Surveying Biodiverse Coral Reef Ecosystems Across the Tree of Life With Environmental DNA

A central challenge in evaluating biodiversity using environmental DNA (eDNA) metabarcoding is that each primer set has biases resulting in selective amplification of taxonomic diversity. Here we test 11 primer pairs to assess Central Pacific coral reef diversity in the five kingdoms of marine eukaryotes: Chromista, Fungi, Metazoa, Plantae and Protozoa. Zero-width OTUs (ZOTUs) were compared to public sequence databases and resolved to the finest taxonomic rank possible. Across all primers, we recovered 421 distinct taxa from five kingdoms in 35 phyla, comprising 62 classes. The number of taxa and taxonomic groups recovered varied widely. Nearly all primers recovered a large fraction of metazoan taxa, and several also recovered a large portion of taxa belonging to Chromista. Based on our results, most primers we tested here can be clustered into four groups: (1) the 'fish' primers, specialized for fishes, (2) the 'diverse' primers, which recover the highest number of identifications at lower taxonomic levels, (3) the 'general' primers, which perform moderately well across the tree of life, and (4) the algae primers. Taxonomic assignments tended to decrease with taxonomic rank, from 86%-100% at the phylum level to 35%-77% at the species level. Primers specialized for specific groups (i.e., fishes) tended to recover fewer taxa at higher taxonomic resolution, while more general primers tended to recover more taxa, though at lower taxonomic resolution. While each primer set proved uniquely informative for some taxa, the taxon recovery profiles provided here should aid researchers in matching the taxonomic specificity of environmental metabarcoding primers with research questions of interest.