Contrasting patterns of genetic structure and phylogeography in the marine agarophytes Gelidiophycus divaricatus and G. freshwateri (Gelidiales, Rhodophyta) from East Asia.

The evolutionary and population demographic history of marine red algae in East Asia is poorly understood. Here, we reconstructed the phylogeographies of two upper intertidal species endemic to East Asia, Gelidiophycus divaricatus and G. freshwateri. Phylogenetic and phylogeographic inferences of 393 mitochondrial cox1, 128 plastid rbcL, and 342 nuclear ITS2 sequences were complemented with ecological niche models. Gelidiophycus divaricatus, a southern species adapted to warm water, is characterized by a high genetic diversity and a strong geographical population structure, characteristic of stable population sizes and sudden reduction to recent expansion. In contrast, G. freshwateri, a northern species adapted to cold temperate conditions, is genetically relatively homogeneous with a shallow population structure resulting from steady population growth and recent equilibrium. The overlap zone of the two species roughly matches summer and winter isotherms, indicating that surface seawater temperature is a key feature influencing species range. Unidirectional genetic introgression was detected at two sites on Jeju Island where G. divaricatus was rare while G. freshwateri was common, suggesting the occurrence of asymmetric natural hybrids, a rarely reported event for rhodophytes. Our results illustrate that Quaternary climate oscillations have left strong imprints on the current day genetic structure and highlight the importance of seawater temperature and sea level change in driving speciation in upper intertidal seaweed species.

Genome of the sea anemone Exaiptasia pallida and transcriptome profiles during tentacle regeneration.

Cnidarians including sea anemones, corals, hydra, and jellyfishes are a group of animals well known for their regeneration capacity. However, how non-coding RNAs such as microRNAs (also known as miRNAs) contribute to cnidarian tissue regeneration is poorly understood. Here, we sequenced and assembled the genome of the sea anemone Exaiptasia pallida collected in Hong Kong waters. The assembled genome size of E. pallida is 229.21 Mb with a scaffold N50 of 10.58 Mb and BUSCO completeness of 91.1%, representing a significantly improved genome assembly of this species. The organization of ANTP-class homeobox genes in this anthozoan further supported the previous findings in jellyfishes, where most of these genes are mainly located on three scaffolds. Tentacles of E. pallida were excised, and both mRNA and miRNA were sequenced at 9 time points (0 h, 6 h, 12 h, 18 h, 1 day, 2, 3, 6, and 8 days) from regenerating tentacles. In addition to the Wnt signaling pathway and homeobox genes that are shown to be likely involved in tissue regeneration as in other cnidarians, we have shown that GLWamide neuropeptides, and for the first time sesquiterpenoid pathway genes could potentially be involved in the late phase of cnidarian tissue regeneration. The established sea anemone model will be useful for further investigation of biology and evolution in, and the effect of climate change on this important group of animals.

Phylogeography of the marine macroalga Sargassum hemiphyllum (Phaeophyceae, Heterokontophyta) in northwestern Pacific.

Sargassum hemiphyllum is commonly found in Japan and Korea, with a variety, var. chinense, that is found distributed in the southern Chinese coast. We previously reported distinct genetic differentiation between the two taxa based on the PCR-RFLP data of plastid RubiscoL-S spacer. The present study aims at elucidating the phylogeographic pattern of S. hemiphyllum based on more markers in the nuclear and extranuclear genomes, with a view to reveal the occurrence of hybridization. The two allopatrically distributed taxa were found to be genetically distinct in nuclear ITS2, plastidial Rubisco (Rbc) and mitochondrial TrnW_I (Trn) spacers. Their divergence was postulated to be attributable to the vicariant event which resulted from the isolation of the Sea of Japan during the late Miocene (6.58-11.25 Mya). Divergence within both S. hemiphyllum and the chinense variety was observed based on Trn spacer, while the divergence in S. hemiphyllum was further confirmed in Rbc spacer. This divergence appears to correspond to the separation of the Japanese populations between the Sea of Japan and the Pacific that occurred around 0.92-2.88 Mya (the early Pleistocene). The presence of an ITS2 clone resembling var. chinense sequences in a Japanese population of S. hemiphyllum (JpNS) raises the possibility of the introgression of var. chinense individuals into S. hemiphyllum population. Compared to that between S. hemiphyllum and the chinense variety, hybridization among the Japanese and Korean populations of S. hemiphyllum is highly probable as all these individuals share a pool of nuclear ITS2 sequences, possibly attributable to incomplete concerted evolution of ITS2.

