Dinoflagellate Amphidinium Carterae Research Articles

Lipidomic profile in three species of dinoflagellates (Amphidinium carterae, Cystodinium sp., and Peridinium aciculiferum) containing very long chain polyunsaturated fatty acids

This study describes the identification of very long chain polyunsaturated fatty acids (VLCPUFAs) in three strains of dinoflagellates (Amphidinium carterae, Cystodinium sp., and Peridinium aciculiferum). The strains were cultivated and their lipidomic profiles were obtained by high resolution mass spectrometry with the aid of positive and negative electrospray ionization (ESI) mode by Orbitrap apparatus. Hydrophilic interaction liquid chromatography (HILIC/ESI) was used to separate major lipid classes of the three genera of dinoflagellates by neutral loss scan showing the ion [M + H-28:8]+, where 28:8 was octacosaoctaenoic acid, and by precursor ion scanning of ions at m/z 407, which was an ion corresponding to the structure of acyl of 28:8 acid (C27H39COO−). Based on these analyzes, it was found that out of more than a dozen lipid classes present in the total lipids, only two classes of neutral lipids, i.e. major triacylglycerols and minor diacylglycerols contain VLCPUFAs. In polar lipids, VLCPUFAs were identified only in phosphatidic acid (PA) and phosphatidyl choline (PC) or in their lyso-forms (LPA and LPC). Further analysis of individual lipid classes by reversed-phase high-performance liquid chromatography (RP-HPLC) showed the presence of triacylglycerols (TAGs) containing VLCPUFAs, i.e. molecular species of the sn-28:7/28:8/28:8, sn-26:7/28:7/28:8, or sn-26:7/28:8/28:8 types. These TAGs are the longest and most unsaturated TAGs isolated from a natural source that have yet been synthesized. In the case of PA and PC, tandem MS identified sn-28:8/16:0-PA and sn-28:8/16:0-PC and the corresponding lyso-forms (28:8-LPC and 28:8-LPA). All these results indicate that TAGs containing VLCPUFAs are biosynthesized in dinoflagellates in the same manner as in higher eukaryotic organisms, which means that the PA, after conversion to DAG, serves as a precursor in the biosynthesis of other phospholipids, e.g. PC, and, after further acylation, also of TAG.

Fatty acid composition and docosahexaenoic acid (DHA) content of the heterotrophic dinoflagellate Oxyrrhis marina fed on dried yeast: compared with algal prey

The heterotrophic dinoflagellate Oxyrrhis marina is known to produce high levels of docosahexaenoic acid (DHA) when fed on diverse algal prey. However, large-scale culturing of algal prey species is not easy and requires a large amount of budget, and thus more easily cultivable and low-cost prey is required. Dried yeast was selected as a strong candidate for an alternative prey in our preliminary tests. Thus, we explored the fatty acid composition and DHA production of O. marina fed on dried yeast and compared these results to those of O. marina fed on two algal prey species: the phototrophic dinoflagellate Amphidinium carterae and chlorophyte Chlorella sp. powder. O. marina fed on dried yeast, which does not contain DHA, produced the same high level of DHA as those fed on DHA-containing A. carterae. This indicates that O. marina is likely to produce DHA by itself regardless of prey items. Furthermore, the DHA content (and portion of total fatty acid methyl esters) of O. marina satiated with dried yeast, 52.40 pg per cell (and 25.9%), was considerably greater than that of O. marina fed on A. carterae (26.91 pg per cell; 15.7%) or powder of Chlorella sp. powder (21.24 pg per cell; 16.7%). The cost of dried yeast (approximately 10 US dollars for 1 kg dried yeast) was much lower than that of obtaining the algal prey (approximately 160 US dollars for 1 kg A. carterae). Therefore, compared to conventional algal prey, dried yeast is a more easily obtainable and lower-cost prey for use in the production of DHA by O. marina. Key words: Amphidinium carterae; Chloroella; DHA production; feeding; lipid; low-cost prey; omega-3

