mkLTG: a command-line tool for taxonomic assignment of metabarcoding sequences using variable identity thresholds.

Area of Science:

• Ecology
• Bioinformatics
• Genetics

Background:

• Metabarcoding is crucial for biodiversity studies, but accurate taxonomic assignment of environmental DNA sequences remains challenging.
• Current methods, like the lowest common ancestor (LCA) approach, often use fixed thresholds unsuitable for diverse taxonomic groups and variable database coverage.

Purpose of the Study:

• To introduce mkLTG, a novel LCA-based method for improved taxonomic assignment in metabarcoding.
• To develop a flexible tool adaptable to different databases, genetic markers, and taxonomic groups.

Main Methods:

• The mkLTG method employs a series of decreasing percentage of identity thresholds, allowing parameter customization for each level.
• Optimization was performed using the COI marker and a comprehensive, non-redundant reference database.
• mkLTG is a command-line application designed for easy integration into existing bioinformatics pipelines.

Main Results:

• The mkLTG method demonstrates adaptability across diverse taxonomic groups and reference databases.
• Benchmarking confirmed the tool's effectiveness and ease of use.

Conclusions:

• mkLTG offers a more robust and adaptable solution for taxonomic assignment in metabarcoding compared to fixed-threshold methods.
• The freely available tool and scripts facilitate its adoption in biodiversity research.