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The toxic dinoflagellate Karenia mikimotoi is a harmful algal bloom-forming species in coastal areas around the world. It produces ichthyotoxins and hemolytic toxins, with deleterious effects on marine ecosystems. In this study, the bacterium Pseudoalteromonas sp. FDHY-MZ2, with high algicidal efficiency against K. mikimotoi , was isolated from a bloom event. Based on the results, it completely lysed K. mikimotoi cells within 24 h 0.5% (vol/vol), with the algicidal activity of the supernatant of the bacterium culture. Algal cell wall fragmentation occurred, leading to cell death. There was a marked decline in various photochemical traits. When treated with the supernatant, cellulase, pheophorbide a oxygenase (PAO) and cyclin B genes were significantly increased, suggesting induced cell wall deterioration, chloroplast degradation and cell cycle regulation of K. mikimotoi cells. In addition, the expression levels of reactive oxygen species (ROS) scavenging gene was significantly inhibited, indicating that the ROS removal system was damaged. The bacterial culture was dried to obtain the spray-dried powder, which showed algicidal activity rates of 92.2 and 100% against a laboratory K. mikimotoi culture and a field microcosm of Karlodinium sp. bloom within 24 h with the addition of 0.04% mass fraction powder. Our results demonstrate that FDHY-MZ2 is a suitable strain for K. mikimotoi and Karlodinium sp. blooms management. In addition, this study provides a new strategy for the anthropogenic control of harmful algal bloom-forming species in situ . IMPORTANCE K. mikimotoi is a noxious algal bloom-forming species that cause damaging of the aquaculture industry and great financial losses. Bacterium with algicidal activity is an ideal agency to inhibit the growth of harmful algae. In this approach application, the bacterium with high algicidal activity is required and the final management material is ideal for easy-to-use. The algicidal characteristics are also needed to understand the effects of the bacterium for managing strategy exploration. In this study, we isolated a novel algicidal bacterium with extremely high lysis efficiency for K. mikimotoi . The algicidal characteristics of the bacterium as well as the chemical and molecular response of K. mikimotoi with the strain challenge were examined. Finally, the algicidal powder was explored for application. The results demonstrate that FDHY-MZ2 is suitable for K. mikimotoi and Karlodinium sp. blooms controlling, and this study provides a new strategy for algicidal bacterium application.

Harmful algal blooms formed by certain dinoflagellate species often occur when environmental nitrogen nutrients (N) are limited. However, the molecular mechanism by which dinoflagellates adapt to low N environments is poorly understood. In this study, we characterized the transcriptomic responses of Prorocentrum shikokuense to N deficiency, along with its physiological impact. Under N deficiency, P. shikokuense cultures exhibited growth inhibition, a reduction in cell size, and decreases in cellular chlorophyll a and nitrogen contents but an increase in carbon content. Accordingly, gene expression profiles indicated that carbon fixation and catabolism and fatty acid metabolism were enhanced. Transporter genes of nitrate/nitrite, ammonium, urea, and amino acids were significantly upregulated, indicating that P. shikokuense cells invest to enhance the uptake of available dissolved N. Notably, upregulated genes included those involved in endocytosis and phagosomes, evidence that P. shikokuense is a mixotrophic organism that activates phagotrophy to overcome N deficiency. Additionally, vacuolar amino acid transporters, the urea cycle, and urea hydrolysis genes were upregulated, indicating N recycling within the cells under N deficiency. Our study indicates that P. shikokuense copes with N deficiency by economizing nitrogen use and adopting multiple strategies to maximize N acquisition and reuse while maintaining carbon fixation. The remarkable low N adaptability may confer competitive advantages to P. shikokuense for forming harmful blooms in DIN-limited environments., Competing Interests: Declaration of competing interest All the authors declare that there are no conflicts of interest regarding this article., (Copyright © 2020 Elsevier B.V. All rights reserved.)

The availability of ambient N nutrient is often correlated with the occurrences of harmful algal bloom formed by certain dinoflagellates, making it important to understand how these species might be responding to such conditions. Here, transcriptome sequencing of Karenia mikimotoi was conducted to understand the underlying molecular mechanisms by which this dinoflagellate copes with nitrogen (N) deficiency. Transcriptomic analysis revealed 8802 unigenes (3.56%) that were differentially expressed with ≥ 2-fold change. Under N-depleted conditions, genes involved in glycolysis, fatty acid metabolism, and the tricarboxylic acid (TCA) cycle as well as lipid accumulation were significantly upregulated. The elevated expression of enzymes used in protein degradation and turnover suggests possible metabolic reconfiguration towards accelerated N recycling. Moreover, a significant increase in urea transporter was observed, indicating increased assimilation of organic nitrogen resources as an alternative in N-depleted cultures of K. mikimotoi. The down-regulated glutamate synthase genes were also identified under N deficiency, suggesting suppression of primary amino acid synthesis to save N resource. Taken together, results of this study show enhanced multiple N resource acquisition and reuse of multiple N resources constitute a comprehensive strategy to cope with N deficiency in a dinoflagellate., (Copyright © 2021. Published by Elsevier B.V.)

