Dinoflagellate Proton-Pump Rhodopsin Genes in Long Island Sound: Diversity and Spatiotemporal Distribution.

Microbial proton-pump rhodopsin (PPR)-based phototrophy, a light-harvesting mechanism different from chlorophyll-based photosystems, may contribute significantly to solar energy entry into the marine ecosystem. PPR transforms solar energy into cellular energy that is used for various metabolic processes in the cells or flagellar movement. Although rhodopsins or their encoding genes have been documented in a wide phylogenetic range of cultured dinoflagellates, information is limited about how widespread and how spatiotemporally dynamical dinoflagellate PPR (DiPPR) are in natural marine ecosystems. In this study, we investigated DiPPR in Long Island Sound (LIS), a temperate estuary of the Atlantic Ocean between Connecticut and Long Island, New York, USA. We isolated six novel full-length dinoflagellate proton-pump rhodopsin cDNAs, expanding the DiPPR database that is crucial to PPR research. Based on these new sequences and existing sequences of DiPPR, we designed primers and conducted quantitative PCR and sequencing to determine the abundance and diversity of DiPPR genes spatially and temporally throughout a year in the water samples collected from LIS. DiPPR genes were found year-round and throughout LIS but with higher abundances in the eutrophic Western Sound and in April and July. The gene diversity data suggest that there are at least five distinct rhodopsin-harboring groups of dinoflagellates throughout the year. The abundance of DiPPR genes, measured as copy number per mL of seawater, appeared not to be influenced by water temperature or nitrogen nutrient concentration but exhibited weak negative correlations with orthophosphate concentration and salinity and a positive correlation with the abundance of DiPPR-harboring dinoflagellates. This first quantitative profiling of PPR in natural plankton reveals the prevalence and dynamics of this plastid-independent photoenergy harvesting mechanism in a temperate estuary and provides efficient DiPPR primers potentially useful for future research. Furthermore, this study shed light on the potential role of DiPPR in phosphor nutrition and dinoflagellate population, which warrants further studies.

In situ community transcriptomics illuminates CO2-fixation potentials and supporting roles of phagotrophy and proton pump in plankton in a subtropical marginal sea.

Lineage-wise physiological activities of plankton communities in the ocean are important but challenging to characterize. Here, we conducted whole-assemblage metatranscriptomic profiling at continental shelf and slope sites in the South China Sea to investigate carbon fixation potential in different lineages. RuBisCO expression, the proxy of Calvin carbon fixation (CCF) potential, was mainly contributed by Bacillariophyta, Chlorophyta, Cyanobacteria, and Haptophyta, which was differentially affected by environmental factors among lineages. CCF potential exhibited positive or negative correlations with phagotrophy gene expression, suggesting phagotrophy possibly enhances or complements CCF. Our data also reveal significant non-Calvin carbon fixation (NCF) potential, as indicated by the active expression of genes in all five currently recognized NCF pathways, mainly contributed by Flavobacteriales, Alteromonadales, and Oceanospirillales. Furthermore, in Flavobacteriales, Alteromonadales, Pelagibacterales, and Rhodobacterales, NCF potential was positively correlated with proton-pump rhodopsin (PPR) expression, suggesting that NCF might be energetically supported by PPR. The novel insights into the lineage-differential potential of carbon fixation, widespread mixotrophy, and PPR as an energy source for NCF lay a methodological and informational foundation for further research to understand carbon fixation and the trophic landscape in the ocean.IMPORTANCEMarine plankton plays an important role in global carbon cycling and climate regulation. Phytoplankton and cyanobacteria fix CO2 to produce organic compounds using solar energy and mainly by the Calvin cycle, whereas autotrophic bacteria and archaea may fix CO2 by non-Calvin cycle carbon fixation pathways. How active individual lineages are in carbon fixation and mixotrophy, and what energy source bacteria may employ in non-Calvin carbon fixation, in a natural plankton assemblage are poorly understood and underexplored. Using metatranscriptomics, we studied carbon fixation in marine plankton with lineage resolution in tropical marginal shelf and slope areas. Based on the sequencing results, we characterized the carbon fixation potential of different lineages and assessed Calvin- and non-Calvin- carbon fixation activities and energy sources. Data revealed a high number of unigenes (4.4 million), lineage-dependent differential potentials of Calvin carbon fixation and responses to environmental conditions, major contributors of non-Calvin carbon fixation, and their potential energy source.

