Comparing metabarcoding and metagenomics to characterize deep-sea benthic biodiversity from sediment-trap environmental DNA on New Caledonian seamounts

INTRODUCTION, SCIENTIFIC CONTEXT:
Seamounts in New Caledonia have been studied for over twenty years by the Tropical Deep Sea Benthos (TDSB) program. The pioneering study of Richer de Forges et al. (2000) showed that neighbouring seamounts, comparable in topology and depth, can host distinct benthic communities. Subsequent work (Samadi et al. 2006; Castelin et al. 2010, 2011, 2012; Pante et al. 2015; O’Hara et al. 2007, 2021, 2025) has kept documenting this heterogeneity without fully explaining its causes. Basic population genetics tools have described demographic structure but cannot, on their own, identify the underlying drivers. One hypothesis relates benthic demography on each seamount to trophic and demographic coupling between the pelagic and benthic compartments, i.e. to the joint influence of the flux of organic matter and the flux of benthic larvae. Direct evidence for this coupling is still lacking.
During the KANADEEP2 cruise, particle traps (rotating carousels of 24 collection cups) were deployed on two nearby but faunistically contrasted seamounts to record organic-matter and larval fluxes through time. Cups are usually fixed in formalin, which allows organic matter to be quantified and swimming, bentho-pelagic and benthic larval organisms to be observed (Arsenault-Pernet et al. 2026), but the resulting material generally cannot be assigned to described species. In one cup out of two, formalin was replaced by RNAlater to allow environmental DNA (eDNA) analysis of the trap content. During a first internship (Intechmer, 2026), DNA was extracted from 8 RNAlater cups (particle and supernatant fractions): the extracts are concentrated, of high molecular weight, and standard barcode markers (COI Folmer, 18S V3-V5) were amplified successfully, with negative procedural blanks.
RESEARCH PROPOSAL/OBJECTIVES:
The internship aims to compare, on the same DNA extracts, two molecular strategies for characterizing the eDNA content of the sediment-trap samples: (1) metabarcoding, targeting one or several marker genes with reference databases (12S « fish » primers already run by W-J. Chen, results expected by the end of 2026; COI Leray et al. 2013 primers, to be run in 2027), and (2) shotgun metagenomics, based on massive sequencing compared against genome databases (a first calibration test was sent to the ICM platform in 2026). The specific objectives are to:
1. Compare the taxonomic resolution and respective biases of metabarcoding and metagenomics on the KANADEEP2 extracts, and assess whether organisms that entered the traps alive are well represented or if environmental DNA dominates (e.g. from marine snow).
2. Cross-validate the taxonomic assignments against the extensive taxonomic and molecular reference collections built over the last 20 years of the TDSB program (fishes, molluscs, decapods, sponges, cnidarians, echinoderms, annelids), to compensate for the poor coverage of public databases for deep-sea bathyal fauna.
3. Contribute to defining a simple, low-impact, low-tech protocol (choice of fixative, sampling frequency) that could be transferred to autonomous observatories such as ScInObs for the long-term monitoring of deep-sea benthic biodiversity.
DESCRIPTION OF DATA :
The candidate will work with DNA extracts already obtained from 8 RNAlater sediment-trap cups from the KANADEEP2 cruise (particle fraction, 0.3 mm sieve; supernatant fraction, 20 µm and 0.22 µm filtration). These extracts have already yielded successful amplification of standard COI and 18S markers. 12S « fish » metabarcoding results (W-J. Chen) are expected by the end of 2026, and a shotgun metagenomics calibration test (2 extracts, ICM platform) is ongoing in 2026. New COI metabarcoding data (Leray et al. 2013 primers) and a selection of shotgun metagenomics libraries will be produced during 2027, with sequencing informed by the 2026 preliminary results. Part of the data will be available before the internship begins in January 2027, and will be completed with further sequencing runs during the internship.
METHODOLOGIES:
Bioinformatic analysis of metabarcoding data (quality filtering, denoising/OTU or ASV clustering, taxonomic assignment against public and MNHN in-house reference databases) using standard pipelines (e.g. OBITools, DADA2/QIIME2).
Comparative analysis of shotgun metagenomics reads (taxonomic profiling against reference genome/protein databases), following approaches such as Baloglu et al. (2021) and Armbrecht et al. (2021).
Cross-validation of taxonomic assignments against the DNA barcode and reference genome resources built by the MNHN taxonomic network for the seamount fauna (fish, molluscs, decapods, sponges, cnidarians, echinoderms, annelids).
Comparative assessment of the taxonomic resolution, cost and technical constraints of the two approaches, in view of proposing a simplified protocol usable in autonomous, low-impact benthic biodiversity observatories.
REFERENCES:
Richer de Forges, B., Koslow, J. A., & Poore, G. C. B. (2000). Diversity and endemism of the benthic seamount fauna in the southwest Pacific. Nature, 405(6789), 944-947.
Leray, M., Yang, J.Y., Meyer, C.P., Mills, S.C., Agudelo, N., Ranwez, V., Boehm, J.T., Machida, R.J. (2013). A new versatile primer set targeting a short fragment of the mitochondrial COI region for metabarcoding metazoan diversity. Frontiers in Zoology, 10, 34.
Baloglu, B., Chen, Z., Elbrecht, V., Braukmann, T., MacDonald, S., Steinke, D. (2021). A workflow for accurate metabarcoding using nanopore MinION sequencing. Methods in Ecology and Evolution, 12, 794-804.
Armbrecht, L., Eisenhofer, R., Utge, J., Sibert, E.C., Rocha, F., Ward, R., Pierella Karlusich, J.J., Tirichine, L., Norris, R., Summers, M., Bowler, C. (2021). Paleo-diatom composition from Santa Barbara Basin deep-sea sediments: a comparison of 18S-V9 and diat-rbcL metabarcoding vs shotgun metagenomics. ISME Communications, 1, 66.
Arsenault-Pernet, E.-J., Brulport, J.-P., Samadi, S., & Olu, K. (2026). Fauna results from sediment traps on the Stylaster and Munida seamounts south of New Caledonia (Kanadeep2 and Kanarecup cruise). SEANOE. https://doi.org/10.17882/112062