Environmental DNA Assessment of Fish Diversity in the White River, Shadyside Lake, and Trail Creek, Indiana

Environmental DNA (eDNA) metabarcoding provides a non-invasive approach for describing fish-associated molecular diversity in freshwater ecosystems. We analyzed one composite water sample from each of four Indiana locations: White River upstream (WR UP), White River downstream (WR DN), Shadyside Lake (SS Lake), and Trail Creek, using MiFish mitochondrial 12S rRNA sequencing. A total of 81 fish-associated exact sequence variants (ESVs) were retained for analysis after excluding non-fish assignments and a positive control. Observed ESV richness was highest in WR DN (56), followed by WR UP (27), SS Lake (22), and Trail Creek (5). Shannon diversity was likewise highest in WR DN (H′ = 2.27) and lowest in Trail Creek (H′ = 0.78). Twenty-one fish-associated ESVs were shared between the White River samples, which also showed the lowest Bray–Curtis dissimilarity (0.204) among site pairs. Processed fish-assigned read totals varied markedly (322–23,127), limiting direct richness comparisons, especially for Trail Creek. These findings provide a descriptive baseline of fish-associated eDNA assemblages but do not establish differences in fish abundance or replicated site-level biodiversity.