Harmful Algal Species in the Tebrau Strait: An SEM Observation of the Dinofl agellate Assemblage

Harmful algal bloom (HAB) is a natural phenomenon due to the increase of algal cell density in the water column that subsequently causes deleterious effects to natural environments as well as mankind. HABs in the country mainly occurred when a particular group of dinofl agellate cells proliferate in the eutrophied semi-enclosed coastal water body. In this study, dinofl agellate species composition in the Tebrau Strait was determined by scanning electron microscope (SEM). Plankton samples were collected by a 20-micron plankton net haul at several locations of the strait. Samples were undergone fi xation, serial dehydration and followed by critical point drying. Samples were then observed under a JEOL analytical SEM. Total of 11 dinofl agellate species were identifi ed, with 7 species known to be associated with HABs events. The occurrence of a fi sh-killing unarmoured dinofl agellate, Karlodinium venefi cum was reported for the fi rst time from Malaysian waters. The presence of this and other potentially harmful dinofl agellate species in the strait should be taken seriously by the respective authorities in future expansion of aquaculture industry in the strait

First Record of Marine Dinoflagellate, Alexandrium Tamutum (Dinophyceae) from Malaysia

Several species of dinoflagellates in the genus Alexandrium are known to be toxic, and have been associated with paralytic shellfish poisoning (PSP) in Malaysia. These Alexandrium species showed high morphological similarity among the toxic and non-toxic species, and detailed observation of the thecal plate’s arrangement is required for precise species identification. Co-occurrence of the toxic and non-toxic species has complicated the plankton monitoring of PSP. In this study, a clone of Alexandrium species was established from plankton samples collected from Kota Belud, Sabah. The specimen was observed under epi-fluorescence microscope, and nucleotide sequences of the nuclear-encoded ribosomal RNA gene obtained. Morphologically, the clone showed relatively wide and large sixth precingular plate (6´´) compared to that of A. minutum. The sulcal posterior plate (Sp) is similar to that of A. minutum, which is wider than long. The first apical plate (1´) is irregularly rhomboidal with a small ventral pore (vp) present on its right margin. The morphological characters resembled to the species description of A. tamutum. Phylogenetic analysis of the ITS rDNA region also revealed a monophyly of this clone with other strains of A. tamutum, and separating them from the A. minutum clade. Species-specific sequence signatures of A. tamutum were obtained in silico, which could be as potential oligonucleotide probe regions for species detection by using molecular tool. This represents the first report of A. tamutum found in Malaysian waters

Benthic Harmful Dinoflagellate Assemblages In A Fringing Reef Of Sampadi Island, Sarawak, Malaysia

A study on the presence and relative abundance of benthic harmful algal bloom (BHAB) forming dinoflagellate species was carried out in the coral reefs of Sampadi Island, Sarawak, Malaysia. The study involved deployment of fiberglass screens as an artificial substrate for the benthic epiphytic microalgae. The screens were placed for 24 h above the seafloor along a 100 m transect at 10 m intervals. BHAB species attached to the screens were identified and cell abundances were enumerated under a light microscope. The BHAB community at the study site was dominated by Prorocentrum spp. and Coolia spp. Other BHAB species collected included Amphidinium spp., Gambierdiscus spp. and Ostreopsis spp. Total cell densities collected on the screens ranged from 5 to 100 cells per 100 cm2. The two BHAB groups of primary concern, Gambierdiscus spp. and Ostreopsis spp. were detected at relatively low abundances of 0.6–4.2% and 1.8–16% respectively. This study has shown that potentially toxic BHAB species were present in the coral reef and the artificial substrate approach could provide a convenient quantitative method for the collection of clean samples for identification and enumeration purposes

Character evolution of the benthic thecate dinoflagellate, gambierdiscus (Dinophyceae), with an introduction of the interactive key to species

The genus Gambierdiscus is one of the benthic marine dinoflagellates that are known to produce biotoxins, causing ciguatera food poisoning (CFP) in humans after consuming the contaminated marine fishes. These benthic marine dinoflagellates are commonly found in the warm tropical and subtropical waters. The phenomena associated with high cell abundance or “blooms” is referred to Benthic Harmful Algal Blooms (BHABs). Precise species identification is crucial as not all of the species in the genus are toxic. In this study, morphological characteristics of Gambierdiscus species widely used in species identification were analyzed, and the character states coded. Taxon sampling on the large subunit (LSU) rDNA of all Gambierdiscus species were carried out, and used for the phylogenetic reconstruction. The character states were mapped onto the Maximum Parsimony (MP) tree to investigate the character state evolution of Gambierdiscus species. Morphological information and the distribution of the 13 species were used to develop a comprehensive taxonomic database of Gambierdiscus, a web-based interactive identification key for species identification is presented