Harmful algal blooms (HABs) have increased as a result of global climate and environmental changes, exerting increasing impacts on the aquatic ecosystem, coastal economy, and human health. Despite great research efforts, our understanding on the drivers of HABs is still limited in part because HAB species' physiology is difficult to probe in situ. Here, we used molecular ecological analyses to characterize a dinoflagellate bloom at Xiamen Harbor, China. Prorocentrum donghaiense was identified as the culprit, which nutrient bioassays showed were not nutrient-limited. Metatranscriptome profiling revealed that P. donghaiense highly expressed genes related to N- and P-nutrient uptake, phagotrophy, energy metabolism (photosynthesis, oxidative phophorylation, and rhodopsin) and carbohydrate metabolism (glycolysis/gluconeogenesis, TCA cycle and pentose phosphate) during the bloom. Many genes in P. donghaiense were up-regulated at night, including phagotrophy and environmental communication genes, and showed active expression in mitosis. Eight microbial defense genes were up-regulated in the bloom compared with previously analyzed laboratory cultures. Furthermore, 76 P. donghaiense microRNA were identified from the bloom, and their target genes exhibited marked differences in amino acid metabolism between the bloom and cultures and the potential of up-regulated antibiotic and cell communication capabilities. These findings, consistent with and complementary to recent reports, reveal major metabolic processes in P. donghaiense potentially important for bloom formation and provide a gene repertoire for developing bloom markers in future research., (Copyright © 2019 Elsevier B.V. All rights reserved.)

Proliferating cell nuclear antigen (PCNA) plays critical roles in eukaryotic DNA replication and replication-associated processes. It is typically encoded by one or two gene copies (pcna) in eukaryotic genomes. Recently reported higher copy numbers of pcna in some dinoflagellates raised a question of how this gene has uniquely evolved in this phylum. Through real-time PCR quantification, we found a wide range of pcna copy number (2-287 copies) in 11 dinoflagellate species (n = 38), and a strong positive correlation between pcna copy number and genome size (log 10 -log 10 transformed). Intraspecific pcna diverged up to 21% and are dominated by nonsynonymous substitutions, indicating strong purifying selection pressure on and hence functional necessity of this gene. By surveying pcna copy numbers in eukaryotes, we observed a genome size threshold at 4 pg DNA, above which more than two pcna copies are found. To examine whether retrotransposition is a mechanism of pcna duplication, we measured the copy number of retroposed pcna, taking advantage of the 22-nt dinoflagellate-specific spliced leader (DinoSL) capping the 5' end of dinoflagellate nuclear-encoded mRNAs, which would exist in the upstream region of a retroposed gene copy. We found that retroposed pcna copy number increased with total pcna copy number and genome size. These results indicate co-evolution of dinoflagellate pcna copy number with genome size, and retroposition as a major mechanism of pcna duplication in dinoflagellates. Furthermore, we posit that the demand of faithful replication and maintenance of the large dinoflagellate genomes might have favored the preservation of the retroposed pcna as functional genes., (© 2018 Phycological Society of America.)

The relationship between algicidal bacteria and harmful-algal-bloom-forming dinoflagellates is understudied and their action modes are largely uncharacterized. In this study, an algicidal bacterium (FDHY-03) was isolated from a bloom of Prorocentrum donghaiense and the characteristics of its action against P. donghaiense was investigated at physiological, molecular, biochemical and cytological levels. 16S rDNA sequence analysis placed this strain in the genus of Alteromonas in the subclass of γ-proteobacteria. Algicidal activity was detected in the bacterial filtrate, suggesting a secreted algicidal principle from this bacterium. Strain FDHY-03 showed algicidal activity on a broad range of HAB-forming species, but the greatest effect was found on P. donghaiense, which showed 91.7% mortality in 24 h of challenge. Scanning electron microscopic analysis indicated that the megacytic growth zone of P. donghaiense cells was the major target of the algicidal action of FDHY-03. When treated with FDHY-03 culture filtrate, P. donghaiense cell wall polysaccharides decreased steadily, suggesting that the algicidal activity occurred through the digestion of cell wall polysaccharides. To verify this proposition, the expression profile of beta-glucosidase gene in FDHY-03 cultures with or without P. donghaiense cell addition was investigated using reverse-transcription quantitative PCR. The gene expression level increased in the presence of P. donghaiense cells, indicative of beta-glucosidase induction by P. donghaiense and the enzyme's role in this dinoflagellate's demise. This study has isolated a new bacterial strain with a strong algicidal capability, documented its action mode and biochemical mechanism, providing a potential source of bacterial agent to control P. donghaiense blooms., (Copyright © 2018 Elsevier B.V. All rights reserved.)

