DOI: 10.5061/dryad.n8pk0p342
title: Data from: Leveraging environmental DNA (eDNA) to optimize targeted removal of invasive fishes

description: Natural resource managers need innovative and cost-effective methodologies
 that enable the targeted removal of aquatic invasive species
 (AIS). Removing AIS before they establish and spread into critical habitat
 for native species can mitigate invasions in freshwater systems
 and preserve ecosystem integrity. To address this need, we
 established protocols using the detection of environmental DNA (eDNA) to
 guide deployment of traditional fisheries trapping methods for
 invasive fish species removal. In a pilot study during spring 2022,
 we set minnow traps (one per site) in a spatially stratified
 random design. We also developed a quantitative polymerase chain
 reaction (qPCR) assay to detect eDNA from multiple closely related
 invasive loach species (family Cobitidae) and compared detections of eDNA
 with detections of fish using minnow traps. At sites where both
 eDNA and minnow traps were deployed, the two methods agreed on the
 presence of loaches approximately 79% of the time (95% CI:
 60%-90%). Based on the rate at which minnow traps failed to
 detect loaches when eDNA samples were positive (22%; 95% CI:
 11%-40%), we estimated that setting three or more replicate traps
 per site would improve detections with gear. This information was used to
 inform a more comprehensive study in spring 2023. This main study
 consisted of two phases: (1) a fixed-point DNA study to calibrate
 a model of dispersal and attenuation rate, and (2) a loach removal phase.
 In the removal phase, we randomly selected sites to sample for
 loach eDNA, plotted eDNA concentration as a GIS layer to
 develop heatmaps, and then placed 10 replicate traps at sites
 with the highest concentrations. A total of 658 loaches were
 removed from 68 of 77 eDNA-positive trapped sites. Our results indicate
 that aquatic invasive species removal is more efficient when eDNA
 detection techniques are combined with traditional
 trapping methods.
description: See briefly summarized methods in the associated ReadMe file of
 this dataset. For more detailed methods see methods described in main
 manuscript.
description: # Data from: Leveraging Environmental DNA (eDNA) to Optimize Targeted
 Removal of Invasive Fishes
 \[[https://doi.org/10.5061/dryad.n8pk0p342](https://doi.org/10.5061/dryad.n8pk0p342)] ## General information ### Author Information Corresponding author: Jennie J. Wiggins Fish Biologist, Lodi Fish and Wildlife Office US Fish and Wildlife Service [jennie_wiggins@fws.gov](mailto:jennie_wiggins@fws.gov) Co-Authors: Vanessa D. Tobias Mathematical Statistician, Lodi Fish and Wildlife Office US Fish and Wildlife Service [vanessa_tobias@fws.gov](mailto:vanessa_tobias@fws.gov) Erika F. Holcombe Supervisory Fish Biologist, Lodi Fish and Wildlife Office US Fish and Wildlife Service [erika_holcombe@fws.gov](mailto:erika_holcombe@fws.gov) Katie Karpenko Research Associate, Genidaqs Laboratory Cramer Fish Sciences [katie.karpenko@fishsciences.net](mailto:katie.karpenko@fishsciences.net) Eric R. Huber Supervisory Fish Biologist, Lodi Fish and Wildlife Office US Fish and Wildlife Service [eric_huber@fws.gov](mailto:eric_huber@fws.gov) Andrew C. Goodman Fish Biologist, Lodi Fish and Wildlife Office US Fish and Wildlife Service [andrew_goodman@fws.gov](mailto:andrew_goodman@fws.gov) Date of data collection: Pilot Study- March 2022; Main Study (including calibration experiment)- February 2023 – June 2023 Geographic location of data collection: All data were collected at San Luis National Wildlife Refuge (Merced County, California, USA) ## Data and file overview This dataset contains 8 CSV files: • All_Variable_descriptions.csv • catch_loach_data.csv • loach_length_data.csv • eDNA_detections_site_data.csv • Resampled_eDNA_detections_site_data.csv • PilotStudy_qPCR_RAW.csv • CalibrationExperiment_qPCR_RAW.csv • MainStudy_qPCR_RAW.csv ### File descriptions: All_Variable_descriptions.csv contains 4 columns and 151 rows of descriptions for all the variables used within the 7 other CSV files associated with this dataset. catch_loach_data.csv contains 21 variables and 897 (898 with column headers) rows of data related to loach capture per minnow trap collected during trapping efforts for the Pilot Study and the Main Study. This file only contains data related to trapped sites. Columns are study, round, site_name, trap_name, habitat_type, trap_num, latitude, longitude, date_set, time_set, date_pulled, time_pulled, total_time_fished, loach_captured, dissolved_oxygen, conductivity, temperature, turbidity, depth, velocity, notes. A complete description of all the variables used in this file can be found in the All_Variable_descriptions.csv file. Environmental data has not been scrubbed for outliers. loach_length_data.csv contains 16 variables and 676 (677 with column headers) rows of length data for each loach collected during trapping efforts for the Pilot Study and the Main Study. Columns are study, round, site_name, trap_name, habitat_type, trap_num, latitude, longitude, date_set, time_set, date_pulled, time_pulled, species_name, species_code, total_length, notes. A complete description of all the variables used in this file can be found in the All_Variable_descriptions.csv file. eDNA_detections_site_data.csv contains 16 variables and 377 (378 with column headers) rows of categorical and environmental data related to all sites sampled for eDNA and/or trapped during the Pilot Study or Main Study. Columns are study, round, site_name, habitat_type, latitude, longitude, date, sampled_eDNA, detection, trapped, dissolved_oxygen, conductivity, temperature, turbidity, depth, velocity. A complete description of all the variables used in this file can be found in the All_Variable_descriptions.csv file. Environmental data has not been scrubbed for outliers. Resampled_eDNA_detections_site_data.csv contains 15 variables