Fish and crustacean biodiversity in an outer maritime estuary of the Pearl River Delta revealed by environmental DNA.

Understanding the faunal community structure in the estuary would be crucial in assessing the health of the ecosystem. The poor visibility in the estuarine area due to the outflow from the Pearl River hinders the conventional visual census in assessing the megafaunal biodiversity. In this study, the fish and crustacean biodiversity of Hong Kong's western waters, i.e. the outer maritime estuary of the PRD, were studied through the metabarcoding of environmental DNA (eDNA). eDNA from the seawater and sediment samples was extracted from five sites in the region. After testing the performance of two genetic markers, amplicons of the cytochrome oxidase I, amplified by polymerase chain reaction, were subjected to Illumina high-throughput sequencing (MiSeq) analysis. A total of 22 fish species from 17 families and 34 crustacean species from 27 families were identified by blasting the sequences against the NCBI GenBank database, demonstrating segregation between samples from different sites. This study provides insight on the detail distribution of fish assembly in PRD, when compared with a previous eDNA study in the inner brackish PRD.

Catastrophic regime shifts in coral communities exposed to physical disturbances: simulation results from object-oriented 3-dimensional coral reef model.

A 3-dimensional individual-based model, the ReefModel, was developed to simulate the dynamical structure of coral reef community using object-oriented techniques. Interactions among functional groups of reef organisms were simulated in the model. The behaviours of these organisms were described with simple mechanistic rules that were derived from their general behaviours (e.g. growing habits, competitive mechanisms, response to physical disturbance) observed in natural coral reef communities. The model was implemented to explore the effects of physical disturbance on the dynamical structure of a 3-coral community that was characterized with three functional coral groups: tabular coral, foliaceous coral and massive coral. Simulation results suggest that (i) the integration of physical disturbance and differential responses (disturbance sensitivity and growing habit) of corals plays an important role in structuring coral communities; (ii) diversity of coral communities can be maximal under intermediate level of acute physical disturbance; (iii) multimodality exists in the final states and dynamic regimes of individual coral group as well as coral community structure, which results from the influence of small random spatial events occurring during the interactions among the corals in the community, under acute and repeated physical disturbances. These results suggest that alternative stable states and catastrophic regime shifts may exist in a coral community under unstable physical environment.

Antiviral activities of extracts from Hong Kong seaweeds.

We extracted six Hong Kong brown seaweed species with hot water for their antiviral properties. The cytotoxicity and antiviral activity of these extracts were tested by MTT [3-(4,5-dimethylthiazol-2-yl)-2,5-diphenlytetrezolium bromide] method, cytopathic effect reduction assay, and plaque reduction assay. The antiviral effect was further determined by flow cytometric analysis. The results showed that most of these extracts inhibited the propagation of herpes simplex virus types 1 and 2 (HSV-1 and HSV-2) standard strains with very low cytotoxicity to the host cells. The extracts of Hydroclathrus clathratus and Lobophora variegata showed more potential anti-HSV activities than the extracts of the other four seaweeds. They also had moderate anti- respiratory syncytial virus (RSV) activities but could not inhibit influenza A virus. Hydroclathrus clathratus was further extracted by diluted acid and alkali and the antiviral effects of the extracts were also detected. The result showed that the hot water extract contained the main carbohydrate components that exhibited the antiviral activities against various strains of HSV, including the acyclovir-resistant strain. HI-3, a compound fractionated from this hot water extract, showed a dose-dependent anti-HSV activity in flow cytometric analysis and plaque reduction assay.

