Meta-Fish-Lib: A generalised, dynamic DNA reference library pipeline for metabarcoding of fishes

Area of Science:

• Ecology
• Genomics
• Bioinformatics

Background:

• DNA metabarcoding accuracy relies on comprehensive and high-quality reference libraries.
• Public repositories like NCBI and BOLD offer vast sequence data but lack curated options for specific research needs.

Purpose of the Study:

• To provide an automated pipeline for downloading, cleaning, and annotating fish mitochondrial DNA sequences.
• To create a curated, versioned, and citable reference library for fish metabarcoding.
• To improve the coverage of U.K. fish species for environmental DNA metabarcoding using the MiFish-12S marker.

Main Methods:

• Developed a pipeline supporting multiple DNA metabarcode markers, species synonym searches, and taxonomic validation.
• Implemented phylogeny-assisted quality control to identify and remove misannotated sequences.
• Generated new MiFish-12S sequences for 88 U.K. marine species and created a curated reference library (Meta-Fish-Lib).

Main Results:

• The pipeline successfully downloads, cleans, and annotates mitochondrial DNA sequence data.
• A ready-to-use reference library for U.K. marine and freshwater fishes is provided.
• New MiFish-12S sequences increased coverage for common U.K. species to 93%.

Conclusions:

• The Meta-Fish-Lib pipeline and library significantly enhance the accuracy and reliability of fish DNA metabarcoding.
• This resource facilitates scalable fish biodiversity assessments using environmental DNA.
• The automated pipeline and curated library address key limitations in current metabarcoding research.