Distance measurements in peridinin-chlorophyll a-protein by light-induced PELDOR spectroscopy. Analysis of triplet state localization

Triplet-triplet energy transfer from chlorophylls to carotenoids is the mechanism underlying the photoprotective role played by carotenoids in many light harvesting complexes, during photosynthesis. The peridinin–chlorophyll-a protein (PCP) is a water-soluble light harvesting protein of the dinoflagellate Amphidinium carterae, employing peridinin as the main carotenoid to fulfil this function.The dipolar coupling of the triplet state of peridinin, populated under light excitation in isolated PCP, to the MTSSL nitroxide, introduced in the protein by site-directed mutagenesis followed by spin labeling, has been measured by Pulse ELectron-electron DOuble Resonance (PELDOR) spectroscopy. The triplet-nitroxide distance derived by this kind of experiments, performed for the first time in a protein system, allowed the assignment of the triplet state to a specific peridinin molecule belonging to the pigment cluster. The analysis strongly suggests that this peridinin is the one in close contact with the water ligand to the chlorophyll a, thus supporting previous evidences based on ENDOR and time resolved-EPR.

Antialgal and antilarval activities of bioactive compounds extracted from the marine dinoflagellate Amphidinium carterae

With the global ban on the application of organotin-based marine coatings by the International Maritime Organization, the development of environmentally friendly, low-toxic and nontoxic antifouling compounds for marine industries has become an urgent need. Marine microorganisms have been considered as a potential source of natural antifoulants. In this study, the antifouling potential of marine dinoflagellate Amphidinium carterae, the toxic and red-tide microalgae, was investigated. We performed a series of operations to extract the bioactive substances from Amphidinium carterae and tested their antialgal and antilarval activities. The crude extract of Amphidinium carterae showed significant antialgal activity and the EC50 value against Skeletonema costatum was 55.4 μg mL−1. After purification, the isolated bioactive substances (the organic extract C) exhibited much higher antialgal and antilarval activities with EC50 of 12.9 μg mL−1 against Skeletonema costatum and LC50 of 15.1 μg mL−1 against Amphibalanus amphitrite larvae. Subsequently, IR, Q-TOFMS, and GC-MS were utilized for the structural elucidation of the bioactive compounds, and a series of unsaturated and saturated 16- to 22-carbon fatty acids were detected. The data suggested the bioactive compounds isolated from Amphidinium carterae exhibited a significant inhibiting effect against the diatom Skeletonema costatum and Amphibalanus amphitrite larvae, and could be substitutes for persistent, toxic antifouling compounds.

Response of polyamine pools in marine phytoplankton to nutrient limitation and variation in temperature and salinity

Previous field observations suggest that the composition of intracellular polyamine pools in phytoplankton determines the profile of polyamines released to the surrounding environ- ment; thus, knowing how these pools vary among species and in response to factors affecting phytoplankton growth provides a basis for understanding fluctuations in dissolved polyamines. Our analyses of the polyamine content of axenic marine phytoplankton cultures show that intra- cellular polyamine pools are large (100s to 1000s µmol l �1 of biovolume) and that putrescine and spermidine are the major compounds present; however, composition varied with species. Norsper- midine and norspermine were the dominant compounds found in the diatom Thalassiosira pseudonana and the dinoflagellate Amphidinium carterae, respectively. We explored the effects of temperature, salinity and nutrient limitation on polyamine pools in T. pseudonana, and found that either increasing temperature or decreasing salinity increased polyamine concentrations in cells and in the growth medium. Nutrient (N, P or Si) limitation caused reductions of intracellular polyamine concentrations, but release was enhanced under N or Si limitation. Polyamine ratios in the media were not the same as in intracellular pools, suggesting selective release or uptake of polyamines by phytoplankton. Thus, the composition of the dissolved polyamine pool in seawater can vary significantly and on short time scales, depending on phytoplankton community composi- tion and environmental factors affecting phytoplankton physiology. Our work provides experi- mental verification that biological mechanisms support inferences derived from environmental correlations about the factors controlling polyamine distributions in the sea.