Exploring coral microbiome assemblages in the South China Sea.

Coral reefs are significant ecosystems. The ecological success of coral reefs relies on not only coral-algal symbiosis but also coral-microbial partnership. However, microbiome assemblages in the South China Sea corals remain largely unexplored. Here, we compared the microbiome assemblages of reef-building corals Galaxea (G. fascicularis) and Montipora (M. venosa, M. peltiformis, M. monasteriata) collected from five different locations in the South China Sea using massively-parallel sequencing of 16S rRNA gene and multivariate analysis. The results indicated that microbiome assemblages for each coral species were unique regardless of location and were different from the corresponding seawater. Host type appeared to drive the coral microbiome assemblages rather than location and seawater. Network analysis was employed to explore coral microbiome co-occurrence patterns, which revealed 61 and 80 co-occurring microbial species assembling the Galaxea and Montipora microbiomes, respectively. Most of these co-occurring microbial species were commonly found in corals and were inferred to play potential roles in host nutrient metabolism; carbon, nitrogen, sulfur cycles; host detoxification; and climate change. These findings suggest that the co-occurring microbial species explored might be essential to maintain the critical coral-microbial partnership. The present study provides new insights into coral microbiome assemblages in the South China Sea.

Urban coral reefs: Degradation and resilience of hard coral assemblages in coastal cities of East and Southeast Asia.

Given predicted increases in urbanization in tropical and subtropical regions, understanding the processes shaping urban coral reefs may be essential for anticipating future conservation challenges. We used a case study approach to identify unifying patterns of urban coral reefs and clarify the effects of urbanization on hard coral assemblages. Data were compiled from 11 cities throughout East and Southeast Asia, with particular focus on Singapore, Jakarta, Hong Kong, and Naha (Okinawa). Our review highlights several key characteristics of urban coral reefs, including "reef compression" (a decline in bathymetric range with increasing turbidity and decreasing water clarity over time and relative to shore), dominance by domed coral growth forms and low reef complexity, variable city-specific inshore-offshore gradients, early declines in coral cover with recent fluctuating periods of acute impacts and rapid recovery, and colonization of urban infrastructure by hard corals. We present hypotheses for urban reef community dynamics and discuss potential of ecological engineering for corals in urban areas.

Antiviral polysaccharides isolated from Hong Kong brown seaweed Hydroclathrus clathratus.

Two relatively pure polysaccharides H3-a1 and H3-b1 had been isolated from the brown seaweed Hydroclathrus clathratus. They were characterized by HPLC, ultraviolet scanning, gas chromatography, infrared spectroscopy and elemental analysis, and shown to be two different sulfated polysaccharides with different monosaccharide content, but both with high relative molecular mass. They contained some proteins and uronic acid respectively. The sulfate content and bioactivity of these polysaccharides varied during purification. The fractions derived from the hot water extract also exhibited low anticoagulant effect. This is the first time that the antiherpetic and anticoagulant activities were evaluated for the polysaccharides from the Hong Kong brown seaweed Hydroclathrus clathratus.

Enhancement of bromophenol levels in aquacultured silver seabream (Sparus sarba).

The effect of the addition of marine algae in fish feed on the levels of bromophenols in fish flesh was studied. These bromophenols include 2-bromophenol, 4-bromophenol, 2,4-dibromophenol, 2,6-dibromophenol, and 2,4,6-tribromophenol. Two types of algae-containing fish feeds with 30% Padina arborescens and 30% Sargassum siliquastrum were developed. The total bromophenol contents of these feeds were 132 and 340 ng/g respectively, which were significantly higher than that of the control feed (8.9 ng/g) (ANOVA, p < 0.05). Silver seabream was used as the model fish for the feeding experiment. Bromophenol contents of both fish gut and flesh were monitored at 2-week intervals throughout the 8-week period. Two-way ANOVA showed that only the 30% S. siliquastrum-containing feed significantly (p < 0.05) increased the total bromophenol content in the fish flesh with time. This also produced sensorial differences in the fish flesh.

