Yasuwo Fukuyo

DOMOIC ACID PRODUCTION IN NITZSCHIA SP. (BACILLARIOPHYCEAE) ISOLATED FROM A SHRIMP-CULTURE POND IN DO SON, VIETNAM

Domoic acid (DA), a neuroexcitatory amino acid, was detected in batch culture of the newly recognized species Nitzschia navis‐varingica Lundholm et Moestrup. The production of DA by this diatom was confirmed by electrospray ionization mass spectrometry. The diatom was collected from a shrimp‐culture pond in Do Son, Vietnam. The production of DA (1.7 pg·cell−1) is within the levels reported for Pseudo‐nitzschia multiseries (Hasle) Hasle. The DA production started during the late exponential growth phase and reached a maximum during the early stationary growth phase. Maximum DA levels in the axenic culture decreased to about half that of the nonaxenic culture (0.9 pg·cell−1 vs. 1.7 pg·cell−1), suggesting that DA production by the new species is influenced by bacteria.

Occurrences of harmful algal blooms (HABs) associated with ocean environments in the South China Sea

Harmful algal blooms (HABs) occur frequently in the South China Sea (SCS), causing enormous economic losses in aquaculture. We analyzed historical HAB records during the period from 1980 to 2003 in SCS. We found that HABs-affected areas have expanded and the frequency of HABs varied during this period. The seasonal and annual variations, as well as causative algal species of HABs are different among the four regions. Areas with frequent HABs include the Pearl River Estuary (China), the Manila Bay (the Philippines), the Masinloc Bay (the Philippines), and the western coast of Sabah (Malaysia). HABs occurred frequently during March–May in the northern region of SCS, May–July in the eastern region, July in the western region, and year-round in the southern region. Among the species that cause HABs, Noctiluca scintillans dominated in the northern region, and Pyrodinium bahamense in the southern and eastern regions. Causative species also varied in different years for the entire SCS. Both P. bahamense and N. scintillans were the dominant species during 1980–2003. Some species not previously recorded formed blooms during 1991–2003, including Phaeocystis globosa, Scrippsiella trochoidea, Heterosigma akashiwo, and Mesodinium rubrum. Variations in HABs are related to various regional conditions, such as a reversed monsoon wind in the entire SCS, river discharges in the northern area, upwelling in Vietnam coastal waters during southwest winds and near Malaysia coastal waters during northeast winds, and eutrophication from coastal aquaculture in the Pearl River estuary, Manila Bay, and Masinloc Bay.

Toxicity of benthic dinoflagellates found in coral reef.

Five species of dinoflagellates and three microalgae of benthic habitat collected in French Polynesia were grown in unialgal state and tested for toxicity on mice and for hemolytic activity. Two diethyl ether soluble toxins (PL toxin-I, II) and one fast-acting toxin soluble in 1-butanlo (PL toxin-III) were found in Prorocentrum lima. The chromatographic behaviors of PL toxin-I and II closely resembled those of scaritoxin and ciguatoxin prepared from ciguateric fishes. Potent hemolytic substance was present in Amphidinium sp. This species was also toxic to mice. No remarkable activity was detected in the following species: Coolia monotis, Amphidinium sp.-2, and Gymnodinium sp. (dinoflagellates); Chloococcopsis sp. and Isochrysis sp. (blue-green algae); and Nitzschia panduriformis (diatom).

Ocean urea fertilization for carbon credits poses high ecological risks

The proposed plan for enrichment of the Sulu Sea, Philippines, a region of rich marine biodiversity, with thousands of tonnes of urea in order to stimulate algal blooms and sequester carbon is flawed for multiple reasons. Urea is preferentially used as a nitrogen source by some cyanobacteria and dinoflagellates, many of which are neutrally or positively buoyant. Biological pumps to the deep sea are classically leaky, and the inefficient burial of new biomass makes the estimation of a net loss of carbon from the atmosphere questionable at best. The potential for growth of toxic dinoflagellates is also high, as many grow well on urea and some even increase their toxicity when grown on urea. Many toxic dinoflagellates form cysts which can settle to the sediment and germinate in subsequent years, forming new blooms even without further fertilization. If large-scale blooms do occur, it is likely that they will contribute to hypoxia in the bottom waters upon decomposition. Lastly, urea production requires fossil fuel usage, further limiting the potential for net carbon sequestration. The environmental and economic impacts are potentially great and need to be rigorously assessed.

Confirmation of domoic acid production of Pseudo-nitzschia multiseries isolated from Ofunato Bay, Japan.

Production of domoic acid (DA), the responsible toxin for amnesic shellfish poisoning, was examined for 44 strains of Pseudo-nitzschia spp. isolated from Ofunato Bay, Japan. Only one strain which was identified as Pseudo-nitzschia multiseries produced DA in a level comparable to Canadian strains. No significant DA was detected in the rest of the strains, indicating that toxic P. multiseries does not bloom in a high density in the bay.

Are prorocentroid dinoflagellates monophyletic? A study of 25 species based on nuclear and mitochondrial genes.