PCP and acpPC are the two major antennae proteins that bind pigments in peridinin-containing dinoflagellates. The relationship between antennae proteins and cellular pigments at molecular level is still poorly understood. Here we identified and characterized the two antennae protein genes in dinoflagellate Prorocentrum donghaiense under different light conditions. The mature PCP protein was 32 kDa, while acpPC was a polyprotein each of 19 kDa. Both genes showed higher expression under low light than under high light, suggesting their possible role in a low light adaptation mechanism. The two genes showed differential diel expression rhythm, with PCP being more highly expressed in the dark than in the light period and acpPC the other way around. HPLC analysis of cellular pigments indicated a diel change of chlorophyll c2, but invariability of other pigments. A stable peridinin: chlorophyll a pigment ratio was detected under different light intensities and over the diel cycle, although the diadinoxanthin:chlorophyll a ratio increased significantly with light intensity. The results suggest that 1) PCP and acpPC genes are functionally distinct, 2) PCP and acpPC can function under low light as an adaptive mechanism in P. donghaiense, 3). the ratios of diadinoxanthin:chlorophyll a and peridinin: chlorophyll a can potentially be used as an indicator of algal photophysiological status and a pigment signature respectively under different light conditions in P. donghaiense., (Copyright © 2018 Elsevier B.V. All rights reserved.)

Although gene regulation can occur at both transcriptional and epigenetic (microRNA) levels, combined transcriptomic and microRNAomic responses to environmental stress are still largely unexplored for marine plankton. Here, we conducted transcriptome and microRNAome sequencing for Prorocentrum donghaiense to understand the molecular mechanisms by which this dinoflagellate copes with phosphorus (P) deficiency. Under P-depleted conditions, G1/S specific cyclin gene was markedly downregulated, consistent with growth inhibition, and genes related to dissolved organic phosphorus (DOP) hydrolysis, carbon fixation, nitrate assimilation, glycolysis, and cellular motility were upregulated. The elevated expression of ATP-generating genes (for example, rhodopsin) and ATP-consuming genes suggests some metabolic reconfiguration towards accelerated ATP recycling under P deficiency. MicroRNAome sequencing revealed 17 microRNAs, potentially regulating 3268 protein-coding genes. Functional enrichment analysis of these microRNA-targeted genes predicted decreases in sulfatide (sulfolipid) catabolism under P deficiency. Strikingly, we detected a significant increase in sulfolipid sulfatide content (but not in sulphoquinovosyldiacylglycerol content) and its biosynthesis gene expression, indicating a different sulfolipid-substituting-phospholipid mechanism in this dinoflagellate than other phytoplankters studied previously. Taken together, our integrative transcriptomic and microRNAomic analyses show that enhanced DOP utilization, accelerated ATP cycling and repressed sulfolipid degradation constitute a comprehensive strategy to cope with P deficiency in a model dinoflagellate.

Dinoflagellates are important components of marine ecosystems and essential coral symbionts, yet little is known about their genomes. We report here on the analysis of a high-quality assembly from the 1180-megabase genome of Symbiodinium kawagutii. We annotated protein-coding genes and identified Symbiodinium-specific gene families. No whole-genome duplication was observed, but instead we found active (retro)transposition and gene family expansion, especially in processes important for successful symbiosis with corals. We also documented genes potentially governing sexual reproduction and cyst formation, novel promoter elements, and a microRNA system potentially regulating gene expression in both symbiont and coral. We found biochemical complementarity between genomes of S. kawagutii and the anthozoan Acropora, indicative of host-symbiont coevolution, providing a resource for studying the molecular basis and evolution of coral symbiosis., (Copyright © 2015, American Association for the Advancement of Science.)

Gene structure and expression regulation of form II RuBisCO (rbcII) in dinoflagellates are still poorly understood. Here we isolated this gene (Pdrbc) and investigated its diel expression pattern in a harmful algal bloom forming dinoflagellate Prorocentrum donghaiense. We obtained cDNA sequences with triple tandem repeats of the coding unit (CU); the 5' region has the sequence of a typical dinoflagellate plastid gene, encoding an N-terminus with two transmembrane regions separated by a plastid transit peptide. The CUs (1,455 bp except 1464 bp in last CU) are connected through a 63 bp spacer. Phylogenetic analysis showed that rbcII CUs within species formed monophyletic clusters, indicative of intraspecific gene duplication or purifying evolution. Using quantitative PCR (qPCR) we estimated 117±40 CUs of Pdrbc in the P. donghaiense genome. Although it is commonly believed that most dinoflagellate genes lack transcriptional regulation, our RT-qPCR analysis on synchronized cultures revealed remarkable diel rhythm of Pdrbc expression, showing significant correlations of transcript abundance with the timing of the dark-to-light transition and cell cycle G2M-phase. When the cultures were shifted to continuous light, Pdrbc expression remained significantly correlated with the G2M-phase. Under continuous darkness the cell cycle was arrested at the G1 phase, and the rhythm of Pdrbc transcription disappeared. Our results suggest that dinoflagellate rbcII 1) undergoes duplication or sequence purification within species, 2) is organized in tandem arrays in most species probably to facilitate efficient translation and import of the encoded enzyme, and 3) is regulated transcriptionally in a cell cycle-dependent fashion at least in some dinoflagellates.