Seasonal variations of bromophenols in brown algae (Padina arborescens, Sargassum siliquastrum, and Lobophora variegata) collected in Hong Kong.

Distributions and seasonal variations of the key seafood flavor compounds including 2-bromophenol, 4-bromophenol, 2,4-dibromophenol, 2,6-dibromophenol, and 2,4,6-tribromophenol in three species of brown algae (Padina arborescens, Sargassum siliquastrum, and Lobophora variegata) found in Hong Kong waters were investigated. Bromophenols were extracted by simultaneous steam distillation and solvent extraction apparatus and analyzed by gas chromatography-mass spectrometry. On a dried weight basis, the total bromophenol content (TBC) determined varied widely with seasons (from 40.9 to 7030 ng/g). The TBCs detected were higher in winter and lower in summer. Except for 2-bromophenol, the rest of the bromophenols were detected in all of the algal samples. The TBC of L. variegata was generally the highest among all of the algae collected. Relatively high concentrations of bromophenols in algae supported the fact that marine algae were major producers of bromophenols in the marine environment.

MORPHOLOGICAL AND GENETIC VARIATION IN THE POPULATIONS OF SARGASSUM HEMIPHYLLUM (PHAEOPHYCEAE) IN THE NORTHWESTERN PACIFIC(1).

Difficulty in species identification of Sargassum (Sargassaceae, Fucales) is partly attributed to the high polymorphism among its individuals and populations. This study aimed at assessing morphological and genetic variations in two varieties, var. hemiphyllum J. Agardh and var. chinense J. Agardh, of Sargassum hemiphyllum (Turner) C. Agardh, a widely distributed species in the northwestern Pacific. We investigated 26 measurable, five numerical, and 33 categorical morphological parameters associated with different branching levels of specimens from each of six localities within its distribution range using cluster analysis (CA) and principal coordinate analysis (PCoA). Leaf size of the primary and secondary branching levels and the vesicle size of the secondary branches of the specimens examined were determined to be the most important morphological parameters that were significantly different among populations. Change in leaf and vesicle length of individuals among the six populations followed a latitudinal gradient, with smaller leaves and vesicles associated with northern populations and larger ones in the southern populations. The possible influence of the gradual change in sea surface temperatures (SSTs) along this gradient in the northwestern Pacific on leaf and vesicle morphologies of this species was suggested. PCR-RFLP analysis of the RUBISCO spacer in the chloroplast genome revealed two distinct and highly homogenous clades, a China clade and a Japan-Korea clade, which corresponded to var. chinense and var. hemiphyllum, respectively. The formation of refugia along the "Paleo-coast" in the East China Sea during glacial periods is suggested to have led to the vicariance of ancestral populations of S. hemiphyllum and thus to have promoted genetic differentiation. The massive freshwater outflow of the Yellow and Yangtze rivers may continue to act as a barrier, prolonging the allopatric distribution of the two varieties.

Isolation and characterization of a sulfated polysaccharide from the brown alga Sargassum patens and determination of its anti-herpes activity.

Bioactivity-guided fractionation of the hot water extract from the brown alga Sargassun patens led to the isolation of a polysaccharide as an antiviral component against herpes simplex viruses which are the cause of cold sores (HSV-1) and genital herpes (HSV-2). The polysaccharide contained a sulfur group that could be present as a sulfate ester. It is thus a sulfated polysaccharide with a molecular mass of about 424 kDa, and is designated SP-2a. Gas chromatographic assay showed that the polysaccharide consisted of fucose, galactose, mannose, xylose, glucose, and galactosamine. The fucose is the major constituent sugar (35.3%), followed by galactose (18.4%). The 50% effective concentration (EC50) against HSV-2, HSV-1, and HSV-1 acyclovir resistant strain was 1.3, 5.5, and 4.1 microg/mL, respectively. The 50% cytotoxic concentration (CC50) of SP-2a on the growth of normal Vero cell line was more than 4000 microg/mL. Therefore SP-2a of S. patens may be a potent agent for treating HSV infections.