Effects and mechanisms of glyphosate as phosphorus nutrient on element stoichiometry and metabolism in the diatom Phaeodactylum tricornutum.

The ability to utilize dissolved organic phosphorus (DOP) gives phytoplankton competitive advantages in P-limited environments. Our previous research indicates that the diatom Phaeodactylum tricornutum could grow on glyphosate, a DOP with carbon-phosphorus (C-P) bond and an herbicide, as sole P source. However, direct evidence and mechanism of glyphosate utilization are still lacking. In this study, using physiological and isotopic analysis, combined with transcriptomic profiling, we demonstrated the uptake of glyphosate by P. tricornutum and revealed the candidate responsible genes. Our data showed a low efficiency of glyphosate utilization by P. tricornutum, suggesting that glyphosate utilization costs energy and that the alga possessed an herbicide-resistant type of 5-enolpyruvylshikimate-3-phosphate (EPSP) synthase. Compared to the P-limited cultures, the glyphosate-grown P. tricornutum cells up-regulated genes involved in DNA replication, cell growth, transcription, translation, carbon metabolism, and many genes encoding antioxidants. Additionally, cellular C and silicon (Si) increased remarkably while cellular nitrogen (N) declined in the glyphosate-grown P. tricornutum, leading to higher Si:C and Si:N ratios, which corresponded to the up-regulation of genes involved in the C metabolism and Si uptake and the down-regulation of those encoding N uptake. This has the potential to enhance C and Si export to the deep sea when P is limited but phosphonate is available. In sum, our study documented how P. tricornutum could utilize the herbicide glyphosate as P nutrient and how glyphosate utilization may affect the element content and stoichiometry in this diatom, which have important ecological implications in the future ocean.IMPORTANCEGlyphosate is the most widely used herbicide in the world and could be utilized as phosphorus (P) source by some bacteria. Our study first revealed that glyphosate could be transported into Phaeodactylum tricornutum cells for utilization and identified putative genes responsible for glyphosate uptake. This uncovers an alternative strategy of phytoplankton to cope with P deficiency considering phosphonate accounts for about 25% of the total dissolved organic phosphorus (DOP) in the ocean. Additionally, accumulation of carbon (C) and silicon (Si), as well as elevation of Si:C ratio in P. tricornutum cells when grown on glyphosate indicates glyphosate as the source of P nutrient has the potential to result in more C and Si export into the deep ocean. This, along with the differential ability to utilize glyphosate among different species, glyphosate supply in dissolved inorganic phosphorus (DIP)-depleted ecosystems may cause changes in phytoplankton community structure. These insights have implications in evaluating the effects of human activities (use of Roundup) and climate change (potentially reducing DIP supply in sunlit layer) on phytoplankton in the future ocean.

Clustered Regularly Interspaced Short Palindromic Repeat/CRISPR-Associated Protein and Its Utility All at Sea: Status, Challenges, and Prospects.

Initially discovered over 35 years ago in the bacterium Escherichia coli as a defense system against invasion of viral (or other exogenous) DNA into the genome, CRISPR/Cas has ushered in a new era of functional genetics and served as a versatile genetic tool in all branches of life science. CRISPR/Cas has revolutionized the methodology of gene knockout with simplicity and rapidity, but it is also powerful for gene knock-in and gene modification. In the field of marine biology and ecology, this tool has been instrumental in the functional characterization of 'dark' genes and the documentation of the functional differentiation of gene paralogs. Powerful as it is, challenges exist that have hindered the advances in functional genetics in some important lineages. This review examines the status of applications of CRISPR/Cas in marine research and assesses the prospect of quickly expanding the deployment of this powerful tool to address the myriad fundamental marine biology and biological oceanography questions.

