Volume 43 Issue 2
Mar.  2022
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Hui-Ting Ruan, Rui-Li Wang, Hong-Ting Li, Li Liu, Tian-Xu Kuang, Min Li, Ke-Shu Zou. Effects of sampling strategies and DNA extraction methods on eDNA metabarcoding: A case study of estuarine fish diversity monitoring. Zoological Research, 2022, 43(2): 192-204. doi: 10.24272/j.issn.2095-8137.2021.331
Citation: Hui-Ting Ruan, Rui-Li Wang, Hong-Ting Li, Li Liu, Tian-Xu Kuang, Min Li, Ke-Shu Zou. Effects of sampling strategies and DNA extraction methods on eDNA metabarcoding: A case study of estuarine fish diversity monitoring. Zoological Research, 2022, 43(2): 192-204. doi: 10.24272/j.issn.2095-8137.2021.331

Effects of sampling strategies and DNA extraction methods on eDNA metabarcoding: A case study of estuarine fish diversity monitoring

doi: 10.24272/j.issn.2095-8137.2021.331
Funds:  This work was supported by the National Natural Science Foundation of China (32102793), National Key R&D Program of China (2018YFD0900802), Central Public-Interest Scientific Institution Basal Research Fund, South China Sea Fisheries Research Institute, CAFS (2019TS13, 2021SD18), Key Special Project for Introduced Talents Team of Southern Marine Science and Engineering Guangdong Laboratory (Guangzhou) (GML2019ZD0605), Open Fund Project of Key Laboratory of Offshore Fishery Development of Ministry of Agriculture and Rural Affairs (LOF 2020-02), and China-ASEAN Maritime Cooperation Fund (CAMC-2018F)
More Information
  • Environmental DNA (eDNA) integrated with metabarcoding is a promising and powerful tool for species composition and biodiversity assessment in aquatic ecosystems and is increasingly applied to evaluate fish diversity. To date, however, no standardized eDNA-based protocol has been established to monitor fish diversity. In this study, we investigated and compared two filtration methods and three DNA extraction methods using three filtration water volumes to determine a suitable approach for eDNA-based fish diversity monitoring in the Pearl River Estuary (PRE), a highly anthropogenically disturbed estuarine ecosystem. Compared to filtration-based precipitation, direct filtration was a more suitable method for eDNA metabarcoding in the PRE. The combined use of DNeasy Blood and Tissue Kit (BT) and traditional phenol/chloroform (PC) extraction produced higher DNA yields, amplicon sequence variants (ASVs), and Shannon diversity indices, and generated more homogeneous and consistent community composition among replicates. Compared to the other combined protocols, the PC and BT methods obtained better species detection, higher fish diversity, and greater consistency for the filtration water volumes of 1 000 and 2 000 mL, respectively. All eDNA metabarcoding protocols were more sensitive than bottom trawling in the PRE fish surveys and combining two techniques yielded greater taxonomic diversity. Furthermore, combining traditional methods with eDNA analysis enhanced accuracy. These results indicate that methodological decisions related to eDNA metabarcoding should be made with caution for fish community monitoring in estuarine ecosystems.
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