Triplet–triplet energy transfer from chlorophylls to carotenoids in two antenna complexes from dinoflagellate Amphidinium carterae

Room temperature transient absorption spectroscopy with nanosecond resolution was used to study quenching of the chlorophyll triplet states by carotenoids in two light-harvesting complexes of the dinoflagellate Amphidinium carterae: the water soluble peridinin–chlorophyll protein complex and intrinsic, membrane chlorophyll a–chlorophyll c2–peridinin protein complex. The combined study of the two complexes facilitated interpretation of a rather complicated relaxation observed in the intrinsic complex. While a single carotenoid triplet state was resolved in the peridinin–chlorophyll protein complex, evidence of at least two different carotenoid triplets was obtained for the intrinsic light-harvesting complex. Most probably, each of these carotenoids protects different chlorophylls. In both complexes the quenching of the chlorophyll triplet states by carotenoids occurs with a very high efficiency (~100%), and with transfer times estimated to be in the order of 0.1ns or even faster. The triplet–triplet energy transfer is thus much faster than formation of the chlorophyll triplet states by intersystem crossing. Since the triplet states of chlorophylls are formed during the whole lifetime of their singlet states, the apparent lifetimes of both states are the same, and observed to be equal to the carotenoid triplet state rise time (~5ns).

Pigment organisation in the membrane-intrinsic major light-harvesting complex of Amphidinium carterae: Structural characterisation of the peridinins and chlorophylls a and c2 by resonance Raman spectroscopy and from sequence analysis

The structures and environments of the protein-bound peridinins (Pers) and chlorophylls (Chls) a/c2 in the membrane-intrinsic major light-harvesting complex of the dinoflagellate Amphidinium carterae (LHCAmph) are characterised using resonance Raman (RR) spectroscopy with 11 excitation wavelengths, at 77K. The excitation-dependent variation in the CC stretching mode (ν1) suggests the presence of three Pers with conjugation lengths over 8 double bonds (dBs), and one diadinoxanthin, between 413.7 and 528.7nm. Two Perred species are revealed on excitation at 550 and 560nm. These Perred species exhibit anomalously low ν1 values, together with notable resonant enhancement of lactone ring-breathing and -deformation modes. To discern protein-specific effects, the RR spectra are compared to that of Per in polar (acetonitrile), polarisable (toluene) and polar-protic (ethanol) solvents. Resonantly enhanced lactone, ring-breathing (942cm−1) and ring-deformation (~650cm−1), modes are identified both in solution, and in the protein, and discussed in the context of the mixing of the S1 and S2 states, and formation of the intramolecular charge-transfer (ICT) state. In the Chl-absorbing region, two sets of Chl c2's, and (at least) six Chl a's can be differentiated. With a pigment ratio of 5–6 (Chl a):2 (Chl c2):5–6 (Per):1 Ddx determined from the fit to the RT absorption and 77K RR spectra, sequence comparison of LHCAmp to LHCII, and the diatom LHC, fucoxanthin–chlorophyll-a/c–protein (FCP), a template for the conserved pigment binding sites is proposed, to fill the paucity of structural information in the absence of a crystal structure for LHCAmph.

Efficient pathways of excitation energy transfer from delocalized S2 excitons in the peridinin-chlorophyll a-protein complex.