A bloom of Karlodinium australe (Gymnodiniales, Dinophyceae) associated with mass mortality of cage-cultured fishes in West Johor Strait, Malaysia

A recent (February 2014) mass mortality of fishes was observed in the cage-farming region of the West Johor Strait of Malaysia, involving over four different species of cultured fishes, numbering ∼50,000 fish. A field investigation at six stations along the West Johor Strait collected water samples and examined for the presence of harmful species. Dead fishes were collected for necropsy. The phytoplankton composition was dominated by a species of Karlodinium, at a considerably high cell concentration (0.31–2.34 × 106 cells l−1), and constituting 68.8–98.6% of the phytoplankton relative abundance at all stations. Detailed morphological assessment by light and scanning electron microscopy revealed that the species was Karlodinium australe de Salas, Bolch and Hallegraeff. This was supported by molecular evidence of the nuclear encoded large subunit ribosomal gene (LSU rDNA) and the second internal transcribed spacer (ITS2) via single-cell PCR. The sequences of LSU rDNA yielded 3.6–4.0% divergence when compared to the sister taxon, K. armiger; and >6.5% when compared to other Karlodinium species. Fish necropsy showed symptoms similar to those affected by karlotoxin ichthyotoxins. This is the first report of a mass mortality of cage-cultured and wild fishes attributed to the unarmored dinoflagellate K. australe

Jerantinine A induces tumor-specific cell death through modulation of splicing factor 3b subunit 1 (SF3B1)

Precursor mRNA (pre-mRNA) splicing is catalyzed by a large ribonucleoprotein complex known as the spliceosome. Numerous studies have indicated that aberrant splicing patterns or mutations in spliceosome components, including the splicing factor 3b subunit 1 (SF3B1), are associated with hallmark cancer phenotypes. This has led to the identification and development of small molecules with spliceosome-modulating activity as potential anticancer agents. Jerantinine A (JA) is a novel indole alkaloid which displays potent anti-proliferative activities against human cancer cell lines by inhibiting tubulin polymerization and inducing G2/M cell cycle arrest. Using a combined pooled-genome wide shRNA library screen and global proteomic profiling, we showed that JA targets the spliceosome by up-regulating SF3B1 and SF3B3 protein in breast cancer cells. Notably, JA induced significant tumor-specific cell death and a significant increase in unspliced pre-mRNAs. In contrast, depletion of endogenous SF3B1 abrogated the apoptotic effects, but not the G2/M cell cycle arrest induced by JA. Further analyses showed that JA stabilizes endogenous SF3B1 protein in breast cancer cells and induced dissociation of the protein from the nucleosome complex. Together, these results demonstrate that JA exerts its antitumor activity by targeting SF3B1 and SF3B3 in addition to its reported targeting of tubulin polymerization

Marine micro-phytoplankton of Singapore, with a review of harmful microalgae in the region

A survey of marine phytoplankton in the Singapore Strait was carried out between May and June 2013, as part of an effort to determine the diversity of phytoplankton in Singapore’s coastal waters. A total of 34 microalgal samples were collected using a 20 μm-mesh plankton net and from coastal sediments. Living samples and preserved samples in Lugol’s solution were identified to species as far as possible under the microscope. A checklist of marine micro-phytoplankton was updated to encompass 270 taxa, including 49 new records from Singapore waters. Some 37 species from 15 families were dinoflagellates, and 233 species from 50 families were diatoms. Harmful microalgae, categorized as biotoxin-producers and fish killers, were also found in this survey. These were in the genera Alexandrium, Amphidinium, Ceratium, Cochlodinium, Coolia, Dinophysis, Gambierdiscus, Karenia, Karlodinium, Ostreopsis, Prorocentrum, Nitzschia, and Pseudo-nitzschia

New scenario for speciation in the benthic dinoflagellate genus Coolia (Dinophyceae)

