 mg/mL, and the ABTS radical scavenging activity (IC50) and ferric reducing antioxidant power (FRAP) value were about 3.36 mg/mL and 561.9 &mu, mol/L, respectively. Finally, the maximum lutein recovery from T. suecica through statistical optimization was estimated to be 22.3 mg/g biomass, which was 3.1-fold improved compared to the control group.

The freshwater green algae Closterium is sensitive to water quality, and hence has been suggested as ideal organisms for toxicity testing. In the present study, we evaluated the photosynthetic and biochemical responses of C. ehrenbergii to the common contaminants, coppers. The 72 h median effective concentrations (EC50) of CuSO4 and CuCl2 on the test organism were calculated to be 0.202 mg/L and 0.245 mg/L, respectively. Exposure to both coppers considerably decreased pigment levels and photosynthetic efficiency, while inducing the generation of reactive oxygen species (ROS) in cells with increased exposure time. Moreover, the coppers significantly increased the levels of lipid peroxidation and superoxide dismutase (SOD) activity, even at relatively lower concentrations. These suggest that copper contaminants may exert deleterious effects on the photosynthesis and cellular oxidative stress of C. ehrenbergii, representing its powerful potential in aquatic toxicity assessments.

Metacaspases (MCAs) are cysteine proteases that share sequence homology with caspases, and may play roles in programmed cell death (PCD). In the present study, we identified a novel MCA gene (CpMCA) from the red tide dinoflagellate Cochlodinium polykrikoides, and examined its molecular characteristics and gene expression in response to algicide-induced cell death. CpMCA cDNA is 1164 bp in length, containing a dinoflagellate spliced leader sequence (dinoSL), an 879-bp open reading frame (ORF), which codes for a 293-aa protein, and a poly (A) tail. Multi-sequence comparison indicated that CpMCA belongs to type I MCA, but it has a different structure at the N-terminal. Phylogenetic analysis showed that C. polykrikoides may have acquired the MCA gene from bacteria by means of horizontal gene transfer (HGT). In addition, expressions of CpMCA significantly increased following exposure to the common algicides copper sulfate and oxidizing chlorine, which trigger cell death in dinoflagellates, suggesting that CpMCA may be involved in cell death.

The marine green microalga, Tetraselmis suecica, is an important food source for aquaculture, lipid source for biofuel production, and a potential model organism for toxicity assays because of its rapid growth and ability to produce useful chemicals. In order to gain molecular toxicogenomic insights, we determined expressed sequence tags (ESTs) of T. suecica by pyrosequencing, and attained 741 K reads, including 290 Mb of cDNA information. Upon data processing, 24,651 contigs and 19,072 non-overlapping fragments were acquired and deposited to the NCBI non-redundant and gene ontology databases. Of these, 11,292 contigs and 1848 singletons were annotated. From the EST data, we found that many previously identified stress-responsive protein-coding genes were included. The data were further investigated with a focus on heat shock protein (HSP) gene families, with most characterized HSP genes present in our ESTs. In addition, the expression of HSP70 and HSP90 was significantly increased after T. suecica cells were exposed to metal contaminants. This work expands our understanding of stress-related genomics of T. suecica and further evaluates a potential use of oxidative stress-related genes as biomarkers, in particular HSP involvement in environmental genomics and marine toxicological assessment.

In this study, we sequenced and analyzed the complete mitochondrial genome (mtgenome) of the hydrozoan jellyfish Turritopsis lata. The mtgenome was a complete linear form (15,047 bp in length, 30.9% A, 42.1% T, 12.5% C, and 14.5% G), including 13 protein coding genes (PCGs) (cox1, cox2, cox3, atp6, atp8, nad1, nad2, nad3, nad4, nad4L, nad5, nad6, and cytb), 2 tRNAs (tRNAMet and tRNATrp), and 2 rRNAs (12S and 16S rRNA). The genome structure of the T. lata was completely identical to those of other species within the subclass Hydroidolina. In addition, our molecular phylogenetic analysis using 13 PCGs within hydrozoans showed that T. lata was the closest to Turritopsis dohrnii.