Excitation energy transfer (EET) in peridinin-chlorophyll-protein (PCP) complexes is dominated by the S1 → Qy pathway, but the high efficiencies cannot be solely explained by this one pathway. We postulate that EET from peridinin S2 excitons may also be important. We use complete active space configuration interaction calculations and the transition density cube method to calculate Coulombic couplings between peridinin and chlorophyll a in the PCP complex of the dinoflagellate Amphidinium carterae and compare monomeric and dimeric delocalized peridinin S2 excited states. Our calculations show that the S2 → Qy EET pathway from peridinin to chlorophyll a is the dominant energy transfer pathway from the S2 excited state in PCP, with several values in the sub-picosecond range. This result suggests that the S2 → Qy EET pathway may be responsible for the reported chlorophyll a bleaching signature seen in experiment at around 200 fs, and not the S2 → Qx pathway as previously suggested.

Excitation Energy Transfer in the Peridinin-Chlorophyll a-Protein Complex Modeled Using Configuration Interaction

We modeled excitation energy transfer (EET) in the peridinin-chlorophyll a-protein (PCP) complex of dinoflagellate Amphidinium carterae to determine which pathways contribute dominantly to the high efficiency of this process. We used complete active space configuration interaction (CAS-CI) to calculate electronic structure properties of the peridinin (PID) and chlorophyll a (CLA) pigments in PCP and the transition density cube (TDC) method to calculate Coulombic couplings between energy transfer donors and acceptors. Our calculations show that the S1 → Qy EET pathway from peridinin to chlorophyll a is the dominant energy transfer pathway in PCP, with two sets of interactions-between PID612 and CLA601 and between PID622 and CLA602-contributing most strongly. EET lifetimes for these two interactions were calculated to be 2.66 and 2.90, with quantum efficiencies of 85.75 and 84.65%, respectively. The calculated Coulombic couplings for EET between two peridinin molecules in the strongly allowed S2 excited states are extremely large and suggest excitonic coupling between pairs of peridinin S2 states. This methodology is also broadly applicable to the study of EET in other photosynthetic complexes and/or organic photovoltaics, where both single and double excitations are present and donor and acceptor molecules are tightly packed.

Antifungal amphidinol 18 and its 7-sulfate derivative from the marine dinoflagellate Amphidinium carterae.

Two new polyketides of the amphidinol family, amphidinol 18 (AM18, 1) and its corresponding 7-sulfate derivative (AM19, 2), have been isolated from the MeOH extract of the dinoflagellate Amphidinium carterae. Structure elucidation of the two polyoxygenated molecules has been accomplished by extensive use of spectroscopic and spectrometric techniques. AM18 exhibited antifungal activity against Candida albicans at 9 μg/mL.

Feeding by the newly described mixotrophic dinoflagellate Gymnodinium smaydae: Feeding mechanism, prey species, and effect of prey concentration

To investigate feeding by the newly described mixotrophic dinoflagellate Gymnodinium smaydae, we explored the feeding mechanism and the kinds of prey species that G. smaydae is able to feed on. In addition, we measured the growth and ingestion rates of G. smaydae on optimal and suboptimal algal prey Heterocapsa rotundata and Heterocapsa triquetra as a function of prey concentration. Among the 19 algal prey species offered, G. smaydae ingested only thecate dinoflagellates H. rotundata, H. triquetra, Heterocapsa sp., and Scrippsiella trochoidea. Among the peduncle-feeding dinoflagellates so far reported, G. smaydae is the only grazer that is able to feed on S. trochoidea and one of the two species that are able to feed on H. triquetra. However, G. smaydae did not feed on the raphidophyte Heterosigma akashiwo, the cryptophytes Teleaulax sp. and Rhodomonas salina, and the small dinoflagellate Amphidinium carterae which all the other peduncle-feeding dinoflagellates except Stoeckeria algicida are able to feed on. G. smaydae fed on algal prey using a peduncle after anchoring the prey by a tow filament. All Heterocapsa species supported high positive growth of G. smaydae, S. trochoidea only helped in merely maintaining the predator population. With increasing mean prey concentration, the growth and ingestion rates for G. smaydae on H. rotundata increased rapidly, but became saturated at concentrations of 455ngCml−1 (3500cellsml−1), while that on H. triquetra increased rapidly, but slowly at concentrations of 293ngCml−1 (945cellsml−1). The maximum specific growth rates (i.e., mixotrophic growth) of G. smaydae on H. rotundata and H. triquetra were 2.226d−1 and 1.053d−1, respectively, at 20°C under a 14:10h light–dark cycle of 20μEm−2s−1, while the growth rates (i.e., phototrophic growth) under the same light conditions without added prey were 0.005 to −0.051d−1. The maximum ingestion rates of G. smaydae on H. rotundata and H. triquetra were 1.59ngCgrazer−1d−1 (12.3cellsgrazer−1d−1) and 0.24ngCgrazer−1d−1 (0.8cellsgrazer−1d−1), respectively. The calculated grazing coefficients for G. smaydae on co-occurring H. rotundata or H. triquetra were up to 0.23h−1 or 0.02h−1, respectively (i.e., 21% or 2% of the population of H. rotundata or H. triquetra was removed by G. smaydae populations in 1h). The results of the present study suggest that G. smaydae can sometimes have a considerable grazing impact on the population of H. rotundata.