and 38 (39 with column headers) rows of categorical and environmental data related to sites that were re-sampled for eDNA if loach were captured there during trapping efforts for the Main Study. Columns are study, round, site_name, habitat_type, latitude, longitude, date, detection, dissolved_oxygen, conductivity, temperature, turbidity, depth, velocity, notes. A complete description of all the variables used in this file can be found in the All_Variable_descriptions.csv file. Environmental data has not been scrubbed for outliers. PilotStudy_qPCR_RAW.csv contains 20 variables and 540 (541 with column headers) rows of qPCR results for all filters collected during the Pilot Study. Columns are projectID, project_type, extraction_order, date, site_name, filter_replicate, FilterID, vol_filtered_ml, location, location_broad, extraction_date, control, month, year, field_notes, technical_replicate, Cq, target, relative_conc_ng/uL, relative_conc_weighted_by_vol. A complete description of all the variables used in this file can be found in the All_Variable_descriptions.csv file. CalibrationExperiment_qPCR_RAW.csv contains 33 variables and 630 (631 with column headers) rows of qPCR results for all filters collected during the eDNA calibration experiment of the Main Study. Columns are projectID, project_type, extraction_order, date, name, filter_replicate, FilterID, vol_filtered_ml, location, location_broad, extraction_date, lat, lon, control, month, year, field_notes,dist_lc_m, lc_deploy_time, lc_remove_time, lc_spp_1, lc_spp_2, spp_1_wt_g, spp_2_wt_g , vel_m_s, vel_ft_sec, turb_ntu, depth_m, technical_replicate, Cq, relative_conc_ng/uL, target. A complete description of all the variables used in this file can be found in the All_Variable_descriptions.csv file. MainStudy_qPCR_RAW.csv contains 22 variables and 2674 (2675 with column headers) rows of qPCR results for all filters collected during the Main Study. Columns are projectID, project_type, extraction_order, date, site_name, filter_replicate, FilterID, vol_filtered_ml, location, location_broad, extraction_date, lat, lon, control, month, year, field_notes, technical_replicate, Cq, target, relative_conc_ng/uL, relative_conc_weighted_by_vol. A complete description of all the variables used in this file can be found in the All_Variable_descriptions.csv file. ### Methodological information Please refer to the associated paper for a more detailed overview of data collection, processing, and analyses for this dataset. eDNA sample collection: Environmental DNA samples were collected by simultaneously filtering water through three Millipore® Sterivex™-HV 0.45 μm sterile PVDF membrane filter units (Millipore Sigma, Burlington, MA, USA) using three Masterflex® L/S Easy-Load II peristaltic pumps. Environmental DNA field controls were collected at the start of each sampling day to confirm that field equipment was free of contamination. Pilot Study: We selected 127 random sampling sites made up of an approximately equal number of habitat types termed ponds (n=62) and canals (n=65). One baited minnow trap was set at each of the selected locations for approximately 24 h. A random subset of sites was also sampled for eDNA, including 20 canal sites and eight pond sites. We measured the total length (TL) of all trapped loaches to the nearest mm. Species identification was genetically confirmed for each specimen captured resulting in 17 *Paramisgurnus dabryanus* (Large-Scale Loach) and one *Misgurnus mizolepis* (Fine-Scale Loach). Calibration of eDNA: To better understand eDNA transport and persistence within the sampling area, we conducted a fixed-point eDNA study in a small shallow canal and a large deep canal. We created a fixed source of eDNA using known quantities of commercially available frozen *Engraulis mordax* (Northern Anchovy; 16 g) and *Hemiramphus brasiliensis* (Ballyhoo; 100 g). Velocity and turbidity were measured prior to sampling using a Global Water FP111 Flow Probe (YSI Inc., Yellow Springs, OH, USA) and a Hach 2100Q Turbidity Meter (Hach Co., Loveland, CO, USA), respectively. After the calculated dispersal time had passed, we collected eDNA samples simultaneously in triplicate at six set distances from the tethered fish. Once complete, eDNA sources were removed from each canal. The eDNA collection process was repeated at the same locations the following day to determine if eDNA from the proxy species remained in the system after 24 h. Main Study: We sampled 250 of 300 randomly generated canal sites for loach eDNA. Pond habitats were not sampled in the main study.  Environmental DNA sampling (week 1, 3, 5) and loach trapping (week 2, 4, 6) alternated weekly to permit strategic deployment of traps based on the previous week’s eDNA results. Each set of an eDNA sampling week and its associated trapping week was referred to as one “round.” Areas containing the highest concentrations of loach eDNA received priority for trapping. Two additional weeks (week 7-8; extra_1, extra_2) of trapping only were conducted after the eDNA filter budget was exhausted. At sites where loach eDNA was detected, we deployed 10 minnow traps over a total of 260 m moving upstream. Traps soaked for approximately 24 h before they were removed. Sites where loach were captured, were resampled for eDNA the following week (designated by an ‘RX’ in the site name). We measured the TL of all trapped loaches to the nearest mm. Species identification for the loaches collected during this study have not been genetically confirmed and thus have been assigned a generic species name and code at the time of this publication. Environmental data were collected at each site one time before eDNA sampling and one time before trapping.  ## Code/Software The statistical code for reproducing the analyses in the associated manuscript can be found at [https://github.com/USFWS/LFWO-Loach-eDNA.git](https://github.com/USFWS/LFWO-Loach-eDNA.git). The scripts were created using R version 4.3.1 "Beagle Scouts".