Spawning and larval development of two tropical cowries (Gastropoda: Cypraeidae), Cypraea tigris and Mauritia arabica under laboratory conditions.

The spawning and larval culture of cowrie (family Cypraeidae) are both difficult and little known, in part due to the long planktonic period of most species. In this study, we describe the captive spawning behaviour and larval development of two tropical cowrie species, Cypraea tigris and Mauritia arabica. Both species brooded over their egg masses before hatching occurred and larvae were collected for culture under laboratory conditions. The brooding period for C. tigris was between 7 and 17 days, and freshly hatched veligers were approximately 200-240 µm in size. Cypraea tigris larvae were reared for up to 37 days in culture but did not achieve successful settlement. The brooding period for M. arabica was between 7 and 10 days, and hatched veligers were approximately 160-205 µm in size. The first settled juvenile M. arabica was observed at 70 days post-hatch. Our findings from this study represent the first comprehensive documentation of successful metamorphosis of Cypraeidae larvae, particularly M. arabica, into early-stage juvenile.

Sediment-driven plastisphere community assembly on plastic debris in tropical coastal and marine environments.

Coastal habitats have been suggested to serve as a sink for unaccounted plastic debris, i.e., "missing plastic" in the sea, and hence, a hotspot of plastic pollution in the marine and coastal environments. Although the accumulation of plastic debris may pose significant threats to coastal ecosystems, we know little about the fate of these plastic debris and their ecological impacts due to the lack of studies on plastic-microbe interactions in coastal habitats, especially for the tropical marine and coastal environments. In this study, we collected plastic debris from 14 sites consisting of various coastal ecosystems (seagrass meadows, mangrove forests, and beaches), and marine ecosystem (coral reef) around Singapore and characterized the prokaryotic and eukaryotic microbial communities colonized on them. Our results showed that the composition of plastisphere communities in these intertidal ecosystems was predominantly influenced by the sediment than by the plastic materials. Compared with surrounding sediment and seawater, the plastic debris enriched potential plastic degraders, such as Muricauda, Halomonas, and Brevundimonas. The plastic debris was also found to host taxa that play significant roles in biogeochemical cycles (e.g., cyanobacteria, Erythrobacter), hygienically relevant bacteria (e.g., Chryseobacterium, Brevundimonas), and potential pathogens that may negatively impact the health of coastal ecosystems (e.g., Thraustochytriaceae, Labyrinthulaceae, Flavobacterium). Taken together, our study provides valuable insights into the plastic-microbe interactions in tropical coastal and marine ecosystems, highlighting the urgent need for plastisphere studies to understand the fate and ecological impacts of plastic debris accumulated in coastal habitats.

Litter traps: A comparison of four marine habitats as sinks for anthropogenic marine macro-litter in Singapore.

The potential for marine litter being trapped in biodiverse marine habitats such as mangrove forests, seagrass meadows and coral reefs is poorly understood. This study presents the first comprehensive investigation on the status of macro-litter across four marine habitats in Singapore during the two monsoonal seasons. Overall, litter density did not vary considerably between the southwest and the northeast monsoon. The litter density in terms of count was generally lower in seagrass meadows and coral reefs compared to mangroves and beaches. Plastic was the major type of litter found across most habitat types. Notably, many fishing-related items were found on coral reefs, while drinking straws were abundant at the mangrove strandlines during the southwest monsoon. Foam fragments and cigarette butts were common at the beach strandlines. These results suggest that mangroves among other habitats examined here should be prioritised for clean-up efforts in order to restore these critical coastal habitats.

Interactions between marine megafauna and plastic pollution in Southeast Asia.

