Kelimpahan Dinoflagellata Bentik Berbahaya di Habitat Lamun dan Makroalga di Pulau Pari, Kepulauan Seribu, Indonesia

<p class="Papertext"><strong>The Abundance of the Harmful Benthic Dinoflagellate in the Seagrass and Macroalgae Habitats on Pari Island, Thousand Islands, Indonesia. </strong>Benthic dinoflagellates are an important part of benthic ecosystems and part of their ecological functions. However, ciguatoxin (CTX) producer dinoflagellates, which could cause Ciguatera Fish Poisoning (CFP) disease, is a known threat to marine biota and the island’s coastal communities, such as in Pari Island. Thus, this research aimed is to study the population of five harmful and toxin producer benthic dinoflagellate genera, namely <span style="text-decoration: underline;">Amphidinium</span>, <span style="text-decoration: underline;">Coolia</span>, <span style="text-decoration: underline;">Gambierdiscus</span>, <span style="text-decoration: underline;">Ostreopsis</span>, and <span style="text-decoration: underline;">Prorocentrum</span>, in macroalgae and seagrass habitats of Pari Island. Benthic dinoflagellate samples were collected in Juni 2019 using artificial substrates in the form of a 10×15 cm nylon screen. Environmental variables, such as nutrient concentration (nitrogen and phosphorus) and water temperature were also measured. The results showed that the average benthic dinoflagellate density in seagrass habitats was four times higher compared to the macroalgae habitats. <em>Gambierdiscus</em> were found as the most abundant benthic dinoflagellate in seagrass habitats, while <em>Prorocentrum</em> were more abundant in macroalgae habitats. Principal Component Analysis (PCA) showed that the N:P ratio was the most important regulating parameter for most benthic dinoflagellate genera in this study, except <em>Ostreopsis</em>, which was found to be more abundant in phosphorus-rich macroalga habitats. The data also shows an increase of benthic dinoflagellate cell density up to seven times compared to the previous research data in 2012 and 2013. That trend indicates an increase in the disturbance and pressure on the macroalgae and seagrass habitats of Pari Island, which require special attention to prevent the occurrence of toxic Benthic Harmful Algal Blooms (bHABs) and CFP cases in the island.</p>

Amphidinium stirisquamtum sp. nov. (Dinophyceae), a new marine sand-dwelling dinoflagellate with a novel type of body scale

Amphidinium species are amongst the most abundant benthic dinoflagellates in marine intertidal sandy ecosystems. Some of them produce a variety of bioactive compounds that have both harmful effects and pharmaceutical potential. In this study, Amphidinium cells were isolated from intertidal sand collected from the East China Sea. The two strains established were subjected to detailed examination by light, and scanning and transmission electron microscopy. The vegetative cells had a minute, irregular, and triangular-shaped epicone deflected to the left, thus fitting the description of Amphidinium sensu stricto. These strains are distinguished from other Amphidinium species by combination characteristics: (1) longitudinal flagellum inserted in the lower third of the cell; (2) icicle-shaped scales, 276 ± 17 nm in length, on the cell body surface; (3) asymmetrical hypocone with the left side longer than the right; and (4) presence of immotile cells. Therefore, they are described here as Amphidinium stirisquamtum sp. nov. The molecular tree inferred from small subunit rRNA, large subunit rRNA, and internal transcribed spacer-5.8S sequences revealed that A. stirisquamtum is grouped together with the type species of Amphidinium, A. operculatum, in a fully supported clade, but is distantly related to other Amphidinium species bearing body scale. Live A. stirisquamtum cells greatly affected the survival of rotifers and brine shrimp, their primary grazers, making them more susceptible to predation by the higher tropic level consumers in the food web. This will increase the risk of introducing toxicity, and consequently, the bioaccumulation of toxins through marine food webs.

Depuration Kinetics and Growth Dilution of Caribbean Ciguatoxin in the Omnivore Lagodon rhomboides: Implications for Trophic Transfer and Ciguatera Risk