Glyphosate (Roundup) as phosphorus nutrient enhances carbon and nitrogen accumulation and up-regulates phosphorus metabolisms in the haptophyte Isochrysis galbana.

Inorganic phosphate limitation for phytoplankton may be intensified with water stratification by global warming, and with the increasing nitrogen: phosphorus (N:P) ratio in coastal zones resulting from continuous anthropogenic N overloading. Under these circumstances, phytoplankton's ability to use dissolved organic phosphorus (DOP) will give species a competitive advantage. In our previous study, we have shown that the haptophyte Isochrysis galbana can use glyphosate (Roundup) as a P nutrient source to support growth, but the mechanism of how remains unexplored. Here, we show that three genes encoding PhnC (IgPhnCs), which exhibit up-regulated expression in glyphosate-grown cultures, are probably responsible for glyphosate uptake, while homologs of PhnK and PhnL (IgPhnK and IgPhnL) probably provide auxiliary support for the intracellular degradation of glyphosate. Meanwhile, we found the use efficiency of glyphosate was low compared with phosphate, probably because glyphosate uptake and hydrolysis cost energy and because glyphosate induces oxidative stress in I. galbana. Meanwhile, genes encoding 5-enolpyruvylshikimate 3-phosphate (EPSP) synthase, the target of the herbicide, were up-regulated in glyphosate cultures. Furthermore, our data showed the up-regulation of P metabolisms (transcription) in glyphosate-grown cultures, which further induced the up-regulation of nitrate/nitrite transport and biosynthesis of some amino acids. Meanwhile, glyphosate-grown cells accumulated more C and N, resulting in remarkably high C:N:P ratio, and this, along with the up-regulated P metabolisms, was under transcriptional and epigenetic regulation. This study sheds lights on the mechanism of glyphosate utilization as a source of P nutrient by I. galbana, and these findings have biogeochemical implications.

Novel Plastid Genome Characteristics in Fugacium kawagutii and the Trend of Accelerated Evolution of Plastid Proteins in Dinoflagellates.

Typical (peridinin-containing) dinoflagellates possess plastid genomes composed of small plasmids named "minicircles". Despite the ecological importance of dinoflagellate photosynthesis in corals and marine ecosystems, the structural characteristics, replication dynamics, and evolutionary forcing of dinoflagellate plastid genomes remain poorly understood. Here, we sequenced the plastid genome of the symbiodiniacean species Fugacium kawagutii and conducted comparative analyses. We identified psbT-coding minicircles, features previously not found in Symbiodiniaceae. The copy number of F. kawagutii minicircles showed a strong diel dynamics, changing between 3.89 and 34.3 copies/cell and peaking in mid-light period. We found that F. kawagutii minicircles are the shortest among all dinoflagellates examined to date. Besides, the core regions of the minicircles are highly conserved within genus in Symbiodiniaceae. Furthermore, the codon usage bias of the plastid genomes in Heterocapsaceae, Amphidiniaceae, and Prorocentraceae species are greatly influenced by selection pressure, and in Pyrocystaceae, Symbiodiniaceae, Peridiniaceae, and Ceratiaceae species are influenced by both natural selection pressure and mutation pressure, indicating a family-level distinction in codon usage evolution in dinoflagellates. Phylogenetic analysis using 12 plastid-encoded proteins and five nucleus-encoded plastid proteins revealed accelerated evolution trend of both plastid- and nucleus-encoded plastid proteins in peridinin- and fucoxanthin-dinoflagellate plastids compared to plastid proteins of nondinoflagellate algae. These findings shed new light on the structure and evolution of plastid genomes in dinoflagellates, which will facilitate further studies on the evolutionary forcing and function of the diverse dinoflagellate plastids. The accelerated evolution documented here suggests plastid-encoded sequences are potentially useful for resolving closely related dinoflagellates.

Alkaline Phosphatase PhoD Mutation Induces Fatty Acid and Long-Chain Polyunsaturated Fatty Acid (LC-PUFA)-Bound Phospholipid Production in the Model Diatom Phaeodactylum tricornutum.

