Marshal S. Hoy Carl O. Ostberg Dorothy M. Chase Austen C. Thomas 20251203 spreadsheet https://doi.org/10.5066/P13SYTQC The content contains metadata from an eDNA survey that assessed the efficacy of an autonomous eDNA sampler for tracking Chinook, sockeye, and coho salmon migrating through the Lake Washington Ship Canal between June and September 2022. The purpose of the data collection was to 1) compare daily sockeye, Chinook and coho salmon eDNA concentrations to the number of adults counted daily at the Ballard Locks to determine how well eDNA concentration estimates track with the adult counts and 2) determine if abiotic covariates affect detection probabilities. Data will be used to produce a package of model outputs which will be included in a manuscript for submission to a peer reviewed journal. 20220615 20221010 ground condition Complete None planned Lake Washington/ Sammamish River/ Cedar River watershed -122.4110 -121.9922 47.7947 47.4439 None Environmental DNA Salmon Autonomous sampling Population ecology USGS Metadata Identifier USGS:6924dbabd4be022d9b2fdf58 None Lake Washington Seattle, Washington State, USA Salmonid Chinook salmon sockeye salmon coho salmon Kingdom Animalia Subkingdom Bilateria Infrakingdom Deuterostomia Phylum Chordata Subphylum Vertebrata Infraphylum Gnathostomata Superclass Actinopterygii Class Teleostei Superorder Protacanthopterygii Order Salmoniformes Family Salmonidae Subfamily Salmoninae Genus Oncorhynchus Species Oncorhynchus tshawytscha TSN: 161980 Species Oncorhynchus nerka TSN: 161979 Species Oncorhynchus kisutch TSN: 161977 None. Please see 'Distribution Info' for details. None. Users are advised to read the dataset's metadata thoroughly to understand appropriate use and data limitations. Marshal Hoy USGS Western Fisheries Research Center mailing and physical

6505 NE 65TH ST

Seattle WA 98115 USA 206 526 6282 mhoy@usgs.gov All data have been cross-checked and are accurate All data match with methodological practices and fall within expected ranges. All data have been checked for duplication and omission and none were found. Data set is considered complete for the information presented, as described in the abstract. Users are advised to read the rest of the metadata record carefully for additional details. No formal positional accuracy tests were conducted. No formal positional accuracy tests were conducted. The intake for the eDNA autosampler was set to ~1M deep, located at the end of the King County Environmental Lab dock which is at approximately 3M total depth (47° 39’ 09.38 N, 122° 21’ 42.28 W). We performed 2 studies for data collection: a temporal study and a contamination carry over study. For the temporal study, we assessed the efficacy of an autonomous eDNA sampler for tracking Chinook, sockeye, and coho salmon migration through the Lake Washington Ship Canal between June and September 2022. Throughout the temporal study, the autosampler was programmed to collect eDNA samples twice per day. The first daily samples were collected 10:00am -3:00 pm (sampling times changed depending on the timing of autosampler servicing), and the second daily sample was always collected 12 hours after the first daily sample. For the eDNA carry over study, we assessed sample-to-sample contamination in the form of residual eDNA in the sampling apparatus that may be carried over from one sample to the next (performed on September 20, 2022). The study was achieved by filtering samples that alternated between the Lake Washington Ship Canal and deionized water, with the aim of identifying whether residual eDNA collected from the Lake Washington Ship Canal contaminated the filter samples collected from deionized water. For both studies, we used 5µm pore size self-preserving polyethersulfone filters (Smith-Root, Inc). Sample volumes were set to 5.0L or until a clogged filter was detected via a flow sensor. The autosampler was serviced once per week, which included cleaning the intake filter, collection and reloading of eDNA filters, changing the battery, and programming the next weeks scheduled deployment. Sample filters were placed in sealed packaging and stored at room temperature for 33-117 days until DNA extraction. 20221010 All laboratory protocols and analyses were designed to avoid cross-contamination. The eDNA workflow and sample preparation were separated into designated clean workstations, each with a UV hood, including a station where DNA was extracted (no amplified PCR products or highly concentrated target DNA sequences allowed), a station where PCR reagents were prepared and loaded, a station where DNA standards were diluted and loaded, and a station dedicated to PCR amplification. Sample preparation was performed in UV hoods using equipment dedicated to processing eDNA samples at each workstation. Workstations were decontaminated with UV and/or 10% bleach before and after each use. 20230111 To extract eDNA from filter samples, each whole filter was cut into eight 2-mm-wide strips with sterile scissors prior to DNA extraction and strips were incubated together in lysis buffer for 1 h at 55°C. DNA was extracted from filters using Qiagen DNeasy Blood & Tissue extraction kits (Qiagen) with the following modifications: 360 μl ATL buffer and 40 μl of Proteinase K were used for cell lysis and the volume of AL buffer and 95% ethanol was adjusted to 400 μl post-lysis. DNA was eluted into 200 μl of AE buffer and stored at −20°C until qPCR analysis. Negative DNA extraction controls (extraction buffers only) were included during the DNA extraction process to identify any contamination of equipment and reagents that may have occurred during this procedure. 