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Enhancing groundwater metazoan metabarcoding through primer performance assessment and reference library updates
Tobias Hoeschel1, Hannah Rau2, Melanie Sinn2
1RPTU University Kaiserslautern-Landau, Institute for Environmental Sciences, Campus Landau, Fortstrasse 7, Landau 76829, Germany; Institute for Groundwater Ecology at the RPTU University Kaiserslautern-Landau, Campus Landau, Fortstrasse 7, Landau 76829, Germany.
This study enhances DNA metabarcoding for groundwater biodiversity by validating a new primer pair and updating reference databases. This improved molecular toolbox better reveals hidden aquatic fauna in subterranean ecosystems.
Area of Science:
- Environmental DNA (eDNA) metabarcoding
- Aquatic ecology
- Molecular biology
Background:
- Groundwater is a vital freshwater resource supporting unique stygofauna crucial for subterranean ecosystem functioning.
- DNA metabarcoding offers a powerful method to study these hidden communities, overcoming taxonomic challenges and identifying cryptic species.
- Current limitations in primer pair validation and reference database coverage hinder eDNA metabarcoding for groundwater metazoans.
Purpose of the Study:
- To strengthen the molecular toolbox for groundwater biodiversity research.
- To address the scarcity of validated primer pairs for groundwater invertebrates.
- To improve the taxonomic coverage of reference databases for subterranean fauna.
Main Methods:
- Evaluation of the primer pair fwhF2 / fwhR2n targeting the mitochondrial COI region for amplifying groundwater invertebrates.
- Development of the Groundwater invertebrate Database (GwinD) with updated taxonomic information.
- Application of the enhanced toolbox in a case study of central European groundwater communities.
Main Results:
- The primer pair fwhF2 / fwhR2n demonstrated effective amplification of central European groundwater invertebrates, especially crustaceans.
- Significant updates were made to public databases, including 30 Copepoda, five Bathynellacea, and five Ostracoda species.
- The study successfully applied the toolbox for eDNA metabarcoding, revealing community composition but noting limitations for non-crustacean groups.
Conclusions:
- The developed molecular toolbox, including the validated primer pair and updated database, significantly advances groundwater biodiversity assessment.
- This research improves the ability to monitor groundwater ecosystems and assess the impacts of environmental changes.
- Further efforts are needed to expand database coverage for non-crustacean groundwater fauna.