With rapid growth and high lipid contents, microalgae have become promising environmentally friendly candidates for renewable biodiesel and health supplements in our era of global warming and energy depletion. Various pathways have been explored to enhance algal lipid production, especially gene editing. Previously, we found that the functional loss of PhoD-type alkaline phosphatase (AP), a phosphorus-stress indicator in phytoplankton, could lead to increased lipid contents in the model diatom Phaeodactylum tricornutum, but how the AP mutation may change lipid composition remains unexplored. This study addresses the gap in the research and investigates the effects of PhoD-type AP mutation on the lipid composition and metabolic regulation in P. tricornutum using transcriptomic and lipidomic analyses. We observed significantly modified lipid composition and elevated production of fatty acids, lysophosphatidylcholine, lysophosphatidylethanolamine, ceramide, phosphatidylinositol bisphosphate, and monogalactosylmonoacylglycerol after PhoD_45757 mutation. Meanwhile, genes involved in fatty acid biosynthesis were upregulated in mutant cells. Moreover, the mutant exhibited increased contents of ω-3 long-chain polyunsaturated fatty acid (LC-PUFA)-bound phospholipids, indicating that PhoD_45757 mutation could improve the potential bioavailability of PUFAs. Our findings indicate that AP mutation could influence cellular lipid synthesis and probably redirect carbon toward lipid production and further demonstrate that AP mutation is a promising approach for the development of high-value microalgal strains for biomedical and other applications.

Species-dependent effects of seawater acidification on alkaline phosphatase activity in dinoflagellates.

Increases of atmospheric CO2 cause ocean acidification (OA) and global warming, the latter of which can stratify the water column and impede nutrient supply from deep water. Phosphorus (P) is an essential nutrient for phytoplankton to grow. While dissolved inorganic phosphorus (DIP) is the preferred form of P, phytoplankton have evolved alkaline phosphatase (AP) to utilize dissolved organic phosphorus (DOP) when DIP is deficient. Although the function of AP is known to require pH > 7, how OA affects AP activity and hence the capacity of phytoplankton to utilize DOP is poorly understood. Here, we examined the effects of pH conditions (5.5-11) on AP activity from six species of dinoflagellates, an important group of marine phytoplankton. We observed a general pattern that AP activity declined sharply at pH 5.5, peaked between pH 7 and 8, and dropped at pH > 8. However, our data revealed remarkable interspecific variations in optimal pH and niche breadth of pH. Among the species examined, Fugacium kawagutii and Prorocentrum cordatum had an optimal pH at 8, and Alexandrium pacificum, Amphidinium carterae, Effrenium voratum, and Karenia mikimotoi showed an optimal pH of 7. However, whereas A. pacificum and K. mikimotoi had the broadest pH niche for AP (7-10) and F. kawagutii the second (8-10), Am. carterae, E. voratum, and P. cordatum exhibited a narrow pH range. The response of Am. carterae AP to pH changes was verified using purified AP heterologously expressed in Escherichia coli. These results in concert suggest OA will likely differentially impact the capacity of different phytoplankton species to utilize DOP in the projected more acidified and nutrient-limited future ocean.

Active viral infection during blooms of a dinoflagellate indicates dinoflagellate-viral co-adaptation.

IMPORTANCE: This study represents the first that investigates in situ virus infection in dinoflagellate blooms. Our findings reveal highly similar viral assemblages that infected the bloom species Prorocentrum shikokuense and a co-adapted metabolic relationship between the host and the viruses in the blooms, which varied between the prolonged and the short-lived blooms of the same dinoflagellate species. These findings fill the gap in knowledge regarding the identity and behavior of viruses in a dinoflagellate bloom and shed light on what appears to be the complex mode of infection. The novel insight will be potentially valuable for fully understanding and modeling the role of viruses in regulating blooms of dinoflagellates and other algae.

Two-sided effects of the organic phosphorus phytate on a globally important marine coccolithophorid phytoplankton.