Southeast (SE) Asia is a highly biodiverse region, yet it is also estimated to cumulatively contribute a third of the total global marine plastic pollution. This threat is known to have adverse impacts on marine megafauna, however, understanding of its impacts has recently been highlighted as a priority for research in the region. To address this knowledge gap, a structured literature review was conducted for species of cartilaginous fishes, marine mammals, marine reptiles, and seabirds present in SE Asia, collating cases on a global scale to allow for comparison, coupled with a regional expert elicitation to gather additional published and grey literature cases which would have been omitted during the structured literature review. Of the 380 marine megafauna species present in SE Asia, but also studied elsewhere, we found that 9.1 % and 4.5 % of all publications documenting plastic entanglement (n = 55) and ingestion (n = 291) were conducted in SE Asian countries. At the species level, published cases of entanglement from SE Asian countries were available for 10 % or less of species within each taxonomic group. Additionally, published ingestion cases were available primarily for marine mammals and were lacking entirely for seabirds in the region. The regional expert elicitation led to entanglement and ingestion cases from SE Asian countries being documented in 10 and 15 additional species respectively, highlighting the utility of a broader approach to data synthesis. While the scale of the plastic pollution in SE Asia is of particular concern for marine ecosystems, knowledge of its interactions and impacts on marine megafauna lags behind other areas of the world, even after the inclusion of a regional expert elicitation. Additional funding to help collate baseline data are critically needed to inform policy and solutions towards limiting the interactions of marine megafauna and plastic pollution in SE Asia.

Genomic Survey and Resources for the Boring Giant Clam Tridacna crocea.

The boring giant clam Tridacna crocea is an evolutionary, ecologically, economically, and culturally important reef-dwelling bivalve targeted by a profitable ornamental fishery in the Indo-Pacific Ocean. In this study, we developed genomic resources for T. crocea. Using low-pass (=low-coverage, ~6×) short read sequencing, this study, for the first time, estimated the genome size, unique genome content, and nuclear repetitive elements, including the 45S rRNA DNA operon, in T. crocea. Furthermore, we tested if the mitochondrial genome can be assembled from RNA sequencing data. The haploid genome size estimated using a k-mer strategy was 1.31-1.39 Gbp, which is well within the range reported before for other members of the family Cardiidae. Unique genome content estimates using different k-mers indicated that nearly a third and probably at least 50% of the genome of T. crocea was composed of repetitive elements. A large portion of repetitive sequences could not be assigned to known repeat element families. Taking into consideration only annotated repetitive elements, the most common were classified as Satellite DNA which were more common than Class I-LINE and Class I-LTR Ty3-gypsy retrotransposon elements. The nuclear ribosomal operon in T. crocea was partially assembled into two contigs, one encoding the complete ssrDNA and 5.8S rDNA unit and a second comprising a partial lsrDNA. A nearly complete mitochondrial genome (92%) was assembled from RNA-seq. These newly developed genomic resources are highly relevant for improving our understanding of the biology of T. crocea and for the development of conservation plans and the fisheries management of this iconic reef-dwelling invertebrate.

A comparative investigation of toxicity of three polymer nanoparticles on acorn barnacle (Amphibalanus amphitrite).

Pollution from plastic waste is increasingly prevalent in the environment and beginning to generate significant adverse impact on the health of living organisms. In this study, we investigate the toxicity of polymer nanoparticles exposed to Acorn Barnacle (Amphibalanus amphitrite) nauplii, as an animal model. Highly stable aqueous dispersion of luminescent nanoparticles from three common polymers: polymethylmethacrylate (PMMA), polystyrene (PS), and polyvinylchloride (PVC), were prepared via nanoprecipitation and fully characterised. Exposure studies of these polymer particles to freshly spawned barnacle nauplii were performed within a concentration range from 1 to 25 mg/L under laboratory-controlled conditions. The exposure to PMMA and PS nanoparticles did not show detrimental toxicity and did not cause sufficient mortality to compute a LC50 value. However, PVC nanoparticles were significantly toxic with a mortality rate of up to 99% at 25 mg/L, and the calculated LC50 value for PVC nanoparticles was 7.66 ± 0.03 mg/L, 95% CI. Interestingly, PVC nanoparticle aggregates were observed to adhere to the naupliar carapace and appendages at higher concentrations and could not be easily removed by washings. To explore the possibility of chemical toxicity of polymer nanoparticles, analysis of the polymer powders which was used to prepare the nanoparticles was conducted. The presence of low molecular weight oligomers such as dimers, trimers and tetramers were observed in all polymer samples. The chemical nature and concentration of such compounds are likely responsible for the observed toxicity to the barnacle nauplii. Overall, our study shows that care should be exercised in generalising the findings of exposure studies performed using one type of plastic particles, as the use of different plastic particles may elicit different responses inside a living organism.

