Characterizing soil eukaryotic diversity from NEON metagenomics datasets - R code and input files

Belowground eukaryotic diversity serves a vital role in soils and thus ecosystem functioning, yet the composition, structure, and macroecology of these communities is significantly under-characterized. The National Ecological Observatory Network (NEON) conducts long-term surveillance of numerous taxa and ecosystem properties and processes (e.g., soil and water-extracted metagenomes, above- and belowground abiotic variables such as soil moisture, temperature, and solar irradiation, and abundance data on meso- and macroscopic organisms, such as beetles and birds) and provides them publicly. However, the soil metagenomics provided by NEON are not routinely evaluated for their micro- and meso-eukaryotic component, in part because analyzing metagenomes for eukaryotic sequences brings unique challenges such as low relative sequence abundance, large genomes, and a more complex gene structure and is not yet mainstream in many molecular ecology pipelines. In this study, we mined the publicly available, well supported, and ecologically comprehensive NEON metagenomics datasets originally collected for soil prokaryotes using a custom-built pipeline and produced a preliminary assessment of biodiversity trends in North American soil eukaryotes. We used data from 1455 samples collected from 495 plots across 45 NEON sites spanning 11 biomes to extract ~800 18S rRNA reads per sample (22,000 reads per site), which corresponded to 5190 OTUs in 35 distinct phyla and represents the first large-scale soil eukaryote analysis using NEON data. We then tested the data to see whether taxonomic richness and community structure reflected similar patterns to previously established ecological trends. For example, we found that eukaryotic richness was negatively correlated with pH. We also found that paired heavily managed sites had 43% lower eukaryotic richness than low management sites. When sorting samples into biomes, we found that most biomes had a distinct eukaryotic community (significant PERMANOVA) from other biomes except for the forest biomes. Lastly, we found that fire decreased eukaryotic richness at one site. These findings are in line with generally accepted ecological trends and support the notion that NEON soil metagenome datasets can and should be used to further explore spatiotemporal patterns in soil fauna diversity, its associations with ecosystem functioning, and responses to environmental changes in North America