RT Journal Article T1 Enhancing metabarcoding efficiency and ecological insights through integrated taxonomy and DNA reference barcoding: A case study on beach meiofauna A1 Macher, Jan Niklas A1 Martínez, Alejandro A1 Çakir, Sude A1 Cholley, Pierre Etienne A1 Christoforou, Eleni A1 Curini Galletti, Marco A1 Van Galen, Lotte A1 García Cobo, Marta A1 Jondelius, Ulf A1 De Jong, Daphne A1 Leasi, Francesca A1 Lemke, Michael A1 Rubio Lopez, Iñigo A1 Sánchez Santos, Nuria A1 Sørensen, Martin Vinther A1 Todaro, M. Antonio A1 Renema, Willem A1 Fontaneto, Diego AB Molecular techniques like metabarcoding, while promising for exploring diversity of communities, are often impeded by the lack of reference DNA sequences available for taxonomic annotation. Our study explores the benefits of combining targeted DNA barcoding and morphological taxonomy to improve metabarcoding efficiency, using beach meiofauna as a case study. Beaches are globally important ecosystems and are inhabited by meiofauna, microscopic animals living in the interstitial space between the sand grains, which play a key role in coastal biodiversity and ecosystem dynamics. However, research on meiofauna faces challenges due to limited taxonomic expertise and sparse sampling. We generated 775 new cytochrome c oxidase I DNA barcodes from meiofauna specimens collected along the Netherlands' west coast and combined them with the NCBI GenBank database. We analysed alpha and beta diversity in 561 metabarcoding samples from 24 North Sea beaches, a region extensively studied for meiofauna, using both the enriched reference database and the NCBI database without the additional reference barcodes. Our results show a 2.5-fold increase in sequence annotation and a doubling of species-level Operational Taxonomic Units (OTUs) identification when annotating the metabarcoding data with the enhanced database. Additionally, our analyses revealed a bell-shaped curve of OTU richness across the intertidal zone, aligning more closely with morphological analysis patterns, and more defined community dissimilarity patterns between supralittoral and intertidal sites. Our research highlights the importance of expanding molecular reference databases and combining morphological taxonomy with molecular techniques for biodiversity assessments, ultimately improving our understanding of coastal ecosystems. PB Wiley SN 1755-098X YR 2024 FD 2024 LK https://hdl.handle.net/20.500.14352/118016 UL https://hdl.handle.net/20.500.14352/118016 LA eng NO Macher, J.-N., Martínez, A., Çakir, S., Cholley, P.-E., Christoforou, E., Curini Galletti, M., van Galen, L., García-Cobo, M., Jondelius, U., de Jong, D., Leasi, F., Lemke, M., Rubio Lopez, I., Sánchez, N., Sørensen, M. V., Todaro, M. A., Renema, W., & Fontaneto, D. (2024). Enhancing metabarcoding efficiency and ecological insights through integrated taxonomy and DNA reference barcoding: A case study on beach meiofauna. Molecular Ecology Resources, 24, e13997. https://doi.org/10.1111/1755-0998.13997 NO This study was funded by the Biology, Ecology, Nature (BEN) grant no. T0206/37197/2021/kg of the Stemmler-Stiftung to J-NM. NO Stemmler Stiftung DS Docta Complutense RD 9 abr 2025