Understanding the biological impact of organic pollutants absorbed by nanoplastics.

Many organisms are consuming food contaminated with micro- and nanoparticles of plastics, some of which absorb persistent organic pollutants (POPs) from the environment and acting as carrier vectors for increasing the bioavailability in living organisms. We recently reported that polymethylmethacrylate (PMMA) nanoparticles at low concentrations are not toxic to animal models tested. In this study, the toxicity of diphenylamine (DPA) incorporated PMMA nanoparticles are assessed using barnacle larvae as a model organism. The absorption capacity of DPA from water for commercially available virgin PMMA microparticles is relatively low (0.14 wt%) during a 48 h period, which did not induce exposure toxicity to barnacle nauplii. Thus, PMMA nanoparticles encapsulated with high concentrations of DPA (DPA-enc-PMMA) were prepared through a reported precipitation method to achieve 40% loading of DPA inside the particles. Toxicity of DPA-enc-PMMA nanoparticles were tested using freshly spawned acorn barnacle nauplii. The observed mortality of nauplii from DPA-enc-PMMA exposure was compared to the values obtained from pure DPA exposure in water. The mortality among the exposed acorn barnacle nauplii did not exceed 50% even at a high concentration of DPA inside the PMMA nanoparticles. The results suggest that the slow release of pollutants from polymer nanoparticles may not induce significant toxicity to the organism living in a dynamic environment. The impact of long-term exposure of DPA absorbed plastic nanoparticles need to be investigated in the future.

Conserving threatened species during rapid environmental change: using biological responses to inform management strategies of giant clams.

Giant clams are threatened by overexploitation for human consumption, their valuable shells and the aquarium trade. Consequently, these iconic coral reef megafauna are extinct in some former areas of their range and are included in the International Union for Conservation of Nature (IUCN) Red List of Threatened Species and Convention on International Trade in Endangered Species of Wild Fauna and Flora. Now, giant clams are also threatened by rapid environmental change from both a suite of local and regional scale stressors and global change, including climate change, global warming, marine heatwaves and ocean acidification. The interplay between local- to regional-scale and global-scale drivers is likely to cause an array of lethal and sub-lethal effects on giant clams, potentially limiting their depth distribution on coral reefs and decreasing suitable habitat area within natural ranges of species. Global change stressors, pervasive both in unprotected and protected areas, threaten to diminish conservation efforts to date. International efforts urgently need to reduce carbon dioxide emissions to avoid lethal and sub-lethal effects of global change on giant clams. Meanwhile, knowledge of giant clam physiological and ecological responses to local-regional and global stressors could play a critical role in conservation strategies of these threatened species through rapid environmental change. Further work on how biological responses translate into habitat requirements as global change progresses, selective breeding for resilience, the capacity for rapid adaptive responses of the giant clam holobiont and valuing tourism potential, including recognizing giant clams as a flagship species for coral reefs, may help improve the prospects of these charismatic megafauna over the coming decades.

Transcriptome-based target-enrichment baits for stony corals (Cnidaria: Anthozoa: Scleractinia).