In this study, we analyzed the complete mitochondrial genome of the hydrozoan jellyfish Blackfordia virginica. The genome was a linear form (15,109 bp long, 73.6% AT), including 13 protein-coding genes (cox2, atp8, atp6, cox3, nad2, nad5, nad6, nad3, nad4L, nad1, nad4, cytB, and cox1), 2 tRNAs (tRNA-Met and tRNA-Trp), and 2 rRNAs (12S and 16S RNA). The genome structure of the B. virginica was completely identical to mitochondrial genomes of other hydrozoans that belonged to Leptothecata and Anthoathecata. Molecular phylogenetic analysis within hydrozoan species showed that B. virginica was the closest to the hydrozoan Laomedea flexuosa.

Red tides caused by the marine dinoflagellate Cochlodinium polykrikoides Margalef pose significant environmental problems worldwide. Recently, the existence of severe blooms attributable to a single Cochlodinium Schutt species has been questioned by many researchers. Herein we investigated the dinoflagellate composition of harmful algal blooms (HABs) attributed to C. polykrikoides in Korean coastal waters at nine different stations (St.). The component species of Cochlodinium blooms were examined by using microscopic and gene-cloning methods. In the nine study areas, C. polykrikoides was the predominant species of HABs in St. 2, 4, 7, and St. 9. Based on the morphological identification, the bloom was initially thought to be caused only by C. polykrikoides; however, we detected additional bloom-forming dinoflagellates (Polykrikos schwartzii Butschli and Polykrikos kofoidii Chatton), and diatoms (Pseudo-nitzschia americana (Hasle) Fryxell) along with C. polykrikoides. The parasitic dinoflagellates Amoebophrya Koeppen and Euduboscquella Coats, Bachvaroff & Delwiche were found to be co-located with Cochlodinium in our study, and for the first time, Cochlodinium fulvescens Iwataki, Kawami & Matsuoka was detected in Korea (west coast). These results suggest co-existence of multiple dinoflagellates in bloom populations of Cochlodinium and describe the composition of other dinoflagellate blooms (e.g., Polykrikos spp.) in Korean coastal regions. This co-occurrence may be considered during efforts to monitor and control HABs.

We firstly described a box jellyfish, Carybdea brevipedalia collected from the southern coasts of Korea. It is morphologically characterized by gastric phacellae, a special digestive system of cubozoan jellyfish, and velarium, the thin muscle flap forming the opening of the subumbrellar cavity. The phacellae are linear-shaped, comprising numerous cirri branched from multiple roots. Each root also has multiple numbers of cirrus bundles. Basis of velarial cannels parts into two branches in each octant of velarium. Its geographic distributions were limited to Jejudo and the middle-southern coasts where the organisms are seriously hazardous to bathers in summer. Numerous individuals and/or large-sized populations were observed specifically from beaches at Jeju-do, Namhae-do and a small fishing port of Namildae, while only a small amount of the individuals was observed in Gamak Bay, Jaran- Goseung Bay, and Geoje-do. We confirmed molecular identity of the Korean C. brevipedalia with comparison of nuclear ribosomal DNA sequences. Until now, Carybdea brevipedalia is the only cubozoan species reported to be distributed in Korean waters.

The marine dinoflagellateCochlodinium polykrikoidesis one of the most common ichthyotoxic species that causes harmful algal blooms (HABs), which leads to ecological damage and huge economic loss in aquaculture industries. Cyclophilins (CYPs) belong to the immunophilin superfamily, and they may play a role in the survival mechanisms of the dinoflagellate in stress environments. In the present study, we identified a novel cyclophilin gene fromC. polykrikoidesand examined physiological and gene transcriptional responses to biocides copper sulphate (CuSO4) and sodium hypochlorite (NaOCl). The full length ofCpCYPwas 903 bp, ranging from the dinoflagellate splice leader (DinoSL) sequence to the polyA tail, comprising a 639 bp ORF, a 117 bp 5′-UTR, and a 147 bp 3′-UTR. Motif and phylogenetic comparisons showed that CpCYP was affiliated to group B of CYP. In biocide stressors, cell counts, chlorophylla, and photosynthetic efficiency (Fv/Fm) ofC. polykrikoideswere considerably decreased in both exposure time- and dose-dependent manners. In addition,CpCYPgene expression was significantly induced after 24 h exposure to the biocide-treated stress conditions. These results indicate an effect of the biocides on the cell physiology and expression profile ofCpCYP, suggesting that the gene may play a role in environmental stress responses.