Feeding by the newly described heterotrophic dinoflagellate Stoeckeria changwonensis: A comparison with other species in the family Pfiesteriaceae

The feeding ecology of the newly described heterotrophic dinoflagellate Stoeckeria changwonensis was explored. The feeding behavior of S. changwonensis, and the kinds of prey species that it feeds on were investigated with several different types of microscopes and high-resolution video-microscopy. Additionally, the growth and ingestion rates of S. changwonensis as a function of prey concentration for perch (Lateolabrax japonicus) blood cells, the raphidophyte Heterosigma akashiwo, the cryptophytes Rhodomonas salina and Teleaulax sp., and the phototrophic dinoflagellate Amphidinium carterae prey were measured. S. changwonensis feeds on prey through a peduncle, after anchoring the prey by using a tow filament. This type of feeding behavior is similar to that of Stoeckeria algicida, Pfiesteria piscicida, and Luciella masanensis in the family Pfiesteriaceae; however, S. changwonensis feeds on various kinds of prey species different from those of the other heterotrophic dinoflagellates. S. changwonensis ingested perch blood cells and diverse algal species, in particular, the large thecate dinoflagellate Lingulodinium polyedrum which are not eaten by the other peduncle feeders. H. akashiwo and the perch blood cells supported positive growth of S. changwonensis, but R. salina, Teleaulax sp., and A. carterae which support positive growth of P. piscicida and L. masanensis did not support positive growth of S. changwonensis. With increasing mean prey concentration the growth rates for S. changwonensis on H. akashiwo and the perch blood cells increased rapidly and then slowly or became saturated. The maximum growth rates of S. changwonensis on H. akashiwo and the perch blood cells were 0.376 and 0.354d−1, respectively. Further, the maximum ingestion rates of S. changwonensis on H. akashiwo and the perch blood cells were 0.35ngCpredator−1d−1 (3.5cellspredator−1d−1) and 0.27ngCpredator−1d−1 (29cellspredator−1d−1), respectively. These maximum growth and ingestion rates of S. changwonensis on H. akashiwo, the perch blood cells, R. salina, Teleaulax sp., and A. carterae differed considerably from those of S. algicida, P. piscicida, and L. masanensis on the same prey species. Thus, the feeding behavior of S. changwonensis may differ from that of other species in the family Pfiesteriaceae.