Despite the ecological and economic significance of stony corals (Scleractinia), a robust understanding of their phylogeny remains elusive due to patchy taxonomic and genetic sampling, as well as the limited availability of informative markers. To increase the number of genetic loci available for phylogenomic analyses in Scleractinia, we designed 15,919 DNA enrichment baits targeting 605 orthogroups (mean 565 ± SD 366 bp) over 1,139 exon regions. A further 236 and 62 barcoding baits were designed for COI and histone H3 genes respectively for quality and contamination checks. Hybrid capture using these baits was performed on 18 coral species spanning the presently understood scleractinian phylogeny, with two corallimorpharians as outgroup. On average, 74% of all loci targeted were successfully captured for each species. Barcoding baits were matched unambiguously to their respective samples and revealed low levels of cross-contamination in accordance with expectation. We put the data through a series of stringent filtering steps to ensure only scleractinian and phylogenetically informative loci were retained, and the final probe set comprised 13,479 baits, targeting 452 loci (mean 531 ± SD 307 bp) across 865 exon regions. Maximum likelihood, Bayesian and species tree analyses recovered maximally supported, topologically congruent trees consistent with previous phylogenomic reconstructions. The phylogenomic method presented here allows for consistent capture of orthologous loci among divergent coral taxa, facilitating the pooling of data from different studies and increasing the phylogenetic sampling of scleractinians in the future.

Quantification of coral sperm collected during a synchronous spawning event.

Most studies of coral reproductive biology to date have focused on oocyte numbers and sizes. Only one (ex situ) study has enumerated sperm numbers, even though these data have multiple potential applications. We quantified total coral sperm and eggs per gamete bundle collected from six species in situ during a synchronous spawning event in Singapore. Egg-sperm bundles were captured midwater as they floated towards the surface after being released by the colony. For each sample, a semi-transparent soft plastic bottle was squeezed and released to create a small suction force that was used to 'catch' the bundles. This technique provided several advantages over traditional methods, including low cost, ease of use, no diving prior to the night of collection needed, and the ability to target specific areas of the colony. The six species sampled were Echinophyllia aspera, Favites abdita, F. chinensis, Merulina ampliata, M. scabricula and Platygyra pini. The mean number of sperm packaged within one egg-sperm bundle ranged from 2.04 × 10(6) to 1.93 × 10(7). The mean number of eggs per egg-sperm bundle ranged from 26.67 (SE ± 3.27) to 85.33 (SE ± 17.79). These data are critical for fertilisation success models, but the collection technique described could also be applied to studies requiring in situ spawning data at the polyp level.

Recruitment constraints in Singapore's fluted giant clam (Tridacna squamosa) population--a dispersal model approach.

Recruitment constraints on Singapore's dwindling fluted giant clam, Tridacna squamosa, population were studied by modelling fertilisation, larval transport, and settlement using real-time hydrodynamic forcing combined with knowledge of spawning characteristics, larval development, behaviour, and settlement cues. Larval transport was simulated using a finite-volume advection-diffusion model coupled to a three-dimensional hydrodynamic model. Three recruitment constraint hypotheses were tested: 1) there is limited connectivity between Singapore's reefs and other reefs in the region, 2) there is limited exchange within Singapore's Southern Islands, and 3) there exist low-density constraints to fertilisation efficacy (component Allee effects). Results showed that connectivity among giant clam populations was primarily determined by residual hydrodynamic flows and spawning time, with greatest chances of successful settlement occurring when spawning and subsequent larval dispersal coincided with the period of lowest residual flow. Simulations suggested poor larval transport from reefs located along the Peninsular Malaysia to Singapore, probably due to strong surface currents between the Andaman Sea and South China Sea combined with a major land barrier disrupting larval movement among reefs. The model, however, predicted offshore coral reefs to the southeast of Singapore (Bintan and Batam) may represent a significant source of larvae. Larval exchange within Singapore's Southern Islands varied substantially depending on the locations of source and sink reefs as well as spawning time; but all simulations resulted in low settler densities (2.1-68.6 settled individuals per 10,000 m(2)). Poor fertilisation rates predicted by the model indicate that the low density and scattered distribution of the remaining T. squamosa in Singapore are likely to significantly inhibit any natural recovery of local stocks.