Danwei Huang’s research while affiliated with National University of Singapore and other places

Atlantic reef-building corals and coral reefs continue to experience extensive decline due to increased stressors related to climate change, disease, pollution, and numerous anthropogenic threats. To understand the impact of ocean warming and reef loss on the estimated extinction risk of shallow water Atlantic reef-building scleractinians and milleporids, all 85 valid species were reassessed under the IUCN Red List Categories and Criteria, updating the previous Red List assessment of Atlantic corals published in 2008. For the present assessment, individual species declines were estimated based on the modeled coral cover loss (1989–2019) and projected onset of annual severe bleaching events (2020–2050) across the Atlantic. Species traits were used to scale species’ relative vulnerability to the modeled cover declines and forecasted bleaching events. The updated assessments place 45.88%–54.12% of Atlantic shallow water corals at an elevated extinction risk compared to the previous assessments conducted in 2008 (15.19%–40.51%). However, coral cover loss estimates indicate an improvement in reef coverage compared to the historic time-series used for the 2008 assessments. Based on this, we infer that, although remaining dangerously high, the rate of Atlantic reef coral cover decline has surprisingly slowed in recent decades. However, based on modeled projections of sea-surface temperature that predict the onset of annual severe bleaching events within the next 30 years, we listed 26 (out of 85) species as Critically Endangered in the IUCN Red List. Each of these species had previously been listed under a lower threatened category and this result alone highlights the severe threat future bleaching events pose to coral survival and the reef ecosystems they support.

Aim As climate change increasingly threatens the world's coral reefs, enhancing their resilience by improving population connectivity for key reef species is crucial for ensuring their persistence. Here, we evaluate the population genomic structure of two common coral species, Pocillopora acuta and Porites sp., chosen due to their divergent life histories. Thousands of single‐nucleotide polymorphisms (SNPs) were sequenced and analysed to infer regional connectivity patterns in Southeast Asia, a region that harbours a tremendous diversity of marine life. Location Coasts of the Malay Peninsula and northern Borneo, covering ~1 million km². Method NextRAD genotyping‐by‐sequencing of 185 Porites sp. and 221 Pocillopora acuta colonies. Libraries were prepared and sequenced on Illumina NovaSeq 6000. Genotyping involved initial quality controls, allele frequency filtering and checks for contamination. Genetic structure was assessed with Bayesian clustering, and relationships between genetic variation and environmental factors were studied through redundancy analysis. Contemporary gene flow was estimated using BayesAss. Results We observed panmixia among the broadcasting Porites sp. populations, while for the brooding Pocillopora acuta, the Malay Peninsula acts a strong barrier to dispersal between the Malacca Strait and the southern South China Sea. Moreover, its genomic structure seems to follow current marine ecoregion delineation. By analysing contemporary migrant movement, we can prioritise reef localities for conservation. In particular, localities at the Andaman Coral Coast are contemporarily isolated from the other localities, and Tioman is identified as a major larval source for both species. Main Conclusion Our analyses highlight contrasting population differentiation patterns between the two species that can be explained by the disparity in their reproductive strategies. These findings are important for biodiversity managers in Southeast Asia; incorporation of regional connectivity considerations into conservation planning can help safeguard ecosystem resilience and persistence.

Background DNA metabarcoding applies high-throughput sequencing approaches to generate numerous DNA barcodes from mixed sample pools for mass species identification and community characterisation. To date, however, most metabarcoding studies employ second-generation sequencing platforms like Illumina, which are limited by short read lengths and longer turnaround times. While third-generation platforms such as the MinION (Oxford Nanopore Technologies) can sequence longer reads and even in real-time, application of these platforms for metabarcoding has remained limited possibly due to the relatively high read error rates as well as the paucity of specialised software for processing such reads. Results We show that this is no longer the case by performing nanopore-based, cytochrome c oxidase subunit I (COI) metabarcoding on 34 zooplankton bulk samples, and benchmarking the results against conventional Illumina MiSeq sequencing. Nanopore R10.3 sequencing chemistry and super accurate (SUP) basecalling model reduced raw read error rates to ~ 4%, and consensus calling with amplicon_sorter (without further error correction) generated metabarcodes that were ≤ 1% erroneous. Although Illumina recovered a higher number of molecular operational taxonomic units (MOTUs) than nanopore sequencing (589 vs. 471), we found no significant differences in the zooplankton communities inferred between the sequencing platforms. Importantly, 406 of 444 (91.4%) shared MOTUs between Illumina and nanopore were also found to be free of indel errors, and 85% of the zooplankton richness could be recovered after just 12–15 h of sequencing. Conclusion Our results demonstrate that nanopore sequencing can generate metabarcodes with Illumina-like accuracy, and we are the first study to show that nanopore metabarcodes are almost always indel-free. We also show that nanopore metabarcoding is viable for characterising species-rich communities rapidly, and that the same ecological conclusions can be obtained regardless of the sequencing platform used. Collectively, our study inspires confidence in nanopore sequencing and paves the way for greater utilisation of nanopore technology in various metabarcoding applications.