Dissolved organic phosphorus (DOP) is a potential source of aquatic eutrophication and pollution because it can potentially stimulate growth in some species and inhibit growth in other species of algae, the foundation of the marine ecosystem. Inositol hexaphosphate (also named phytic acid or PA), an abundant organophosphate, is presumably ubiquitous in the marine environment, but how it affects marine primary producers is poorly understood. Here, we investigated the bioavailability of this DOP to the cosmopolitan coccolithophore Emiliania huxleyi. Our results showed that E. huxleyi cells can take up PA and dissolved inorganic phosphorus (DIP) simultaneously. Absorbed PA can efficiently support algal growth, producing cell yield between DIP and phosphorus (P)-depleted conditions. Accordingly, PA supply as the sole P source highly influences cellular metabolism and nutrient stoichiometry. Particularly, PA-grown cultures exhibited enhanced carbon fixation, increased lipid content, activated energy metabolism, and induced nitrogen assimilation. However, our data suggest that PA may also exert some levels of toxic effects on E. huxleyi. This study provides novel insights into the variable effects of a DOP on marine phytoplankton, which will inform new inquiries about how the complex DOP constituencies in the ocean will shape phytoplankton community structure and function. IMPORTANCE The dissolved organic phosphorus (DOP) utilization in phytoplankton plays vital roles in cellular P homeostasis, P-nutrient niche, and the dynamics of community structure in marine ecosystems, but its mechanisms, potentially varying with species, are far from clear. In this study, we investigated the utilization of a widespread DOP species, which is commonly produced by plants (land plants and marine macrophytes) and released into coastal areas, in a globally distributed bloom-forming coccolithophore species in various phosphorus environments. Using a combination of physiological and transcriptomic measurements and analyses, our experimental results revealed the complex mechanism and two-sided effects of DOP (major algal growth-supporting and minor toxic effects) in this species, providing a novel perspective on phytoplankton nutrient regulation.

Phosphorus nutrition strategies in a Symbiodiniacean species: Implications in coral-alga symbiosis facing increasing phosphorus deficiency in future warmer oceans.

Coral reefs thrive in the oligotrophic ocean and rely on symbiotic algae to acquire nutrients. Global warming is projected to intensify surface ocean nutrient deficiency and anthropogenic discharge of wastes with high nitrogen (N): phosphorus (P) ratios can exacerbate P nutrient limitation. However, our understanding on how symbiotic algae cope with P deficiency is limited. Here, we investigated the responses of a coral symbiotic species of Symbiodiniaceae, Cladocopium goreaui, to P-limitation by examining its physiological performance and transcriptomic profile. Under P stress, C. goreaui exhibited decreases in algal growth, photosynthetic efficiency, and cellular P content but enhancement in carbon fixation, N assimilation, N:P ratio, and energy metabolism, with downregulated expression of carbohydrate exporter genes. Besides, C. goreaui showed flexible mechanisms of utilizing different dissolved organic phosphorus to relieve P deficiency. When provided glycerol phosphate, C. goreaui hydrolyzed it extracellularly to produce phosphate for uptake. When grown on phytate, in contrast, C. goreaui upregulated the endocytosis pathway while no dissolved inorganic phosphorus was released into the medium, suggesting that phytate was transported into the cell, potentially via the endocytosis pathway. This study sheds light on the survival strategies of C. goreaui and potential weakening of its role as an organic carbon supplier in P-limited environments, underscoring the importance of more systematic investigation on future projections of such effects.

Role of diatom-derived oxylipins in organic phosphorus recycling during coastal diatom blooms in the northern South China Sea.