The dinoflagellate sub-class Prorocentrophycidae has a distinct morphology, lacking the typical dinoflagellate cell structure of a clear cingulum and sulcus. It includes species that produce the toxin okadaic acid. Despite its uniqueness, the group has been found polyphyletic in some previous molecular phylogenetic studies. We have re-investigated the phylogeny of this sub-class by culturing and sequencing new strains, comparing sequences from three genes, the mitochondrial cytochrome c oxidase subunit 1 (cox 1) and the nuclear large and small subunit rRNA (LSU and SSU) encoding genes. We analyzed sequences from twenty-five named and two still undescribed species of Prorocentrophycidae. We used newly recognized features of the secondary structure to align regions of the LSU rRNA. The phylogeny based on cox 1 provided the most well-supported tree and showed strong support for the monophyly of prorocentroid dinoflagellates, while the LSU phylogeny was inconclusive. As in previous studies, phylogeny based on SSU shows the group to appear paraphyletic, however, support values were low. Two strongly supported sub-clades were consistently identified. Benthic and planktonic modes appear to have evolved on multiple occasions within both clades of Prorocentriphycidae. The capability to synthesize toxins appears to have arisen early in prorocentroid evolution and, in particular, okadaic acid synthesis is present in some, but not all, members of Clade 2. The D2a region of the LSU rRNA appears to have developed a deletion in three definable steps during prorocentroid evolution. While the phylogenies inferred from the three genes were not congruent, our results give reserved support to the monophyly of the group.

Frequent occurrence of paralytic shellfish poisoning toxins as dominant toxins in marine puffer from tropical water

Considerably high toxicity was detected in marine puffers collected from Masinloc Bay, Philippines. The toxicity was detected in the liver, intestine, muscle and skin. Noteworthy, the specimens, the muscle of which showed high toxicity, appeared in high frequency, indicating that puffers from this area is not safe for human consumption. These puffer specimens contained paralytic shellfish poisoning (PSP) toxins, often as major toxin components, the profile of which was similar to that of freshwater puffers reported from tropical areas. These results indicate that PSP toxins are common in tropical puffers both from marine and freshwater.

SPECIES BOUNDARIES IN THE TOXIC DINOFLAGELLATE PROROCENTRUM LIMA (DINOPHYCEAE, PROROCENTRALES), BASED ON MORPHOLOGICAL AND PHYLOGENETIC CHARACTERS(1).

Wild and cultured specimens of Prorocentrum lima (Ehrenb.) F. Stein from 26 widely different areas in 13 countries were examined in order to determine consistent characters for delimiting species boundaries in this taxon. The morphological characters valve shape, valve size, valve ornamentation, number and shape of valve pores, number and shape of marginal pores, and periflagellar platelets were observed using LM and SEM, and two molecular genetic regions were sequenced. We identified stable morphological characters that were consistent among wild specimens and all cultures, which were valve shape, valve ornamentation, and number and arrangement of periflagellar platelets. All cultures of P. lima identified by these characters formed a monophyletic group in phylogenetic analyses based on the two genes, which, however, included the species Prorocentrum arenarium. P. arenarium was determined to be within the range of morphological variation of P. lima, and therefore we synonymize the two taxa. Within this monophyletic group, P. lima was divided into several subclades in the all phylogenetic analyses. There were no morphological characters specifically related to any one subclade. The subclades appeared to correlate broadly to sample collection regions, suggesting that geographically separated populations may have become genetically distinct within this epi‐benthic species. We have emended species boundaries in P. lima.

Morphological Observation of Alexandrium tanarense (Lebour) Balech, A. catenella (Whedon et Kofoid) Balech and One Related Morphotype (Dinophyceae) in Korea

Twenty-nine culture strains belonging to the genus Alexandrium Halim (Dinophyceae) were established from water column or sediments in Korea. Seventeen isolates were identified as A. tamarense (Lebour) Balech, eight isolates as A. sp. cf. catenella and one as A. catenella (Whedon et Kofoid) Balech according to the presence or absence of a ventral pore, the shape of the posterior sulcal plate and the sulcal width. Three isolates were unable to be identified due to considerable distortion of thecal plates and lack of enough materials, but typical of A. tamarense and/or A. catenella. The overall cell shape of A. tamarense was usually longer than wide. The posterior sulcal plate was definitely longer than wide dorsoventrally, and sulcus extended posteriorly without apparent widening. They were distributed in three major coasts of Korea. In contrast, the cell shape of A. sp.cf. catenella was generally anterior-posteriorly flattened. The transversal axis of the posterior sulcal plate was always longer than the longitudinal, or both axes were nearly equal in length. Its sulcus was broader than that of A. tamarense and widened in the direction of antapex about 1.5 times. This morphotype existed in nearshore and offshore waters of the southern Korea sea. One of A. catenella isolates from Jinhae Bay showed no conspicuous differences with A. sp. cf. catenella except for the consistent absence of a ventral pore.

Protogonyaulax cohorticula, a toxic dinoflagellate found in the Gulf of Thailand

Two clones of Protogonyaulax cohorticula were isolated from the Gulf of Thailand. The extracts of these clones killed mice with typical signs of paralytic shellfish poisoning. The toxicities corresponded to those of strongly toxic clones of P. tamarensis. In the HPLC and electrophoretic analyses, gonyautoxins and saxitoxin were detected. About 80% of the toxins consisted of gonyautoxin I. These results show that P. cohorticula is a toxic species of Protogonyaulax and that it is at least one of the causative organisms of paralytic shellfish poisoning in Thailand.