Identification of fish larvae based on morphology is typically limited to higher taxonomic ranks (e.g., family or order), as larvae possess few morphological diagnostic characters for precise discrimination to species. When many samples are presented at any one time, the use of morphology to identify such specimens can be laborious and time‐consuming. Using a reverse workflow for specimen sorting and identification leveraging high‐throughput DNA sequencing, thousands of fish larvae can be DNA barcoded and sorted into molecular operational taxonomic units (mOTUs) in a single sequencing run with the nanopore sequencing technology (e.g., MinION). This process reduces the time and financial costs of morphology‐based sorting and instead deploys experienced taxonomists for species taxonomic work where they are needed most. In this study, a total of 3022 fish larval specimens from plankton tows across four sites in Singapore were collected and sorted based on this workflow. Eye tissue from individual samples was used for DNA extraction and sequencing of cytochrome c oxidase subunit I. We generated a total of 2746 barcodes after quality filtering (90.9% barcoding success), identified 2067 DNA barcodes (75.3% identification success), and delimited 256 mOTUs (146 genera, 52 families). Our analyses identified specific challenges to species assignment, such as the potential misidentification of publicly available sequences used as reference barcodes. We highlighted how the conservative application and comparison of a local sequence database can help resolve identification conflicts. Overall, this proposed approach enables and expedites taxonomic identification of fish larvae, contributing to the enhancement of reference barcode databases and potentially better understanding of fish connectivity.

Gerromorpha Popov, 1971 is a fascinating and diverse insect lineage that evolved about 200 Mya to spend their entire life cycle on the air-water interface and have since colonized all types of aquatic habitats. The sub-family Halobatinae Bianchi, 1896 is particularly interesting because some species have adapted to life on the open ocean-a habitat where insects are very rarely found. Several attempts have been made to reconstruct the phylogenetic hypotheses of this subfamily, but the use of a few partial gene sequences recovered only a handful of well-supported relationships, thus limiting evolutionary inferences. Fortunately, the emergence of high-throughput sequencing technologies has enabled the recovery of more genetic markers for phylogen-etic inference. We applied genome skimming to obtain mitochondrial and nuclear genes from low-coverage whole-genome sequencing of 85 specimens for reconstructing a well-supported phylogeny, with particular emphasis on Halobatinae. Our study confirmed that Metrocorini Matsuda, 1960, is paraphyletic, whereas Esakia Lundblad, 1933, and Ventidius Distant, 1910, are more closely related to Halobatini Bianchi, 1896, than Metrocoris Mayr, 1865, and Eurymetra Esaki, 1926. We also found that Ventidius is paraphyletic and in need of a taxonomic revision. Ancestral state reconstruction suggests that Halobatinae evolved progressively from limnic to coastal habitats, eventually attaining a marine lifestyle, especially in the genus Halobates Eschscholtz, 1822, where the oceanic lifestyle evolved thrice. Our results demonstrate that genome skimming is a powerful and straightforward approach to recover genetic loci for robust phylogenetic analysis in non-model insects.

Species identification of stony corals (Scleractinia), which are regulated under the Convention on International Trade in Endangered Species of Wild Fauna and Flora, is critical for effective control of harvest quotas, enforcement of trade regulations and species conservation in general. DNA barcoding has the potential to enhance species identification success, depending on the specific taxon concerned and genetic markers used. For Acropora, DNA barcoding, based on the mitochondrial putative control region (mtCR) and the nuclear PaxC intron (PaxC), has been commonly used for species identification and delimitation, but the reliability and robustness of these loci remain contentious. Therefore, we sought to verify the applicability of this approach. In this study, we obtained 127 Acropora colonies from the aquarium trade to test the effectiveness of barcoding mtCR and PaxC for species identification. We were able to recover sequences for both loci in over half of the samples (n = 68), while gene amplification and sequencing of mtCR (n = 125) outperformed PaxC (n = 70). Amongst the 68 samples with both loci recovered, just a single sample could be unambiguously identified to species. Preliminary identities, based on only one gene, were assigned for 40 and 65 samples with mtCR and PaxC, respectively. Further analyses of 110 complete mitochondrial genomes obtained from GenBank showed that, despite the full length of the sequences, only eight species were delimited, of which only three species were correspondingly monophyletic. Therefore, we conclude that the commonly used DNA barcoding markers for Acropora are ineffective for accurate species assignments due to limited variability in both markers and even across the entire mitochondrial genome. Therefore, we propose that barcoding markers should generally not be the only means for identifying corals.

Coral reefs worldwide confront escalating threats, characterized by heightened sedimentation and diminished light penetration due to extensive land reclamation and coastal development. This critical issue is exemplified by the drastic reduction in underwater visibility, notably in Singapore, where levels have dwindled from over 10 meters in the 1960s to approximately 2 meters today. This study endeavours to explore the repercussions of acute low-light stress on the photo-physiology and transcriptomics of the common tropical coral Pachyseris speciosa. The research implemented a 4-week experiment involving three distinct light treatments: control, representing normal light conditions; intermittent light, with 5 days of normal light conditions and 2 days of total darkness; dark, maintaining constant total darkness. Comprehensive photo-physiological data, encompassing parameters like zooxanthellae density, chlorophyll a concentration, colour score, and chlorophyll fluorescence (EQY and MQY), were recorded at the experiment's onset and conclusion using a DPAM-II. Additionally, coral samples collected at the beginning and conclusion of the experiment underwent RNA sequencing via Illumina for transcriptomic analysis, enhancing the depth of understanding regarding molecular responses. Post-experiment analysis revealed the resilience of all P. speciosa colonies, albeit with notable disparities in photo-physiological status. The dark treatment induced substantial reductions in chlorophyll a concentration, zooxanthellae density, and MQY. Interestingly, colonies subjected to intermittent light treatment exhibited signs of photo-acclimation, evident in a significant increase in EQY. Transcriptomic analysis unveiled unique gene expression patterns, particularly in intermittent light-treated colonies, showcasing differential expression of genes associated with low-light adaptations. Pathway analysis emphasized enrichment in photosynthesis-related genes, indicating enhanced light capture and utilization in corals exposed to intermittent light. Overall, our findings suggest that P. speciosa may possess the photo-physiological and molecular capacity to adapt to low-light stress, positioning it as a promising candidate for use in restoration projects in low light environments affected by reduced light penetration.