Measurements of diel variation of acoustic backscatter power from phytoplankton

Phytoplankton is a primary producer in the ocean, and its vegetative activity plays an important role in controlling the global environment. In this study, the high-frequency acoustic signals were collected to evaluate the photosynthetic activity of phytoplankton during a day. The integrated backscatter power (IBP) from a dinoflagellate Cochlodinium polykrikoides was measured by a 5 MHz acoustic system during a 5-day cultivation period with a 14 h:10 h light:dark cycle. IBP increased by 0.6 dB in five days, but varied by 0.83 ± 0.1 dB during an irradiance cycle. The daily increase in IBP was a result of an increase in the number of cells during cultivation, while the diel variation was partly resulted from the variation of volume of cells by photosynthesis. In addition, cell division and separation might affect IBP. IBP of another dinoflagellate Amphidinium carterae Hulburt was also measured by a 10MHz transducer during a 3-day cultivation period while cell volume and photosynthetic capacity were measured four times a day (07:00, 12:00, 19:00, 24:00). This study suggests that high-frequency acoustics may be a meaningful tool to investigate the photosynthetic metabolism of a phytoplankton cell. [This work was supported by Defense Acquisition Program Administration and Agency for Defense Development under the contract UD130007DD.]

Influence of diet on chitobiase-based production rates for the harpacticoid copepod Tigriopus californicus

The influence of diet on copepod production is important when examining energy transfer within marine food webs, yet a direct link between diet and copepod production is lacking. Recently, measurements of the crustacean moulting enzyme chitobiase have allowed for rapid estimates of production rates for zooplankton communities. We tested the sensitivity of this method as a tool for measuring the productivity response of copepods to different diets by rearing the harpacticoid splash-pool copepod, Tigriopus californicus, on single and mixed species phytoplankton diets. Chitobiase-based productivity measurements demonstrate that a single species algal diet of the diatom Thalassiosira weissflogii was the best food item for T. californicus. In contrast, the dinoflagellate Amphidinium carterae was likely toxic given that copepods reared on this diet spent more time in early developmental stages and ultimately failed to survive. Copepod production and daily growth rates of T. californicus were higher on the T. weissflogii and natural food assemblage diets compared with the A. carterae or mixed (A. carterae/T. weissflogii) diets. Our use of the chitobiase method in a lab setting demonstrates the potential utility and sensitivity of this approach for field studies examining the impact and significance of short-term shifts in food quality on entire crustacean zooplankton communities.

Polyuridylylation and processing of transcripts from multiple gene minicircles in chloroplasts of the dinoflagellate Amphidinium carterae

Although transcription and transcript processing in the chloroplasts of plants have been extensively characterised, the RNA metabolism of other chloroplast lineages across the eukaryotes remains poorly understood. In this paper, we use RT-PCR to study transcription and transcript processing in the chloroplasts of Amphidinium carterae, a model peridinin-containing dinoflagellate. These organisms have a highly unusual chloroplast genome, with genes located on multiple small 'minicircle' elements, and a number of idiosyncratic features of RNA metabolism including transcription via a rolling circle mechanism, and 3' terminal polyuridylylation of transcripts. We demonstrate that transcription occurs in A. carterae via a rolling circle mechanism, as previously shown in the dinoflagellate Heterocapsa, and present evidence for the production of both polycistronic and monocistronic transcripts from A. carterae minicircles, including several regions containing ORFs previously not known to be expressed. We demonstrate the presence of both polyuridylylated and non-polyuridylylated transcripts in A. carterae, and show that polycistronic transcripts can be terminally polyuridylylated. We present a model for RNA metabolism in dinoflagellate chloroplasts where long polycistronic precursors are processed to form mature transcripts. Terminal polyuridylylation may mark transcripts with the correct 3' end.

Feeding by the Newly Described, Nematocyst‐Bearing Heterotrophic Dinoflagellate Gyrodiniellum shiwhaense