In this study, inter- and intraspecific genetic diversity within the marine harmful dinoflagellate genus Coolia Meunier was evaluated using isolates obtained from the tropics to subtropics in both Pacific and Atlantic Ocean basins. The aim was to assess the phylogeographic history of the genus and to clarify the validity of established species including Coolia malayensis. Phylogenetic analysis of the D1-D2 LSU rDNA sequences identified six major lineages (L1–L6) corresponding to the morphospecies Coolia malayensis (L1), C. monotis (L2), C. santacroce (L3), C. palmyrensis (L4), C. tropicalis (L5), and C. canariensis (L6). A median joining network (MJN) of C. malayensis ITS2 rDNA sequences revealed a total of 16 haplotypes; however, no spatial genetic differentiation among populations was observed. These MJN results in conjunction with CBC analysis, rDNA phylogenies and geographical distribution analyses confirm C. malayensis as a distinct species which is globally distributed in the tropical to warm-temperate regions. A molecular clock analysis using ITS2 rDNA revealed the evolutionary history of Coolia dated back to the Mesozoic, and supports the hypothesis that historical vicariant events in the early Cenozoic drove the allopatric differentiation of C. malayensis and C. monotis

Clearance rate of two commercially important bivalves feeding on pseudo-nitzschia species (Bacillariophyceae)

The chain-forming pennate diatom, Pseudo-nitzschia is a cosmopolitan diatom. Contamination of domoic acid (DA), derived from several toxic species of Pseudo-nitzschia in filtered feeding bivalves, has subsequently caused seafood poisoning which is commonly known as amnesic shellfish poisoning (ASP). In this study, a feeding experiment was carried out to determine the clearance rates (CR) of two commercial bivalves, blood cockles, Anadara granosa and benthic clams, Polymesoda expansa feeding on Pseudo-nitzschia species. Mass clonal culture of Pseudo-nitzschia sp. strain PnMt43 from Muara Tebas was used as prey in this experiment. The bivalves were fed with different initial cell density ranging from 100 to 5000 cell mL-1. Subsamples were taken half-hourly for a period of six hours for cell counts. Our results showed two distinct clearance responses between the two bivalves (P = 0.0020). P. expansa had the higher CR with 7.316 cell gram-1 mL-1 day than A.granosa with 0.389 cell gram-1 mL-1. The half-saturation concentrations (K) for A. granosa and P. expansa were 955 and 437 cell mL-1, respectively. The higher clearance rate of P. expansa indicates the tendency to accumulate higher level of toxin during a Pseudo-nitzschia bloom. However, P. expansa with higher CR has lower K due to the difference in consumption pattern. Simulation of toxin kinetic of the two bivalves was not able to be conducted due to the absence of toxic Pseudo-nitzschia strains. Further studies on feeding responses and toxin accumulation-depuration rates are important to understand the mechanism of toxin kinetic of DA in these two commercially important bivalves

Harmful marine dinoflagellates (Dinophyceae) of Malaysia including new records of taxa based on morphology and molecular

Dinoflagellates are eukaryotic biflagellated, unicellular organisms that can be found ,in both freshwater and marine habitats. Several species are harmful as they produce potent neurotoxins; cause unusual blooming phenomena in the marine environment. In Malaysia, harmful algal blooms (HABs) event was first reported in 1976. Since then several studies on HABs have been undertaken, focusing mainly in areas where poisoning cases were reported.) In this study, samplings were conducted in several locations including sites where' no poisoning case has been reported, but intensive aquaculture activity is ongoing. Plankton and seaweed samples were collected and brought back to the laboratory. Single-cell of dinoflagellates were isolated and clonal cultures established. Both field and cultured specimens were observed under light and scanning electron microscopy (SEM). Identification to species level was based on detailed investigation of the thecal plates. Nucleotide sequences of the nuclearencoded ribosomal DNA were obtained from clonal cultures, and preliminary toxin analysis was performed by ELISA and LC-MS. A total of 46 taxa of harmful and/or potentially harmful dinoflagellates were identified and documented from four locations (Cherating, Pahang, Port Dickson, Negeri Sembi lan, Tebrau Strait, Johor, and Kuching, Sarawak). Two taxa are new records of occurrence in Malaysian waters, namely Karlodinium veneficum from Tebrau Strait and Protoceratiumfukuyoii sp. nov from Semariang. Notably the PST-producing dinoflagellates, Alexandrium lamiyavanichii and Pyrodinium bahamense were found, for the first time, in Sarawak waters and the Straits of Malacca, respectively. Molecular data obtained from the present study, and the molecular signatures identified for each species will serve as preliminary data for DNA barcoding of the harmful dinoflagellates. Inventory of the harmful dinoflagellate species wiIl provide useful baseline infonnation of the harmful species present in Malaysian waters. Furthermore, distribution data on harmful dinoflagellates could be used to identify the hotspots of potential HABs incidences for early warning and HABs monitoring purposes