DBP metabarcoding pipeline: An integrated tool for eDNA analysis of ONT data using the Indonesian marine fish database
Al Malik, M.D.; Aulya, M.; Cahyani, N.K.D. (2026). DBP metabarcoding pipeline: An integrated tool for eDNA analysis of ONT data using the Indonesian marine fish database. Regional Studies in Marine Science 95: 104868. https://dx.doi.org/10.1016/j.rsma.2026.104868
In: Regional Studies in Marine Science. Elsevier: Amsterdam. ISSN 2352-4855, more
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| Author keywords |
EDNA metabarcoding; Nanopore; 12S; Curated database; Biodiversity; Indonesia |
| Authors | | Top |
- Al Malik, M.D.
- Aulya, M.
- Cahyani, N.K.D.
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| Abstract |
Environmental DNA (eDNA) metabarcoding with ONT Nanopore has been increasingly applied in Indonesia, a global megabiodiversity hotspot, as a complement to traditional biodiversity surveys such as visual census. However, the development of user-friendly bioinformatics pipelines remains an ongoing challenge, particularly for processing and interpreting ONT-based eDNA metabarcoding data. Here, we present the Diponegoro Biodiversity Project (DBP) Metabarcoding Pipeline, designed to streamline the processing of ONT sequence outputs for metabarcoding applications. The pipeline integrates sequential steps of quality filtering with NanoFilt, primer trimming with Cutadapt, OTU clustering and chimera removal with VSEARCH, and taxonomic assignment using BLASTn followed by a Lowest Common Ancestor (LCA) approach. To facilitate application in the context of the Indonesian region, we developed a region-specific reference database (“LocalDB”) comprising 1885 marine fish species, and a combined database (“LocalDB + Pre-made”) covering 2163 marine fish species from the 12S MiFish region, representing Actinopteri and Chondrichthyes. We applied the pipeline with three reference sets (LocalDB, Pre-made, and LocalDB + Pre-made) to eDNA datasets from five marine water samples to evaluate taxonomic resolution and alpha diversity difference. Results showed no significant differences in taxonomic resolution at the genus and species levels, nor in biodiversity metrics across the three databases, indicating that the use of a local database did not alter ecological inferences compared with non–region-specific databases. However, the local database offers the advantage of being curated with native species, reducing misidentifications and false positives. Overall, the DBP pipeline and curated databases provide an integrated protocol for rapid assessment and marine biodiversity monitoring in Indonesia. The resources are available at https://github.com/mdaniealmalik/DBP_Metabarcoding_Pipeline.git |
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