In the present study, we sequenced and analyzed the complete mitochondrial genome of the hydrozoan jellyfish Spirocodon saltatrix. The mitochondrial genome was a linear form (15,752 bp long, 70.4% AT), consisting of 13 protein coding genes (cox1, cox2, atp8, atp6, cox3, nad2, nad5, nad6, nad3, nad4L, nad1, nad4, and cytB), two tRNAs (tRNA-Met and tRNA-Trp), and two rRNAs (12S and 16S). Mitochondrial gene arrangement of the S. saltatrix was completely identical to already-known mitochondrial genomes of hydrozoans. Molecular phylogenetic analysis using 13 protein-coding genes showed that S. saltatrix was closely related to the hydrozoan Clava multicornis.

In this study, we sequenced the complete mitochondrial genome of the jellyfish Aurelia coerulea (A. coerulea). The mitochondrial genome was a linear form (16,748 bp long, 65.7% AT), consisting of 1...

The marine dinoflagellate Cochlodinium polykrikoides is a harmful algal bloom (HAB) species that severely impacts the environment and causes huge economic losses. Yellow clay (YC), considered to be a non-toxic and naturally-occurring material, represents an important step towards the direct control of HABs. In the present study, we evaluated the physiological and biochemical effects of YC on C. polykrikoides after exposures of up to 72 h. We observed little physiological changes in growth rate, chlorophyll a, lipid peroxidation, antioxidant enzymatic activities of superoxide dismutase and catalase, and activity of alkaline phosphatase after exposure to YC. Interestingly, YC significantly increased total carbohydrate and glutathione levels, affecting the physiology of the cells. These results indicate that total carbohydrate content may play an important role in cell–clay aggregation and it could be the main underlying mechanism that mitigates HAB cells via sedimentation.

Dinoflagellate algae are microeukaryotes that have distinct genomes and gene regulation systems, making them an interesting model for studying protist evolution and genomics. In the present study, we discovered a novel manganese superoxide dismutase (PmMnSOD) gene from the marine dinoflagellate Prorocentrum minimum, examined its molecular characteristics, and evaluated its transcriptional responses to the oxidative stress-inducing contaminants, CuSO4 and NaOCl. Its cDNA was 1238 bp and contained a dinoflagellate spliced leader sequence, a 906 bp open reading frame (301 amino acids), and a poly (A) tail. The gene was coded on the nuclear genome with one 174 bp intron; signal peptide analysis showed that it might be localized to the mitochondria. Real-time PCR analysis revealed an increase in gene expression of MnSOD and SOD activity when P. minimum cells were separately exposed to CuSO4 and NaOCl. In addition, both contaminants considerably decreased chlorophyll autofluorescence, and increased intracellular reactive oxygen species. These results suggest that dinoflagellate MnSOD may be involved in protecting cells against oxidative damage.

Bacteria are remarkably associated with the growth of green algae Tetraselmis which are used as a feed source in aquaculture, but Tetraselmis-associated bacterial community is characterized insufficiently. Here, as a first step towards characterization of the associated bacteria, we investigated the community composition of free-living (FLB) and particle-associated (PAB) bacteria in each growth phase (lag, exponential, stationary, and death) of Tetraselmis suecica P039 culture using pyrosequencing. The percentage of shared operational taxonomic units (OTUs) between FLB and PAB communities was substantially high (≥92.4%), but their bacterial community compositions were significantly (p = 0.05) different from each other. The PAB community was more variable than the FLB community depending on the growth phase of T. suecica. In the PAB community, the proportions of Marinobacter and Flavobacteriaceae were considerably varied in accordance with the cell number of T. suecica, but there was no clear variation in the FLB community composition. This suggests that the PAB community may have a stronger association with the algal growth than the FLB community. Interestingly, irrespective of the growth phase, Roseobacter clade and genus Muricauda were predominant in both FLB and PAB communities, indicating that bacterial communities in T. suecica culture may positively affect the algae growth and that they are potentially capable of enhancing the T. suecica growth.

Calmodulin (CaM) is one of the major Ca2+-binding proteins in the cells, and it plays multiple roles in several Ca2+ signaling pathways and regulating the activities of other proteins. In the present study, we characterized CaM genes from the marine dinoflagellates Amphidinium carterae, Cochlodinium polykrikoides, Prorocentrum micans, and P. minimum, and examined their expression patterns upon the addition and chelation of calcium. Their cDNAs had same ORF length (450 bp) and encoded the same protein, but with few nucleotide differences in the ORF and different 3'- and 5' untranslated regions (UTRs). The four CaM proteins consist of four EF-hand Ca2+-binding motifs, two N-terminal domains and two C-terminal domains, and they were highly conserved within eukaryotes. The CaM gene expressions in the tested species increased by calcium treatments; however, they were significantly down-regulated by the calcium-chelator EGTA. The CaM genes of the test species were inducible and regulated by different calcium doses, suggesting their major role in calcium regulation in dinoflagellates.