Diatom-bacteria interactions and the associated bloom dynamics have not been fully understood in the coastal oceans. Here, we focus on the polyunsaturated aldehydes (PUAs) produced by diatoms in the post-bloom phase and look into their roles in microbial phosphorus (P) recycling outside of a P-limited estuary. The phytoplankton community in the bloom was dominated by PUAs-producing diatoms (Skeletonema costatum, Thalassiosira spp., and Pesudonitzschia delicates) with elevated concentrations of biogenic particulate PUAs. In addition, there were micromolar levels of particle-adsorbed PUAs hotspots with distinct compositions in and out of the bloom determined by a combining large-volume filtration and on-site derivation method. Field experiments were conducted to further assess the responses of particle-attached bacteria (PAB) to different PUAs amendments. We found no differences in the alkaline phosphatase (APase) activity and the abundance of PAB between inside and outside the bloom at a low PUAs dosage (<30 µM). However, for a high PUAs dosage (300 µM), APase activity and PAB growth were reduced significantly outside the bloom but no influences within the bloom. Our findings indicate that the hotspot-level oxylipins may play essential roles in bacterial P-remineralization in P-limited coastal areas. PAB can adapt to the high level of PUAs released by diatoms (or their resulting detritus) and potentially maintain a high rate of organic P recycling during the late stages of diatom blooms. Consequently, the interaction between oxylipin-rich diatoms and bacteria may affect phytoplankton blooms and carbon sequestration in the coastal oceans.

A comparative study reveals the relative importance of prokaryotic and eukaryotic proton pump rhodopsins in a subtropical marginal sea.

Proton-pump rhodopsin (PPR) in marine microbes can convert solar energy to bioavailable chemical energy. Whereas bacterial PPR has been extensively studied, counterparts in microeukaryotes are less explored, and the relative importance of the two groups is poorly understood. Here, we sequenced whole-assemblage metatranscriptomes and investigated the diversity and expression dynamics of PPR in microbial eukaryotes and prokaryotes at a continental shelf and a slope site in the northern South China Sea. Data showed the whole PPRs transcript pool was dominated by Proteorhodopsins and Xanthorhodopsins, followed by Bacteriorhodopsin-like proteins, dominantly contributed by prokaryotes both in the number and expression levels of PPR unigenes, although at the continental slope station, microeukaryotes and prokaryotes contributed similarly in transcript abundance. Furthermore, eukaryotic PPRs are mainly contributed by dinoflagellates and showed significant correlation with nutrient concentrations. Green light-absorbing PPRs were mainly distributed in >3 µm organisms (including microeukaryotes and their associated bacteria), especially at surface layer at the shelf station, whereas blue light-absorbing PPRs dominated the <3 µm (mainly bacterial) communities at both study sites, especially at deeper layers at the slope station. Our study portrays a comparative PPR genotype and expression landscape for prokaryotes and eukaryotes in a subtropical marginal sea, suggesting PPR's role in niche differentiation and adaptation among marine microbes.

Building consensus around the assessment and interpretation of Symbiodiniaceae diversity.

Within microeukaryotes, genetic variation and functional variation sometimes accumulate more quickly than morphological differences. To understand the evolutionary history and ecology of such lineages, it is key to examine diversity at multiple levels of organization. In the dinoflagellate family Symbiodiniaceae, which can form endosymbioses with cnidarians (e.g., corals, octocorals, sea anemones, jellyfish), other marine invertebrates (e.g., sponges, molluscs, flatworms), and protists (e.g., foraminifera), molecular data have been used extensively over the past three decades to describe phenotypes and to make evolutionary and ecological inferences. Despite advances in Symbiodiniaceae genomics, a lack of consensus among researchers with respect to interpreting genetic data has slowed progress in the field and acted as a barrier to reconciling observations. Here, we identify key challenges regarding the assessment and interpretation of Symbiodiniaceae genetic diversity across three levels: species, populations, and communities. We summarize areas of agreement and highlight techniques and approaches that are broadly accepted. In areas where debate remains, we identify unresolved issues and discuss technologies and approaches that can help to fill knowledge gaps related to genetic and phenotypic diversity. We also discuss ways to stimulate progress, in particular by fostering a more inclusive and collaborative research community. We hope that this perspective will inspire and accelerate coral reef science by serving as a resource to those designing experiments, publishing research, and applying for funding related to Symbiodiniaceae and their symbiotic partnerships.

In Situ Molecular Ecological Analyses Illuminate Distinct Factors Regulating Formation and Demise of a Harmful Dinoflagellate Bloom.