Modeling ciguatoxin (CTX) trophic transfer in marine food webs has significant implications for the management of ciguatera poisoning, a circumtropical disease caused by human consumption of CTX-contaminated seafood. Current models associated with CP risk rely on modeling abundance/presence of CTX-producing epi-benthic dinoflagellates, e.g., Gambierdiscus spp., and are based on studies showing that toxin production is site specific and occurs in pulses driven by environmental factors. However, food web models are not yet developed and require parameterizing the CTX exposure cascade in fish which has been traditionally approached through top-down assessment of CTX loads in wild-caught fish. The primary goal of this study was to provide critical knowledge on the kinetics of C-CTX-1 bioaccumulation and depuration in the marine omnivore Lagodon rhomboides. We performed a two-phase, 17 week CTX feeding trial in L. rhomboides where fish were given either a formulated C-CTX-1 (n = 40) or control feed (n = 37) for 20 days, and then switched to a non-toxic diet for up to 14 weeks. Fish were randomly sampled through time with whole muscle, liver, and other pooled viscera dissected for toxin analysis by a sodium channel-dependent MTT-based mouse neuroblastoma (N2a) assay. The CTX levels measured in all tissues increased with time during the exposure period (days 1 to 20), but a decrease in CTX-specific toxicity with depuration time only occurred in viscera extracts. By the end of the depuration, muscle, liver, and viscera samples had mean toxin concentrations of 189%, 128%, and 42%, respectively, compared to fish sampled at the start of the depuration phase. However, a one-compartment model analysis of combined tissues showed total concentration declined to 56%, resulting in an approximate half-life of 97 d (R2 = 0.43). Further, applying growth dilution correction models to the overall concentration found that growth was a major factor reducing C-CTX concentrations, and that the body burden was largely unchanged, causing pseudo-elimination and a half-life of 143–148 days (R2 = 0.36). These data have important implications for food web CTX models and management of ciguatera poisoning in endemic regions where the frequency of environmental algal toxin pulses may be greater than the growth-corrected half-life of C-CTX in intermediate-trophic-level fish with high site fidelity.

Ciguatoxin-Producing Dinoflagellate Gambierdiscus in the Beibu Gulf: First Report of Toxic Gambierdiscus in Chinese Waters

Ciguatera poisoning is mainly caused by the consumption of reef fish that have accumulated ciguatoxins (CTXs) produced by the benthic dinoflagellates Gambierdiscus and Fukuyoa. China has a long history of problems with ciguatera, but research on ciguatera causative organisms is very limited, especially in the Beibu Gulf, where coral reefs have been degraded significantly and CTXs in reef fish have exceeded food safety guidelines. Here, five strains of Gambierdiscus spp. were collected from Weizhou Island, a ciguatera hotspot in the Beibu Gulf, and identified by light and scanning electron microscopy and phylogenetic analyses based on large and small subunit rDNA sequences. Strains showed typical morphological characteristics of Gambierdiscus caribaeus, exhibiting a smooth thecal surface, rectangular-shaped 2′, almost symmetric 4′’, and a large and broad posterior intercalary plate. They clustered in the phylogenetic tree with G. caribaeus from other locations. Therefore, these five strains belonged to G. caribaeus, a globally distributed Gambierdiscus species. Toxicity was determined through the mouse neuroblastoma assay and ranged from 0 to 5.40 fg CTX3C eq cell−1. The low level of toxicity of G. caribaeus in Weizhou Island, with CTX-contaminated fish above the regulatory level in the previous study, suggests that the long-term presence of low toxicity G. caribaeus might lead to the bioaccumulation of CTXs in fish, which can reach dangerous CTX levels. Alternatively, other highly-toxic, non-sampled strains could be present in these waters. This is the first report on toxic Gambierdiscus from the Beibu Gulf and Chinese waters and will provide a basis for further research determining effective strategies for ciguatera management in the area.

Detection of the Benthic Dinoflagellates, Ostreopsis cf. ovata and Amphidinium massartii (Dinophyceae), Using Loop-Mediated Isothermal Amplification

For the in situ and sensitive detection of benthic dinoflagellates, we have established an integrated loop-mediated isothermal amplification (LAMP) assay based on Ostreopsis cf. ovata and Amphidinium massartii. To detect the two species, a set of species-specific primers was constructed between the ITS gene and D1–D6 LSU gene, and the reaction temperature, time, and buffer composition were optimized to establish this method. In addition, the specificity of the LAMP primers was verified both in strains established in the laboratory and in field samples collected from the Jeju coastal waters, Korea. With the LAMP assay, the analysing time was within 45 to 60 min, which may be shorter than that with the conventional PCR. The detection sensitivity of the LAMP assay for O. cf. ovata or A. massartii was comparable to other molecular assays (PCR and quantitative PCR (qPCR)) and microscopy examination. The detection limit of LAMP was 0.1 cell of O. cf. ovata and 1 cell of A. massartii. The optimized LAMP assay was successfully applied to detect O. cf. ovata and A. massartii in field samples. Thus, this study provides an effective method for detecting target benthic dinoflagellate species, and could be further implemented to monitor phytoplankton in field surveys as an altenative.

