Topeka Shiner (Notropis topeka) is an endangered fish species that inhabits oxbows, floodplains, and small headwater streams throughout the central United States. Habitat loss and alterations represent the main threats to this species and conservation actions have included restoring oxbows that were historically occupied. Information on species occupancy is critical for evaluating the success of habitat restoration efforts, however, collection efforts can be difficult. Traditionally, sampling for Topeka Shiners has relied on seining. The effectiveness of seine surveys depends largely on habitat characteristics including oxbow size, depth, and substrate. Environmental DNA surveys have emerged as a method to effectively survey across broad landscapes and collect data on species occupancy. In this study our objectives were to: 1) Refine eDNA laboratory methods for Topeka Shiner (specifically increase the sensitivity of existing methods); and 2) Evaluate multiple eDNA field collection methods in order to make recommendations for future Topeka Shiner monitoring efforts. We modified an existing eDNA assay by adding a hydrolysis probe, which resulted in a highly sensitive tool for Topeka Shiner eDNA detection. From 2021 to 2022, we compared three different eDNA collection methods to evaluate their effectiveness: surface water grabs with centrifugation, surface water grabs with vacuum manifold filtration, and on-site surface water filtration using Smith Root filter packs. eDNA sampling at oxbows was followed by seine surveys to confirm species presence/absence. At each site where Topeka Shiners were collected in seine surveys, we also detected Topeka Shiner eDNA. Control sites where Topeka Shiners were presumed absent and no individuals were collected in seine surveys, had no eDNA detections. At one site with unknown Topeka Shiner status, no individuals were collected via seine surveys, but we did detect eDNA. Logistic regression showed that sampling method (i.e., centrifuge, vacuum filtration, Smith Root filtration) was a significant factor predicting detection rate, with centrifugation and Smith Root filtration having higher overall detection rates compared to vacuum filtration. We used linear regression to evaluate which sampling method collected the greatest amount of eDNA. Once again, collection method did have a significant effect on the number of copies per liter collected. At sites sampled in 2021, centrifugation had much higher copies detected than vacuum filtration. In 2022, there was much greater variability in the number of copies detected, and the comparison between vacuum filtration and Smith Root filtration was much more site dependent. For future sampling events, we suggest using Smith Root filters as the most efficient and economical means for collecting eDNA samples for Topeka Shiners.
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