Potential PCR amplification bias in identifying complex ecological patterns: Higher species compositional homogeneity revealed in smaller-size coral reef zooplankton by metatranscriptomics.

PCR-based high-throughput sequencing has permitted comprehensive resolution analyses of zooplankton diversity dynamics. However, significant methodological issues still surround analyses of complex bulk community samples, not least as in prevailing PCR-based approaches. Marine drifting animals-zooplankton-play essential ecological roles in the pelagic ecosystem, transferring energy and elements to higher trophic levels, such as fishes, cetaceans and others. In the present study, we collected 48 size-fractionated zooplankton samples in the vicinity of a coral reef island with environmental gradients. To investigate the spatiotemporal dynamics of zooplankton diversity patterns and the effect of PCR amplification biases across these complex communities, we first took metatranscriptomics approach. Comprehensive computational analyses revealed a clear pattern of higher/lower homogeneity in smaller/larger zooplankton compositions across samples respectively. Our study thus suggests changes in the role of dispersal across the sizes. Next, we applied in silico PCR to the metatranscriptomics datasets, in order to estimate the extent of PCR amplification bias. Irrespective of stringency criteria, we observed clear separations of size fraction sample clusters in both metatranscriptomics and in silico datasets. In contrast, the pattern-smaller-fractioned communities had higher compositional homogeneity than larger ones-was observed in the metatranscriptomics data but not in the in silico datasets. To investigate this discrepancy further, we analysed the mismatches of widely used mitochondrial CO1 primers and identified priming site mismatches likely driving PCR-based biases. Our results suggest the use of metatranscriptomics or, although less ideal, redesigning the CO1 primers is necessary to circumvent these issues.

Utilisation of Oxford Nanopore sequencing to generate six complete gastropod mitochondrial genomes as part of a biodiversity curriculum.

High-throughput sequencing has enabled genome skimming approaches to produce complete mitochondrial genomes (mitogenomes) for species identification and phylogenomics purposes. In particular, the portable sequencing device from Oxford Nanopore Technologies (ONT) has the potential to facilitate hands-on training from sampling to sequencing and interpretation of mitogenomes. In this study, we present the results from sampling and sequencing of six gastropod mitogenomes (Aplysia argus, Cellana orientalis, Cellana toreuma, Conus ebraeus, Conus miles and Tylothais aculeata) from a graduate level biodiversity course. The students were able to produce mitogenomes from sampling to annotation using existing protocols and programs. Approximately 4 Gb of sequence was produced from 16 Flongle and one MinION flow cells, averaging 235 Mb and N50 = 4.4 kb per flow cell. Five of the six 14.1-18 kb mitogenomes were circlised containing all 13 core protein coding genes. Additional Illumina sequencing revealed that the ONT assemblies spanned over highly AT rich sequences in the control region that were otherwise missing in Illumina-assembled mitogenomes, but still contained a base error of one every 70.8-346.7 bp under the fast mode basecalling with the majority occurring at homopolymer regions. Our findings suggest that the portable MinION device can be used to rapidly produce low-cost mitogenomes onsite and tailored to genomics-based training in biodiversity research.

A dataset of sea turtle occurrences around the Taiwan coast.

Background: We describe a dataset of sea turtle sightings around the coast of Taiwan and its islands (Hoh and Fong 2022). This data collection was initiated by TurtleSpot Taiwan, a citizen-science project that collects sea turtle sighting data. This dataset includes 3,515 sighting data dated from March 2010, except most of the data (n = 3,128; 89%) were collected between June 2017 to December 2021. Sightings were reported by citizen scientists to the Facebook Group of TurtleSpot Taiwan by providing occurrence information. We also requested photos and videos for species identification and to record any physical abnormality of the turtle, if observable. In addition to recording data often associated with an occurrence, TurtleSpot aims to identify each sea turtle up to the individual level using the Photo Identification (Photo ID) method. Hence, if photos of left facial scutes were available, the sighted individual can be identified and given a unique turtle ID. In total, 762 individuals were assigned a turtle ID, comprising 723 Greens (Cheloniamydas), 38 Hawksbills (Eretmochelysimbricata) and one Olive Ridley (Lepidochelysolivacea) turtle. This dataset is now publicly opened in Global Biodiversity Information Facility (GBIF) and available for download. It is hoped that the data may assist in future ecological studies and the development of conservation measures. New information: This dataset contains 3,515 occurrence records of sea turtles (Cheloniidae) and is currently the largest public dataset of sea turtle sighting records in Taiwan. Post-publication of this dataset to the GBIF platform demonstrated that the number of Green sea turtle Cheloniamydas records in Taiwan is one of the largest in the world (last accessed date: 15-10-2022). The data served as the foundation for understanding biogeography and sea turtle ecology in Taiwan's coastal waters.

Grazing effects of sea urchin Diadema savignyi on algal abundance and coral recruitment processes.

Herbivores control algae and promote coral dominance along coral reefs. However, the majority of previous studies have focused on herbivorous fish. Here we investigated grazing effects of the sea urchin Diadema savignyi on algal abundance and coral recruitment processes. We conducted an in situ cage experiment with three density conditions of D. savignyi (0, 8, 16 indiv. m-2) for three months during the main coral recruitment season in Taiwan. Results demonstrated a strong algal control by D. savignyi. At the end of the experiment, average algal cover was 95% for 0 indiv. m-2, compared to 47% for 8 indiv. m-2 and 16% for 16 indiv. m-2. Average algal biomass at 8 indiv. m-2 declined by one third compared to 0 indiv. m-2 and almost zero at 16 indiv. m-2. On the other hand, a negative grazing effect of D. savignyi was observed on coral recruitment processes. Notably, at 16 indiv. m-2, the density of coral recruits declined and mortality of small coral fragments (proxy of coral juveniles) increased. Our results confirm findings of previous studies and indicate the need to balance both positive (strong algal control) and negative (physical damage) influences of Diadema grazing to facilitate the coral recruitment process.

Shifting in the Dominant Bacterial Group Endozoicomonas Is Independent of the Dissociation With Coral Symbiont Algae.

The coral-associated Endozoicomonas are dominant bacteria in the coral holobiont. Their relative abundance usually decreases with heat-induced coral bleaching and is proposed to be positively correlated with Symbiodiniaceae abundance. It remains unclear whether this phenomenon of decreased Endozoicomonas abundance is caused by temperature stress or a decreased abundance of Symbiodiniaceae. This study induced bleaching in the coral Euphyllia glabrescens using a dark treatment over 15 weeks. We examined shifts in Endozoicomonas abundance and experimentally reduced Symbiodiniaceae density. 16S rRNA gene amplicon sequencing was used to characterize the changes in bacterial community (incl. Endozoicomonas) over time, and the 16S rRNA gene copy number of Endozoicomonas was quantified by qPCR. We detected a high abundance of Endozoicomonas in E. glabrescens that underwent dark-induced bleaching. The results reveal that changes in the relative abundance of Endozoicomonas are unrelated to Symbiodiniaceae abundance, indicating that Endozoicomonas can be independent of Symbiodiniaceae in the coral holobiont.