Fig 1.
Map of primers and amplicons tested in this study.
The reference sequence shown in black is Drosophila yakuba, cytochrome c oxidase region 1470–3009 bp (1540 nt). Secondary structure is shown for reference, comprised of six alpha helices in the standard DNA barcode region shown here.
Table 1.
COI amplicons used in this study.
Table 2.
Arthropoda ESV and read counts vary by COI amplicon.
Fig 2.
ESV richness continues to increase as COI amplicons are added but species—Order richness reaches a plateau.
For the primer comparison experiment that used the soil DNA extraction kit, we pooled the results from the six sites and show the top COI amplicon combinations that detected the greatest richness. We report the recovered richness when up to six amplicons are combined at the 1) ESV, 2) species, 3) genus, 4) family, and 5) order ranks. ESV = exact sequence variant; A = BR5; B = F230R; C = ml-jg; D = BF1R2; E = BF2R2; F = fwh1.
Fig 3.
Each amplicon differentially recovers site and water quality indicators.
In the top panel, the number of site indicator taxa from across the Arthropoda are shown. In the bottom panel, the number of typical water quality indicator taxa from the EPTC are shown. This analysis was based on normalized data. ESV = exact sequence variant; EPTC = Ephemeroptera, Plecoptera, Trichoptera, Chironomidae.
Fig 4.
Site indicator taxa chosen based on metabarcode sequencing are comprised of Coleoptera, Diptera, Ephemeroptera, and Trichoptera.
Presence is indicated by a dark square, absence by a white square. The total number of Arthropoda site indicator taxa detected by each amplicon is shown in the bottom row.
Fig 5.
Samples cluster mainly by site despite differences in amplicons and replicates.
Results are based normalized data. COI amplicons are labelled directly in the plot. Amplicons shown twice represent the two PCR replicates. Sites are grouped by color according to the legend.