We explored the feeding ecology of the newly described, nematocyst-bearing heterotrophic dinoflagellate Gyrodiniellum shiwhaense (GenBank accession number=FR720082). Using several different types of microscopes and high-resolution video-microscopy, we investigated feeding behavior and types of prey species that G. shiwhaense feeds upon. Additionally, we measured its growth and ingestion rates on its optimal algal prey, the cryptophyte Teleaulax sp. and the dinoflagellate Amphidinium carterae, as a function of prey concentration. These rates were measured for other edible prey at single prey concentrations at which the growth and ingestion rates of G. shiwhaense were saturated. After anchoring the prey with a tow filament, G. shiwhaense fed using a peduncle, ingesting small algal species with equivalent spherical diameters (ESDs) of <13 μm. However, it did not feed on larger algal species that had ESDs≥13 μm or the small diatom Skeletonema costatum. The specific growth rates for G. shiwhaense feeding upon Teleaulax sp. and A. carterae increased rapidly with increasing mean prey concentration before saturating at concentrations of ca. 180-430 ng C/ml. The maximum specific growth rate of G. shiwhaense on Teleaulax sp. and A. carterae were 1.05 and 0.82/d, respectively. However, Heterosigma akashiwo did not support positive growth of G. shiwhaense. The maximum ingestion rates of G. shiwhaense on Teleaulax sp. and A. carterae were 0.35 and 0.54 ng C/grazer/d, respectively. The calculated grazing coefficients attributable to G. shiwhaense on co-occurring cryptophytes and Amphidinium spp. were 0.01-1.87/d and 0.08-2.60/d, respectively. Our results suggest that G. shiwhaense can have a considerable grazing impact on algal populations.

Alkaline Phosphatase Gene Sequence And Transcriptional Regulation By Phosphate Limitation In Amphidinium Carterae (Dinophyceae)(1).

Alkaline phosphatase (AP) in phytoplankton facilitates the utilization of dissolved organic phosphorus (DOP) when the dissolved inorganic phosphorus (DIP) is limited in the environment. The AP gene sequence and its expression under DIP limitation has not been studied in dinoflagellates. In this study, we isolated the full-length cDNA of AP from the toxic dinoflagellate Amphidinium carterae Hulburt (2,112 bp, named as acaap). The deduced amino acid sequence of acaap (ACAAP, 704 amino acid residues) was identified as a membrane-associated protein, in agreement with the dominantly cell surface localization of the AP activity shown with enzyme-labeled fluorescence (ELF) labeling. ACAAP shares sequence similarity in the key domains with APs from diatoms, proteobacteria, and cyanobacteria. In accordance, phylogenetic reconstruction showed clustering of ACAAP with counterparts in those organisms, although branches were long as a result of the generally high variability of the gene sequence. The expression levels of acaap were studied for A. carterae cultured in media with different phosphate concentrations using quantitative reverse-transcription PCR (RT-qPCR) method. The result showed that the transcription level of acaap was elevated in the DIP-depleted cultures relative to the DIP-replete cultures and repressed upon resupply of DIP. The transcription level of acaap exhibited a positive correlation with AP enzyme activity. Taken together, these results demonstrate that AP activity and gene expression are regulated by the availability of DIP in A. carterae, suggesting that AP expression is a promising indicator of DIP stress in this and possibly other species of dinoflagellates.

Gyrodiniellum shiwhaense n. gen., n. sp., A New Planktonic Heterotrophic Dinoflagellate from the Coastal Waters of Western Korea: Morphology and Ribosomal DNA Gene Sequence