The development and demise of a harmful algal bloom (HAB) are generally regulated by multiple processes; identifying specific critical drivers for a specific bloom is important yet challenging. Here, we conducted a whole-assemblage molecular ecological study on a dinoflagellate bloom to address the hypothesis that energy and nutrient acquisition, defense against grazing and microbial attacks, and sexual reproduction are critical to the rise and demise of the bloom. Microscopic and molecular analyses identified the bloom-causing species as Karenia longicanalis and showed that the ciliate Strombidinopsis sp. was dominant in a nonbloom plankton community, whereas the diatom Chaetoceros sp. dominated the after-bloom community, along with remarkable shifts in the community structure for both eukaryotes and prokaryotes. Metatranscriptomic analysis indicated that heightened energy and nutrient acquisition in K. longicanalis significantly contributed to bloom development. In contrast, active grazing by the ciliate Strombidinopsis sp. and attacks by algicidal bacteria (Rhodobacteracea, Cryomorphaceae, and Rhodobacteracea) and viruses prevented (at nonbloom stage) or collapsed the bloom (in after-bloom stage). Additionally, nutrition competition by the Chaetoceros diatoms plausibly contributed to bloom demise. The findings suggest the importance of energy and nutrients in promoting this K. longicanalis bloom and the failure of antimicrobial defense and competition of diatoms as the major bloom suppressor and terminator. This study provides novel insights into bloom-regulating mechanisms and the first transcriptomic data set of K. longicanalis, which will be a valuable resource and essential foundation for further elucidation of bloom regulators of this and related species of Kareniaceae in the future. IMPORTANCE HABs have increasingly occurred and impacted human health, aquatic ecosystems, and coastal economies. Despite great efforts, the factors that drive the development and termination of a bloom are poorly understood, largely due to inadequate in situ data about the physiology and metabolism of the causal species and the community. Using an integrative molecular ecological approach, we determined that heightened energy and nutrient acquisition promoted the bloom, while resource allocation in defense and failure to defend against grazing and microbial attacks likely prevented or terminated the bloom. Our findings reveal the differential roles of multiple abiotic and biotic environmental factors in driving the formation or demise of a toxic dinoflagellate bloom, suggesting the importance of a balanced biodiverse ecosystem in preventing a dinoflagellate bloom. The study also demonstrates the power of whole-assemblage metatranscriptomics coupled to DNA barcoding in illuminating plankton ecological processes and the underlying species and functional diversities.

Phytoplankton and Microzooplankton Community Structure and Assembly Mechanisms in Northwestern Pacific Ocean Estuaries with Environmental Heterogeneity and Geographic Segregation.

Phytoplankton and microzooplankton are crucial players in marine ecosystems and first responders to environmental changes, but their community structures and how they are shaped by environmental conditions have rarely been studied simultaneously. In this study, we conducted an eDNA metabarcoding sequencing combined with multiple statistical methods to simultaneously analyze the phytoplankton and microzooplankton in Liaohe (LH) and Yalujiang (YLJ) estuaries. The major objective was to examine how plankton community structure and assembly mechanism may differ between two estuaries with similar latitudinal position and climate but geographical segregation and differential level of urbanization (more in LH). Clear differences in diversity and composition of phytoplankton and microzooplankton communities between LH and YLJ estuaries were observed. Richness of phytoplankton was significantly higher in LH than YLJ, while richness of microzooplankton was higher in YLJ. The magnitude of intrahabitat variations in phytoplankton communities was significantly stronger than that of microzooplankton. Some phytoplankton and microzooplankton taxa also showed interhabitat differences in their relative abundances. Phytoplankton showed a stronger geographic distance-decay of similarity than microzooplankton, while significant environmental distance-decay of similarity in microzooplankton was found in the less urbanized YLJ estuary. Community assembly of phytoplankton was, based on the neutral community models, driven primarily by stochastic processes, while deterministic processes contributed more for microzooplankton. Furthermore, we detected wider habitat niche breadths and stronger dispersal abilities in phytoplankton than in microzooplankton. These results suggest that passive dispersal shapes the phytoplankton community whereas environmental selection shapes the microzooplankton community. IMPORTANCE Understanding the underlying mechanisms shaping a metacommunity is useful to management for improving the ecosystem function. The research presented in the manuscript mainly tried to address the effects of habitat geography and environmental conditions on the phytoplankton and microzooplankton communities, and the underlying mechanisms of community assembly in temperate estuaries. In order to achieve this purpose, we developed a metabarcoding sequencing method based on 18S rRNA gene. The phytoplankton and microzooplankton communities from two estuaries with similar latitude and climatic conditions but obvious geographical segregation and significant environmental heterogeneity were investigated. The results of our study could lay a solid foundation for ascertaining phytoplankton and microzooplankton communities in estuaries with obvious environmental heterogeneity and geographic segregation and mechanisms underlying community assembly.