Critical Review and Conceptual and Quantitative Models for the Transfer and Depuration of Ciguatoxins in Fishes

We review and develop conceptual models for the bio-transfer of ciguatoxins in food chains for Platypus Bay and the Great Barrier Reef on the east coast of Australia. Platypus Bay is unique in repeatedly producing ciguateric fishes in Australia, with ciguatoxins produced by benthic dinoflagellates (Gambierdiscus spp.) growing epiphytically on free-living, benthic macroalgae. The Gambierdiscus are consumed by invertebrates living within the macroalgae, which are preyed upon by small carnivorous fishes, which are then preyed upon by Spanish mackerel (Scomberomorus commerson). We hypothesise that Gambierdiscus and/or Fukuyoa species growing on turf algae are the main source of ciguatoxins entering marine food chains to cause ciguatera on the Great Barrier Reef. The abundance of surgeonfish that feed on turf algae may act as a feedback mechanism controlling the flow of ciguatoxins through this marine food chain. If this hypothesis is broadly applicable, then a reduction in herbivory from overharvesting of herbivores could lead to increases in ciguatera by concentrating ciguatoxins through the remaining, smaller population of herbivores. Modelling the dilution of ciguatoxins by somatic growth in Spanish mackerel and coral trout (Plectropomus leopardus) revealed that growth could not significantly reduce the toxicity of fish flesh, except in young fast-growing fishes or legal-sized fishes contaminated with low levels of ciguatoxins. If Spanish mackerel along the east coast of Australia can depurate ciguatoxins, it is most likely with a half-life of ≤1-year. Our review and conceptual models can aid management and research of ciguatera in Australia, and globally.

Asynchrony of Gambierdiscus spp. Abundance and Toxicity in the U.S. Virgin Islands: Implications for Monitoring and Management of Ciguatera

Ciguatera poisoning (CP) poses a significant threat to ecosystem services and fishery resources in coastal communities. The CP-causative ciguatoxins (CTXs) are produced by benthic dinoflagellates including Gambierdiscus and Fukuyoa spp., and enter reef food webs via grazing on macroalgal substrates. In this study, we report on a 3-year monthly time series in St. Thomas, US Virgin Islands where Gambierdiscus spp. abundance and Caribbean-CTX toxicity in benthic samples were compared to key environmental factors, including temperature, salinity, nutrients, benthic cover, and physical data. We found that peak Gambierdiscus abundance occurred in summer while CTX-specific toxicity peaked in cooler months (Feb–May) when the mean water temperatures were approximately 26–28 °C. These trends were most evident at deeper offshore sites where macroalgal cover was highest year-round. Other environmental parameters were not correlated with the CTX variability observed over time. The asynchrony between Gambierdiscus spp. abundance and toxicity reflects potential differences in toxin cell quotas among Gambierdiscus species with concomitant variability in their abundances throughout the year. These results have significant implications for monitoring and management of benthic harmful algal blooms and highlights potential seasonal and highly-localized pulses in reef toxin loads that may be transferred to higher trophic levels.

Toxicity Bioassay and Cytotoxic Effects of the Benthic Marine Dinoflagellate Amphidinium operculatum

Benthic dinoflagellates produce a wide array of bioactive compounds, primarily polyketides, that cause toxic effects on human consumers of seafood and perhaps mediate species interactions in the benthic microenvironment. This study assesses toxic and other bioactive effects of the benthic dinoflagellate Amphidinium operculatum (strain AA60) in two targeted bioassays. The brine shrimp (Artemia salina) bioassay revealed lethal effects of direct exposure to live dinoflagellate cells (Treatment A) and even higher potency with ethanolic extracts of lysed cells (Treatment D). There were no inimical bioactive effects of components released to the aqueous growth medium (Treatment B) or from aqueous cell lysates (Treatment C). The hypothesis that released bioactive compounds provide a chemical defense against metazoan grazers is therefore not supported by these results. The cytotoxic effect of ethanolic crude extracts of this dinoflagellate exhibited mild to high growth reduction effects on six human cancer cell lines. In particular, crude cell-free extracts proved highly growth-inhibitory activity towards breast and lung cancer cell lines MCF-7 and SKLU-1, respectively. Preliminary anti-cancer results indicate that natural bioactive compounds from Amphidinium are worthy of structural characterization and further toxicological investigation as potential therapeutants.

Morphotype diversity of Prorocentrum lima in the western part of Indonesian waters

Widiarti R, Zamani NP, Bengen DG, Madduppa H. 2021. Morphotype diversity of Prorocentrum lima in the western part of Indonesian waters. Biodiversitas 22: 609-616. Prorocentrum lima is one of the toxic benthic dinoflagellates, known to produce various toxins, including okadaic acid and dinophysis toxins. The species have a wide range of morphological variability, and possess morphotype diversity, which makes it essential to have detailed morphology observation for identification and other purposes. However, such comprehensive observation has never been reported from Indonesian waters. This study aims to determine the morphological characteristics (including pore size and number of pores) of P. lima morphotypes, in the western part of Indonesian waters (Bintan Island, Belitung Island, Seribu Islands, and Karimunjawa Islands). The results showed three different and unique morphotypes of P. lima, namely morphotype 1, morphotype 2, and morphotype 3. Three clusters were presented by Cluster Analysis, corresponded to the three morphotypes, which were Belitung Island clusters, Seribu Islands and Karimunjawa Islands cluster, and Bintan Island cluster, respectively. These findings support distribution of P. lima and its potential risk of toxicity in Indonesian waters, which prompts the necessity of conducting future research, to avoid the negative impact.