The heterotrophic dinoflagellate Gyrodiniellum shiwhaense n. gen., n. sp. is described from live cells and from cells prepared for light, scanning electron, and transmission electron microscopy. Also, sequences of the small subunit (SSU) and large subunit (LSU) of rDNA have been analyzed. The episome is conical, while the hyposome is ellipsoid. Cells are covered with polygonal amphiesmal vesicles arranged in 16 horizontal rows. Unlike other Gyrodinium-like dinoflagellates, the apical end of the cell shows a loop-shaped row of five elongate amphiesmal vesicles. The cingulum is displaced by 0.3-0.5 × cell length. Cells that were feeding on the dinoflagellate Amphidinium carterae Hulburt were 9.1-21.6 μm long and 6.6-15.7 μm wide. Cells of G. shiwhaense contain nematocysts, trichocysts, a peduncle, and pusule systems, but they lack chloroplasts. The SSU rDNA sequence is >3% different from that of the six most closely related species: Warnowia sp. (FJ947040), Lepidodinium viride Watanabe, Suda, Inouye, Sawaguchi & Chihara, Gymnodinium aureolum (Hulburt) Hansen, Gymnodinium catenatum Graham, Nematodinium sp. (FJ947039), and Gymnodinium sp. MUCC284 (AF022196), while the LSU rDNA is 11-12% different from that of Warnowia sp., G. aureolum, and Nematodinium sp. (FJ947041). The phylogenetic trees show that the species belongs in the Gymnodinium sensu stricto clade. However, in contrast to Gymnodinium spp., cells lack nuclear envelope chambers and a nuclear fibrous connective. Unlike Polykrikos spp., cells of which possess a taeniocyst-nematocyst complex, G. shiwhaense has nematocysts but lacks taeniocysts. It differs from Paragymnodinium shiwhaense Kang, Jeong, Moestrup & Shin by possessing nematocysts with stylets and filaments. Gyrodiniellum shiwhaense n. gen., n. sp. furthermore lacks ocelloids, in contrast to Warnowia spp., Nematodinium spp., and Proterythropsis spp. Based on morphological and molecular data, we suggest that the taxon represents a new species within a new genus.

Isolation and Characterization of Exopolysaccharide Secreted by a Toxic Dinoflagellate, Amphidinium carterae Hulburt 1957 and Its Probable Role in Harmful Algal Blooms (HABs)

Extracellular polymeric substances (EPS) produced by a toxic dinoflagellate Amphidinium carterae Hulburt 1957 was isolated and characterized. Molecular masses of the EPS were about 233 and 1,354kDa. Spectral analyses by (1)H nuclear magnetic resonance and Fourier Transformed-Infrared Spectroscopy revealed the characteristic of the functional groups viz. primary amine, carboxyl, halide, and sulfate groups present in the EPS. However, five elements (C, O, Na, S, and Ca) were detected by scanning electron microscopy - energy dispersive X-ray spectroscopy (SEM-EDX) analysis. X-ray diffraction and differential scanning calorimetric analysis confirmed the amorphous nature of EPS, which was comprised of an average particle size of 13.969μm (d 0.5) with 181nm average roughness. Two monosaccharide constituents, galactose (73.13%) and glucose (26.87%) were detected by gas chromatography-mass spectroscopy analysis. Thermal gravimetric analysis revealed that degradation of EPS obtained from A. carterae takes place in three steps. The EPS produced by A. carterae was found to be beneficial for the growth of both A. carterae and Bacillus pumilus. The potential heterogeneous properties of EPS may play an important role in harmful algal bloom.

Retinal very long-chain PUFAs: new insights from studies on ELOVL4 protein

Compared with other mammalian tissues, retina is highly enriched in PUFA. Long-chain PUFA (LC-PUFA; C18-C24) are essential FAs that are enriched in the retina and are necessary for maintenance of normal retinal development and function. The retina, brain, and sperm also contain very LC-PUFA (VLC-PUFA; >C24). Although VLC-PUFA were discovered more than two decades ago, very little is known about their biosynthesis and functional roles in the retina. This is due mainly to intrinsic difficulties associated with working on these unusually long polyunsaturated hydrocarbon chains and their existence in small amounts. Recent studies on the FA elongase elongation of very long chain fatty acids-4 (ELOVL4) protein, however, suggest that VLC-PUFA probably play some uniquely important roles in the retina as well as the other tissues. Mutations in the ELOVL4 gene are found in patients with autosomal dominant Stargardt disease. Here, we review the recent literature on VLC-PUFA with special emphasis on the elongases responsible for their synthesis. We focus on a novel elongase, ELOVL4, involved in the synthesis of VLC-PUFA, and the importance of these FAs in maintaining the structural and functional integrity of retinal photoreceptors.