Putting eDNA to the Test: A Field Comparison of eDNA Metabarcoding to Established Protocols for Assessing Biodiversity in Missouri’s Ozark Highland Streams

Alternative Title

eDNA Metabarcoding in Ozark Highland Streams


Aquatic biodiversity monitoring to inform conservation and management efforts in stream systems has increasingly begun to incorporate eDNA-based sampling methods. We conducted a comparison of eDNA metabarcoding to a traditional protocol of combined seining and electrofishing methods to assess fish biodiversity of wadeable stream sites in six separate drainages in the Ozark Highlands of Missouri. The study further focused on the headwaters of the Meramec River, which included eleven sites and seasonal sampling (summer and winter). We compared estimates of diversity across sampling methods, assessed hypothesized relationships of habitat (depth and velocity) and season (summer vs winter) to eDNA detection, and tested the effects of sampling method and site locality on fish assemblage structural dynamics. eDNA sampling detected approximately double the number of species detected compared to traditional methods, providing higher diversity estimates while maintaining the relative ranking of sites. eDNA detection probabilities were positively associated with stream depth and velocity and were generally higher in summer than in winter but not for all species. Assemblage differences between tributary and mainstem sites were attributable to a small number of species that were found predominantly in one stream size category or the other, indicating that eDNA was sensitive to within-drainage assemblage structure relationships. Improved species detection and a more comprehensive understanding of assemblage structural dynamics are important benefits encouraging the use of eDNA metabarcoding as a primary collection method in future stream biodiversity assessment and monitoring programs.


Biological Sciences

Keywords and Phrases

DNA, Environmental; Biological Monitoring; Biodiversity; Stream; Fresh Water, Seine, Electrofishing, Metabarcoding

Document Type

Article - Preprint

Document Version


File Type





© 2023 The Authors, all rights reserved.

Publication Date


File 1_ Moreau sample reads.xlsx (11 kB)
File 2_notes on contents of files.txt (1 kB)
File 3_ StFrancis and Whitewater sample reads.xlsx (23 kB)
File 4_ Beaver Creek sample reads.xlsx (12 kB)
File 5_ Current sample reads.xlsx (15 kB)
File 6_ Meramec sample reads.xlsx (49 kB)
File 7_ Mifish_species_table_meramec_all.xlsx (87 kB)
File 8_ Mifish_species_table_petsmartallsamples.xlsx (18 kB)
File 9_ Mifish_species_table_SMCr.xlsx (11 kB)
File 10_ Mifish_species_table_StFrancis.xlsx (26 kB)
File 11_ Mifish_species_table_WW.xlsx (13 kB)
File 12_ Chord16s_species_table_LBC.xlsx (14 kB)
File 13_ Chord16s_species_table_meramec_summer.xlsx (42 kB)
File 14_ Chord16s_species_table_meramec_winter.xlsx (57 kB)
File 15_ Chord16s_species_table_petsmartallsamples.xlsx (21 kB)
File 16_ Chord16s_species_table_SMCr.xlsx (12 kB)
File 17_ Chord16s_species_table_WW.xlsx (15 kB)
File 18_ Mifish_species_table_Current.xlsx (17 kB)
File 19_ Mifish_species_table_LBC.xlsx (12 kB)
File 20_ Reference_StFrancis12s (002).fasta.txt (139 kB)
File 21_ Reference_moreau16s.fasta.txt (12 kB)
File 22_ Reference_moreau12s.fasta.txt (86 kB)
File 23_ Reference_Meramec16s.fasta.txt (21 kB)
File 24_ Reference_Meramec12s.fasta.txt (132 kB)
File 25_ Reference_JamesFinley16s.fasta.txt (12 kB)
File 26_ Reference_JamesFinley12s.fasta.txt (86 kB)
File 27_ Reference_Current16s.fasta.txt (20 kB)
File 28_ Reference_Current12s.fasta.txt (130 kB)
File 29_ Missing species Whitewater 16s.txt (1 kB)
File 30_ Missing species StFrancis16s.txt (1 kB)
File 31_ Missing species StFrancis12s.txt (1 kB)
File 32_ Missing species moreau16s.txt (1 kB)
File 33_ Missing species moreau12s.txt (1 kB)
File 34_ Missing species Meramec16s.txt (1 kB)
File 35_ Missing species Meramec12s.txt (1 kB)
File 36_ Missing species JamesFinley16s.txt (1 kB)
File 37_ Missing species JamesFinley12s.txt (1 kB)
File 38_ Missing species Current16s.txt (1 kB)
File 39_ Missing species Current12s.txt (1 kB)
File 40_ Species list Whitewater.txt (3 kB)
File 41_ Species list StFrancis.txt (4 kB)
File 42_ Species list Moreau.txt (2 kB)
File 43_ Species list Meramec.txt (3 kB)
File 44_ Species list JamesFinley.txt (2 kB)
File 45_ Species list Current.txt (4 kB)
File 46_ Reference_Whitewater16s.fasta.txt (19 kB)
File 47_ Reference_Whitewater12s.fasta.txt (126 kB)
File 48_ Reference_StFrancis16s.fasta.txt (21 kB)

This document is currently not available here.