The nomenclature of the sea nettle jellyfish from Korea was initially described as Dactylometra quinquecirrha Agassiz, 1862. However its identity has been questioned on the basis of its local distribution and molecular data. Here, we examined morphology and DNA sequences of nuclear rDNA using specimens collected from southern Korean waters in August 2014. Based on morphological characteristics (bell size, umbrella pattern, number of tentacles and lappets) and distribution locality, we reassign the Korean D. quinquecirrha to Chrysaora pacifica (Goette, 1886), and provide a re-description accordingly. The molecular identity of C. pacifica was further confirmed by comparison of nuclear ribosomal DNA sequences.

Chlorine (Cl2) is widely used as a disinfectant in water treatment plants and for cleaning swimming pools; it is finally discharged into aquatic environments, possibly causing damage to the non-target organisms in the receiving water bodies. Present study evaluated the effects of the biocide Cl2 to the green alga Closterium ehrenbergii (C. ehrenbergii). Growth rate, chlorophyll a levels, carotenoids, chlorophyll autofluorescence, and antioxidant enzymes were monitored up to 72-h after Cl2 exposure. C. ehrenbergii showed dose-dependent decrease in growth rate and cell division after exposure to Cl2. By using cell counts, the median effective concentration (EC50)-72-h was calculated to be 0.071mgL(-1). Cl2 significantly decreased the pigment levels and chlorophyll autofluorescence intensity, indicating that the photosystem was damaged in C. ehrenbergii. In addition, it increased the production of reactive oxygen species (ROS) in the cells. This stressor significantly increased the activities of antioxidant enzymes, including superoxide dismutase (SOD), catalase, and glutathione, and affected the physiology of the cells. These results indicate that Cl2 induces oxidative stress in the cellular metabolic process and leads to physiological and biochemical damages in the green algae. Cl2 discharged in industrial effluents and from water treatment plants may cause harmful effects to the C. ehrenbergii a common freshwater microalgae and other non-target organisms.

Most scyphomedusae jellyfishes recorded in Korean waters are temperate and subtropical species. In the present study, two cold-water jellyfishes from Korean waters are first described. Scyphomedusae were collected from the coasts of Gangneung and Ulsan of the East Sea, and Boryeong, and the eastern area of the Yellow Sea from June 2006 to May 2015. Scyphomedusae collected in Ulsan and Gangneung were identified morphologically as Aurelia limbata Brandt, 1835. Their umbrella was 250-500 mm in width and 30-50 mm in height; the exumbrella was white and the subumbrella was dark brown in color. The vascular system was complex with anastomosed branches. Individuals of this species lived at about 6.6-9.9°C and 30-50 m depth in Korean waters. Scyphomedusae collected from the center of the Yellow Sea and from Boryeong were identified as Parumbrosa polylobata Kishinouye, 1910. Their umbrella was disc-shaped, 80-200 mm in width and 20-40 mm in height. Its milky white gonads could be seen through the transparent exumbrella. The stomach cavity was round and flat with four interradial cavities connected by subgenital cavities, and the four oral arms were bifurcated and spear-head shaped. Individuals were found at about 6.4-10.0°C and 40-90 m depth in Korean waters. The identities of both species were confirmed by molecular analysis using nuclear ribosomal DNA sequences.

Microalgae are the potential bioindicators of environmental changes, for the environmental risk assessment as well as to set limits for toxic chemical release in the aquatic environment. Here, we evaluated the effects of two endocrine disrupting chemicals (EDCs), namely bisphenol A (BPA) and Aroclor 1016, on the green algae Tetraselmis suecica, diatom Ditylum brightwellii, and dinoflagellate Prorocentrum minimum. Each species showed wide different sensitivity ranges when exposed to these two EDCs; the 72 h effective concentration (EC 50 ) for these test species showed that Aroclor 1016 was more toxic than BPA. EC 50 values for the diatom D. birghtwellii were calculated at 0.037 mg/L for BPA and 0.002 mg/L for Aroclor 1016, representing it was the most sensitive when compared to the other species. In addition, these results suggest that these EDC discharge beyond these concentrations into the aquatic environments may cause harmful effect to these marine species.

