Molecular diversity and assemblages of coral symbionts (Symbiodiniaceae) in diverse scleractinian coral species.

The scleractinian coral-associated symbiotic algae Symbiodiniaceae plays an important role in bleaching tolerance and coral resilience. In this study, coral-associated Symbiodiniaceae communities of 14 reef sites of Perhentian and Redang Islands Marine Parks (Malaysia, South China Sea) were characterized using the high-throughput next-generation amplicon sequencing on the ITS2 rDNA marker to inventory the Symbiodiniaceae diversity from a healthy tropical reef system and to generate a baseline for future studies. A total of 64 coral-Symbiodiniaceae associations were characterized in 18 genera (10 families) of scleractinian corals using the SymPortal analytical framework. The results revealed the predominance of Symbiodiniaceae genera Cladocopium (average 82%) and Durusdinium (18%), while Symbiodinium, Breviolum, Fugacium, and Gerakladium were found as minor groups (<0.01%). Of the 39 Cladocopium and Durusdinium major ITS2 sequences, 14 were considered dominant/sub-dominant, with C3u as the predominant type (63.3%), followed by D1 (15%), C27 (10.1%), and C15 (6.9%). A total of 19 and 13 Cladocopium and Durusdinium ITS2-type profiles were detected across the coral species, respectively. Symbiodiniaceae diversity and richness recorded in this study were higher when compared to other reefs in the proximity. With the increasing coral-Symbiodiniaceae associations archived, the database would provide a baseline to assess the changes of Symbiodiniaceae communities in the coral hosts and to explore the potential adaptive roles of this coral-algal association.

Identification of Fish Species and Toxins Implicated in a Snapper Food Poisoning Event in Sabah, Malaysia, 2017.

In the coastal countries of Southeast Asia, fish is a staple diet and certain fish species are food delicacies to local populations or commercially important to individual communities. Although there have been several suspected cases of ciguatera fish poisoning (CFP) in Southeast Asian countries, few have been confirmed by ciguatoxins identification, resulting in limited information for the correct diagnosis of this food-borne disease. In the present study, ciguatoxin-1B (CTX-1B) in red snapper (Lutjanus bohar) implicated in a CFP case in Sabah, Malaysia, in December 2017 was determined by single-quadrupole selected ion monitoring (SIM) liquid chromatography/mass spectrometry (LC/MS). Continuous consumption of the toxic fish likely resulted in CFP, even when the toxin concentration in the fish consumed was low. The identification of the fish species was performed using the molecular characterization of the mitochondrial cytochrome c oxidase subunit I gene marker, with a phylogenetic analysis of the genus Lutjanus. This is the first report identifying the causative toxin in fish-implicated CFP in Malaysia.

Diverse harmful microalgal community assemblages in the Johor Strait and the environmental effects on its community dynamics.

Coastal ecosystems are often subjected to anthropogenic disturbances that lead to water quality deterioration and an increase in harmful algal bloom (HAB) events. Using the next-generation molecular tool of 18S rDNA metabarcoding, we examined the community assemblages of HAB species in the Johor Strait, Malaysia between May 2018 and September 2019, covering 19 stations across the strait. The molecular operational taxonomic units (OTUs) of HAB taxa retrieved from the dataset (n = 194) revealed a much higher number of HAB taxa (26 OTUs) than before, with 12 taxa belong to new records in the strait. As revealed in the findings of this study, the diversity and community structure of HAB taxa varied significantly over time and space. The most common and abundant HAB taxa in the strait (frequency of occurrence >70%) comprised Heterosigma akashiwo, Fibrocapsa japonica, Pseudo-nitzschia pungens, Dinophysis spp., Gymnodinium catenatum, Alexandrium leei, and A. tamiyavanichii. Also, our results demonstrated that the HAB community assemblages in the strait were dependent on the interplay of environmental variables that influence by the monsoonal effects. Different HAB taxa, constitute various functional types, occupied and prevailed in different environmental niches across space and time, leading to diverse community assemblages and population density. This study adds to the current understandings of HAB dynamics and provides a robust overview of temporal-spatial changes in HAB community assemblages along the environmental gradients in a tropical eutrophic coastal ecosystem.

Effects of substratum and depth on benthic harmful dinoflagellate assemblages.

