Effects of operational taxonomic unit inference methods on soil microeukaryote community analysis using long-read...

Area of Science:

• Soil ecology
• Microbial ecology
• Molecular biology

Background:

• Long amplicon metabarcoding enables phylogenetic analysis of soil microeukaryotes.
• Soil harbors vast, largely uncharacterized microeukaryotic communities.

Purpose of the Study:

• To evaluate the impact of three operational taxonomic unit (OTU) generation methods on soil microeukaryote community analysis.
• To assess how different OTU methods affect richness estimates, taxa identification, and recovery of ecological patterns.

Main Methods:

• PacBio SMRT sequencing of ITS and LSU rDNA regions from grassland soil samples.
• Application of three distinct OTU generation methods.
• Sampling across a moisture gradient with contrasting plant communities (wet, Carex acuta-dominated vs. mesic-dry, grass-dominated).

Main Results:

• Soil communities were dominated by protists (including Ciliophora) and fungi (30-40% of reads).
• Ecological patterns related to environmental conditions were consistently recovered across all OTU methods.
• OTU generation methods significantly influenced richness estimates and taxonomic/phylogenetic resolution.

Conclusions:

• While ecological patterns are robust, the choice of OTU method affects the perceived diversity and identification accuracy of soil microeukaryotes.
• Understanding these methodological differences is critical for accurate characterization of belowground biodiversity.