Chlorine (Cl2) is a commonly used biocide; however, the cellular mechanism and pathways associated with Cl2 exposure in algae have not been elucidated. Here, we investigated the physiological and gene transcriptional responses in the dinoflagellate Prorocentrum minimum upon chlorination, with emphasis on the photosynthetic system. The time-course stress response of P. minimum was assessed by measuring transcription of photosynthesis-related genes, chlorophyll autofluorescence (CAF), and photosynthesis efficiency (F v /F m ), in response to a series of Cl2 doses (0.1, 0.5, and 1.0 mg L−1). CAF data showed a marked reduction in fluorescence intensity with increasing Cl2 doses and exposure time (0, 6, 12, and 24 h). F v /F m dramatically decreased after 1 mg L−1 Cl2 exposure. Transcriptional responses of the photosystem related genes (PmpsbA, PmatpB, PmrbcL, and PmpsaA) were analysed using quantitative real-time (qRT)-PCR. PmpsbA transcription was significantly upregulated in 0.5 and 1.0 mg L−1 of Cl2 after a 24 h exposure. However, the transcriptional expressions of PmatpB and PmpsaA were downregulated. Our results suggest that photosynthesis-related genes such as PmpsbA may act as indicators of environmental stress response and can be useful and rapid biomarker candidates for ecotoxicogenomic assessments.

In the present study, we characterized the potential toxin genes for polyketide synthase (PKS) and saxitoxin (STX) biosynthesis using the transcriptomes of two non-STX producing dinoflagellates Amphidinium carterae and Prorocentrum micans. RNA sequencing revealed 94 and 166 PKS contigs in A. carterae and P. micans, respectively. We first detected type III PKS, which was closely related to bacteria. In addition, dozens of homologs of 20 STX biosynthesis genes were identified. Interestingly, the core STX-synthesizing genes sxtA and sxtB were only found in P. micans, whereas sxtD was detected in A. carterae alone. Bioinformatic analysis showed that the first two core genes (sxtA and sxtG) had a low sequence similarity (37.0-67.6%) and different domain organization compared to those of other toxigenic dinoflagellates, such as Alexandrium pacificum. These might result in the breakdown of the initial reactions in STX production and ultimately the loss of the ability to synthesize the toxins in both dinoflagellates. Our findings suggest that toxin-related PKS and sxt genes are commonly found in non-STX producing dinoflagellates. In addition to their involvement in the synthesis of toxins, our result indicates that genes may also have other molecular metabolic functions.

The green microalgae Closterium ehrenbergii is an ideal organism for ecotoxicology assessments; however, its toxicogenomics has been insufficiently examined. Here, we identified three iron/manganese superoxide dismutase (SOD) genes (designated as CeFeSOD1, CeFeSOD2, and CeMnSOD) from C. ehrenbergii and examined their expressional patterns for four metals (iron, manganese, copper, and nickel). These genes encoded 362, 224, and 245 amino acids, respectively; signal-peptide analysis showed that they were differentially located in chloroplasts, cytosol, or mitochondria. Real-time PCRs revealed differential expression patterns according to metal and doses. Interestingly, CeSODs displayed no noticeable changes to treatment with their corresponding cofactor metals, iron or manganese, even at high doses. However, they were obviously up-regulated under toxic metal (copper and nickel) exposure, exhibiting approximately 10.8- and 4.4-fold increases, respectively. Copper (0.2 mg/L) dramatically stimulated intracellular reactive oxygen species (ROS) formation, increased SOD activity, and reduced photosynthetic efficiency in C. ehrenbergii. These results suggest that CeFeSODs and CeMnSOD might be involved in protecting cells against damage and oxidative stress caused by non-cofactor metals, such as copper and nickel. These genes were sensitively responsive at levels well below the EC50, showing that they can be used as molecular biomarkers to assess the toxicity of specific metal contaminants.