Microhabitats influence the distribution and abundance of benthic harmful dinoflagellate (BHAB) species. Currently, much of the information on the relationships between BHABs and microhabitat preferences is based on non-quantitative anecdotal observations, many of which are contradictory. The goal of this study was to better quantify BHAB and microhabitat relationships using a statistically rigorous approach. Between April 2016 to May 2017, a total of 243 artificial substrate samplers were deployed at five locations in the Perhentian Islands, Malaysia while simultaneous photo-quadrat surveys were performed to characterize the benthic substrates present at each sampling site. The screen samplers were retrieved 24 h later and the abundances of five BHAB genera, Gambierdiscus, Ostreopsis, Coolia, Amphidinium, and Prorocentrum were determined. Substrate data were then analyzed using a Bray-Curtis dissimilarity matrix to statistically identify distinct microhabitat types. Although BHABs were associated with a variety of biotic and abiotic substrates, the results of this study demonstrated differing degrees of microhabitat preference. Analysis of the survey results using canonical correspondence analysis explained 70.5% (horizontal first axis) and 21.6% (vertical second axis) of the constrained variation in the distribution of various genera among microhabitat types. Prorocentrum and Coolia appear to have the greatest range being broadly distributed among a wide variety of microhabitats. Amphidinium was always found in low abundances and was widely distributed among microhabitats dominated by hard coral, turf algae, sand and silt, and fleshy algae and reached the highest abundances there. Gambierdiscus and Ostreopsis had more restricted distributions. Gambierdiscus were found preferentially associated with turf algae, hard coral and, to a lesser extent, fleshy macroalgae microhabitats. Ostreopsis, almost always more abundant than Gambierdiscus, preferred the same microhabitats as Gambierdiscus and were found in microbial mats as well. With similar habitat preferences Ostreopsis may serve as an indicator organism for the presence of Gambierdiscus. This study provides insight into how BHAB-specific microhabitat preferences can affect toxicity risks.

Growth and epiphytic behavior of three Gambierdiscus species (Dinophyceae) associated with various macroalgal substrates.

Species of the benthic dinoflagellate Gambierdiscus produce polyether neurotoxins that caused ciguatera fish/shellfish poisoning in human. The toxins enter marine food webs by foraging of herbivores on the biotic substrates like macroalgae that host the toxic dinoflagellates. Interaction of Gambierdiscus and their macroalgal substrate hosts is believed to shape the tendency of substrate preferences and habitat specialization. This was supported by studies that manifested epiphytic preferences and behaviors in Gambierdiscus species toward different macroalgal hosts. To further examine the supposition, a laboratory-based experimental study was conducted to examine the growth, epiphytic behaviors and host preferences of three Gambierdiscus species towards four macroalgal hosts over a culture period of 40 days. The dinoflagellates Gambierdiscus balechii, G. caribaeus, and a new ribotype, herein designated as Gambierdiscus type 7 were initially identified based on the thecal morphology and molecular characterization. Our results showed that Gambierdiscus species tested in this study exhibited higher growth rates in the presence of macroalgal hosts. Growth responses and attachment behaviors, however, differed among different species and strains of Gambierdiscus over different macroalgal substrate hosts. Cells of Gambierdiscus mostly attached to substrate hosts at the beginning of the experiments but detached at the later time. Localized Gambierdiscus-host interactions, as demonstrated in this study, could help to better inform efforts of sampling and monitoring of this benthic toxic dinoflagellate.

Taxonomy and toxicity of Prorocentrum from Perhentian Islands (Malaysia), with a description of a non-toxigenic species Prorocentrum malayense sp. nov. (Dinophyceae).

Thirteen isolates of Prorocentrum species were established from the coral reefs of Perhentian Islands Marine Park, Malaysia and underwent morphological observations and molecular characterization. Six species were found: P. caipirignum, P. concavum, P. cf. emarginatum, P. lima, P. mexicanum and a new morphotype, herein designated as P. malayense sp. nov. Prorocentrum malayense, a species closely related to P. leve, P. cf. foraminosum, P. sp. aff. foraminossum, and P. concavum (Clade A sensu Chomérat et al. 2018), is distinguished from its congeners as having larger thecal pore size and a more deeply excavated V-shaped periflagellar area. Platelet arrangement in the periflagellar area of P. malayense is unique, with the presence of platelet 1a and 1b, platelet 2 being the most anterior platelet, and a broad calabash-shaped platelet 3. The species exhibits consistent genetic sequence divergences for the nuclear-encoded large subunit ribosomal RNA gene (LSU rDNA) and the second internal transcribed spacer (ITS2). The phylogenetic inferences further confirmed that it represents an independent lineage, closely related to species in Clade A sensu Chomérat et al. Pairwise comparison of ITS2 transcripts with its closest relatives revealed the presence of compensatory base changes (CBCs). Toxicity analysis showed detectable levels of okadaic acid in P. lima (1.0-1.6 pg cell-1) and P. caipirignum (3.1 pg cell-1); this is the first report of toxigenic P. caipirignum in the Southeast Asian region. Other Prorocentrum species tested, including the new species, however, were below the detection limit.

Habitat complexity affects benthic harmful dinoflagellate assemblages in the fringing reef of Rawa Island, Malaysia.