High Heterotrophic Plasticity of Massive Coral Porites pukoensis Contributes to Its Tolerance to Bioaccumulated Microplastics.

Scleractinian corals have been observed to be capable of accumulating microplastics from reef environments; however, the tolerant mechanism is poorly known. Here, we examined the response of Porites pukoensis to microplastic pollution by analyzing algal symbiont density, energetic metabolism, and caspase3 activities (representing the apoptosis level) in the coral-Symbiodiniaceae association. The environments of three fringing reef regions along the south coast of Sanya City, Hainan Province of China, were polluted by microplastics (for example, microplastic concentrations in the seawater ranged from 3.3 to 46.6 particles L-1), resulting in microplastic accumulation in P. pukoensis (0.4-2.4 particles cm-2). The accumulation of microplastics was negatively correlated to algal symbiont density in the corals but not to caspase3 activities in the two symbiotic partners, demonstrating that P. pukoensis could tolerate accumulated microplastics despite the decrease of algal symbiont density. Furthermore, results from the carbon stable isotope and cellular energy allocation assay indicated that P. pukoensis obtained energy availability (mainly as lipid reserves) using the switch between heterotrophy and autotrophy to maintain energy balance and cope with accumulated microplastics. Collectively, P. pukoensis achieved tolerance to microplastic pollution by maintaining energy availability, which was largely attributed to its high heterotrophic plasticity.

Molecular phylogeny of the spiny-surfaced species of the dinoflagellate Prorocentrum with the description of P. thermophilum sp. nov. and P. criophilum sp. nov. (Prorocentrales, Dinophyceae).

Spiny-surfaced species of Prorocentrum form harmful algal blooms, and its taxonomic identity is obscure due to the size and shape variability. Molecular phylogenies reveal two major clades: one for P. cordatum with sequences mainly retrieved as P. minimum, and the other for P. shikokuense with sequences also retrieved as P. dentatum and P. donghaiense. Several closely related clades still need to be characterized. Here, we provide nuclear SSU and LSU rRNA genes, and nuclear ITS region (ITS1-5.8S gene-ITS2) sequences of the strain CCMP3122 isolated from Florida (initially named P. donghaiense) and strains Prorocentrum sp. RCC6871-2 from the Ross Sea, Antarctica. We describe Prorocentrum thermophilum sp. nov. based on the strain CCMP3122, a species also distributed in the open waters of the Gulf of Mexico, New Zealand, and the Arabian Gulf; and Prorocentrum criophilum sp. nov. based on the strain RCC6872, which is distributed in the Antarctic Ocean and Arctic Sea. Prorocentrum thermophilum is roundish (~14 µm long, ~12 µm wide), with an inconspicuous anterior spine-like prolongation under light microscopy, valves with tiny, short knobs (5-7 per µm2 ), and several (<7) large trichocyst pores (~0.3 µm) in the right valve, as well as smaller pores (~0.15 µm). Prorocentrum criophilum is round in valve view (~11 µm long, 10 µm wide) and asymmetrically roundish in lateral view, the periflagellar area was not discernible under light microscopy, valves with very tiny, short knobs (6-10 per µm2 ), and at least 12 large pores in the right valve. Other potentially undescribed species of spiny-surfaced Prorocentrum are discussed.