In the present study, we extensively characterized potential toxin-related genes, including polyketide synthase (PKS), saxitoxin (STX) and fatty acid synthase (FAS) from the non-toxin producing marine dinoflagellate Cochlodinium polykrikoides, comparing to those of a toxigenic dinoflagellate Alexandrium pacificum. RNA sequencing revealed 50 and 271 PKS contigs from C. polykrikoides and A. pacificum, respectively. According to domain constitute and amino acid alteration, we further classified the dinoflagellate type I PKS genes into 4 sub-groups. Type III PKS was first identified in C. polykrikoides. Interestingly, we detected a large number (242 and 288) of homologs of 18 sxt genes from two studied dinoflagellates. Most of the eight key genes (sxtA, sxtB, sxtD, sxtG, sxtH/T, sxtI, sxtS and sxtU) for STX synthesis were detected in both dinoflatellates, whereas a core STX biosynthesis gene sxtG was not detected in C. polykrikoides. This may partially explain the absence of saxitoxin production in C. polykrikoides. In addition, we identified several type I and type II FAS genes, including FabD, FabF, FabG, FabH, FabI, and FabZ, whereas FabB was not found in C. polykrikoides. Overall, the numbers of the toxin-related genes in C. polykrikoides were less than that of A. pacificum. Phylogenetic analyses showed that type I PKS/FASs of dinoflagellates had close relationships with apicomplexans and bacteria. These suggest that the toxin-related PKS and sxt genes are commonly present in toxigenic and non-toxin producing dinoflagellates, and may be involved not only in the toxin synthesis, but also in other related molecular metabolic functions.

Microalgae are capable of tolerating variations in water temperature and sudden exposures to toxic substances, and cellular heat shock proteins (HSPs) help to protect cells from such stress. Here, we determined the complete open reading frames (ORF) of small TsHSP20 and large TsHSP70 and 100 in the chlorophyte Tetraselmis suecica, and examined the expression levels of these genes after exposure to thermal stressors, redox-active metals, and non-redox-active metals. Putative TsHSP20, TsHSP70, and TsHSP100 proteins had conserved HSP-family motifs with different C-terminus motifs. Phylogenetic analyses of individual HSPs showed that T. suecica clustered well with other chlorophytes. Real-time PCR analysis showed that thermal stress did not significantly change the expression of all the tested TsHSPs. In addition, TsHSP20 showed little gene expression after being exposed to copper, whereas TsHSP70 and 100 genes greatly responded to the redox-active metals in CuSO4 followed by CuCl2, but not to the non-redox active metals. Redox-active metals strongly affected the physiology of the cells, as judged by cell counting, reactive oxygen species imaging and photosynthetic efficiency. These findings suggest that small and large HSPs are differentially involved in the response against environmental stressors. Moreover, metal toxicity may be specifically controlled by the anions in the metal compounds.

Superoxide dismutase (SOD) acts as the first line of defence against reactive oxygen species (ROS) within cells. In this study, we characterized a novel SOD gene (PmCuZnSOD) from the dinoflagellate Prorocentrum minimum, and examined its structural features, putative origin and gene expression. The SOD cDNA is 895 bp in length, containing dinoflagellate splice-leader (dinoSL) sequence, 714-bp ORF (237 aa), and poly (A) tail. In addition, PmCuZnSOD is coded on the dinoflagellate nuclear genome without introns and in a non-tandem repeat manner; however, the encoded protein is probably localized in chloroplasts. Phylogenetic analysis indicated that it might be acquired from cyanobacteria via horizontal gene transfer (HGT) and then the gene possibly relocated from the chloroplast to the nuclear genome. Excess copper dramatically increased the PmCuZnSOD transcripts and SOD activity in cells, caused by ROS generation and decrease of photosynthetic efficiency in the treated cells. These suggest that CuZnSOD may function to defend against oxidative stress for the survival of the dinoflagellate.

In this study, the complete mitogenome of brownbranded moon jellyfish Aurelia limbata (Cnidaria, Semaeostomeae, Ulmaridae) was sequenced and analyzed. The linear mitogenome is 16,953 bp long with 3...

The East China Sea (ECS) has long been considered an important monitoring site for oceanic ecosystem changes because many water currents and river discharges constantly influence this area. In this study, the community structure and diversity of phytoplankton in the northern part of the ECS adjacent to Jeju Island were explored using small subunit ribosomal RNA (SSU) pyrosequencing. We analysed samples collected from four stations from the surface and at 30-m and 50-m depths during April and September 2011. We observed spatial and temporal variations in the phytoplankton community. Among phytoplankton, diatoms and dinoflagellates constituted a major portion at all stations (60–90%). However, comparison of the April and September samples showed seasonal variation and shifts in the dispersion of diatom and dinoflagellates among stations. Among stations, diatoms dominated St. 1 and others were dominated by dinoflagellates. Furthermore, phylotypes of potentially toxin-producing genera such as Karlodinium, Heterocapsa, Gymnodinium, Gyrodinium, and Pseudo-nitzschia were dominant in this area.