Few studies have investigated the effect of fine-scale habitat differences on the dynamics of benthic harmful dinoflagellate assemblages. To determine how these microhabitat differences affect the distribution and abundance of the major benthic harmful dinoflagellate genera in a tropical coral reef ecosystem, a field study was undertaken between April-September 2015 and January 2016 on the shallow reef flat of the fringing reef of Rawa Island, Terengganu, Malaysia. Sampling of benthic dinoflagellates was carried out using an artificial substrate sampling method (fiberglass screens). Benthic microhabitats surrounding the sampling screens were characterized simultaneously from photographs of a 0.25-m2 quadrat based on categories of bottom substrate types. Five taxonomic groups of benthic dinoflagellates, Ostreopsis, Gambierdiscus, Prorocentrum, Amphidinium, and Coolia were identified, and cells were enumerated using a light microscope. The results showed Gambierdiscus was less abundant than other genera throughout the study period, with maximum abundance of 1.2 × 103 cells 100 cm-2. While most taxa were present on reefs with high coral cover, higher cell abundances were observed in reefs with high turf algal cover and coral rubble, with the exception of Ostreopsis, where the abundance reached a maximum of 3.4 × 104 cells 100 cm-2 in habitats with high coral cover. Microhabitat heterogeneity was identified as a key factor governing the benthic harmful dinoflagellate assemblages and may account for much of the observed variability in dominant taxa. This finding has significant implications for the role of variability in the benthic harmful algal bloom (BHAB) outbreaks and the potential in identifying BHAB-related toxin transfer pathways and the key vectors in the food webs.

Responses of phytoplankton community to eutrophication in Semerak Lagoon (Malaysia).

Effects of aquaculture activities on the environmental parameters and phytoplankton community structure were investigated in a semi-enclosed lagoon located at Semerak River, Malaysia. Elevated concentrations of phosphate and ammonia were observed at the aquaculture area and the inner lagoon. Relatively low dissolved oxygen, high total chlorophyll a, and high phytoplankton abundances but low species richness were recorded. Chaetoceros, Pseudo-nitzschia brasiliana, Blixaea quinquecornis, and Skeletonema blooms were observed, and some were associated with anoxia condition. Eutrophication level assessed by UNTRIX suggests that the water quality in the lagoon is deteriorating. Dissolved inorganic phosphorus and nitrogen at the impacted area were 15 and 12 times higher than the reference sites, respectively. Such trophic status indices could provide a useful guideline for optimal aquaculture management plan to reduce the environmental impact caused by aquaculture.

Pentaplacodinium saltonense gen. et sp. nov. (Dinophyceae) and its relationship to the cyst-defined genus Operculodinium and yessotoxin-producing Protoceratium reticulatum.

Strains of a dinoflagellate from the Salton Sea, previously identified as Protoceratium reticulatum and yessotoxin producing, have been reexamined morphologically and genetically and Pentaplacodinium saltonense n. gen. et sp. is erected to accommodate this species. Pentaplacodinium saltonense differs from Protoceratium reticulatum (Claparède et Lachmann 1859) Bütschli 1885 in the number of precingular plates (five vs. six), cingular displacement (two widths vs. one), and distinct cyst morphology. Incubation experiments (excystment and encystment) show that the resting cyst of Pentaplacodinium saltonense is morphologically most similar to the cyst-defined species Operculodinium israelianum (Rossignol, 1962) Wall (1967) and O. psilatum Wall (1967). Collections of comparative material from around the globe (including Protoceratium reticulatum and the genus Ceratocorys) and single cell PCR were used to clarify molecular phylogenies. Variable regions in the LSU (three new sequences), SSU (12 new sequences) and intergenic ITS 1-2 (14 new sequences) were obtained. These show that Pentaplacodinium saltonense and Protoceratium reticulatum form two distinct clades. Pentaplacodinium saltonense forms a monophyletic clade with several unidentified strains from Malaysia. LSU and SSU rDNA sequences of three species of Ceratocorys (C. armata, C. gourreti, C. horrida) from the Mediterranean and several other unidentified strains from Malaysia form a well-supported sister clade. The unique phylogenetic position of an unidentified strain from Hawaii is also documented and requires further examination. In addition, based on the V9 SSU topology (bootstrap values >80%), specimens from Elands Bay (South Africa), originally described as Gonyaulax grindleyi by Reinecke (1967), cluster with Protoceratium reticulatum. The known range of Pentaplacodinium saltonense is tropical to subtropical, and its cyst is recorded as a fossil in upper Cenozoic sediments. Protoceratium reticulatum and Pentaplacodinium saltonense seem to inhabit different niches: motile stages of these dinoflagellates have not been found in the same plankton sample.