20230111 All DNA extracts were tested first for the presence of PCR inhibitors by performing an internal positive control (IPC) assay using TaqMan Exogenous Internal Positive Control Reagents (EXO-IPC) (Applied Biosystems). Each DNA sample was run in triplicate, and each IPC assay was performed in 12 μl volumes consisting of 6 μl of TaqPath ProAmp Master Mix (Thermo Fisher Scientific), 1 μl EXO-IPC mix, 0.2 μl EXO-IPC DNA, 1.8 μl Nanopure sterile H2O, and 3 μl DNA template or sterile water for the nontemplate control (NTC) using the default cycle parameters with 40 cycles. Environmental samples were considered inhibited when samples displayed a >2 cyclethreshold (Ct) shift relative to the mean NTC. Samples that were inhibited were treated with OneStep PCR Inhibitor Removal Kit (Zymo Research Corporation) and retested with the IPC assay to confirm that PCR inhibition was alleviated. 20230118 We tested the DNA extracts for the presence of Chinook and coho salmon using the qPCR assays developed in Duda et al. (2020) and for sockeye salmon using the qPCR assay developed in Tillotsen et al. (2018). All assays were performed in triplicate on each DNA extract in 12 μl reaction volumes consisting of 3 μl DNA template and 6 μl TaqPath ProAmp Mastermix. Primer and probe concentrations for each assay were based on the methods reported in the respective publications. PCRs were run on the ViiA 7 Real-Time PCR system (Applied Biosystems) with cycling parameters consisting of an initial step of 2 min at 50°C, then 10 min at 95°C, followed by 45 cycles of denaturing at 95°C for 15 s and annealing/ extension at 60°C for 1 min. Results were analyzed using ViiA 7 RUO 1.2.4 software (Applied Biosystems). To estimate DNA concentrations, we included a five-point dilution (10,000, 2,000, 400, 80 and 16 copies per reaction) of a gBlock double-stranded DNA fragment (Integrated DNA Technologies) representing the target amplicons on each PCR run and each dilution was run in quadruplicate. Negative controls consisting of DNA extraction controls and no-template PCR controls were included and each were run with 3 replicates. All negative controls yielded no positive detections. We considered a positive detection as any sample amplifying at less than 40 Ct with a uniform curve morphology, as suggested by Klymus et al. (2019).   References Duda JJ, Hoy MS, Chase DM, Pess GR, Brenkman SJ, McHenry MM, Ostberg CO. Environmental DNA is an effective tool to track recolonizing migratory fish following large‐scale dam removal. Environmental DNA. 2021 Jan;3(1):121-41. Klymus KE, Merkes CM, Allison MJ, Goldberg CS, Helbing CC, Hunter ME, Jackson CA, Lance RF, Mangan AM, Monroe EM, Piaggio AJ. Reporting the limits of detection and quantification for environmental DNA assays. Environmental DNA. 2020 Jul;2(3):271-82. Tillotson MD, Kelly RP, Duda JJ, Hoy M, Kralj J, Quinn TP. Concentrations of environmental DNA (eDNA) reflect spawning salmon abundance at fine spatial and temporal scales. Biological Conservation. 2018 Apr 1;220:1-1. 20230118 Salmon temporal eDNA survey data from autosampler Comma Separated Value (CSV) file containing data. Producer Defined Sample name Unique identifier code for each eDNA sample collected from the Lake Washington Ship Canal for the temporal study based on the sequential order in which they were taken. Samples with a numerical sample name were collected using the autosampler. SBS samples were collected at the same location using a Smith-Root eDNA sampler to serve as controls. Producer Defined Sample name ranges from 132-373, and SBS1-SBS22. Date Date of sample collection. Producer Defined Date of sample collection. time of day Time of day of sample collection. Producer Defined Time of day of sample collection. water temperature (degrees C) Lake Washington Ship Canal subsurface water temperature at the time of sampling. Producer Defined NA No Data Producer defined 16.2 25.0 Celcius sample volume Volume of water filtered (L). Producer Defined 0.3 5.16 Liters CKCO3 qPCR Ct rep 1 A value that specifies the Ct of the1st PCR replicate of the CKCO3 assay. Producer Defined Undetermined No detectable qPCR amplification. Producer defined 35.871 41.653 CKCO3 qPCR Ct rep 2 A value that specifies the Ct of the 2nd PCR replicate of the CKCO3 assay. Producer Defined Undetermined No detectable qPCR amplification. Producer defined 34.74 42.535 CKCO3 qPCR Ct rep 3 A value that specifies the Ct of the 3rd PCR replicate of the CKCO3 assay. Producer Defined Undetermined No detectable qPCR amplification. Producer defined 34.954 44.009 CKCO3 qPCR assay y-intercept Y-intercept value of the qPCR standard curve for the CKCO3 assay. Producer Defined 40.995 41.243 CKCO3 qPCR assay R2 R2 value of the qPCR standard curve for the CKCO3 assay. Producer Defined 0.989 0.996 CKCO3 qPCR assay slope Slope value of the qPCR standard curve for the CKCO3 assay. Producer Defined -3.038 -2.988 CKCO3 qPCR assay % efficiency Percent efficiency value of the qPCR standard curve for the CKCO3 assay. Producer Defined 113.406 116.125 COCytb qPCR Ct rep 1 A value that specifies the Ct of the 1st PCR replicate of the COCytb assay. Producer Defined Undetermined No detectable qPCR amplification. Producer defined 36.535 42.181 COCytb qPCR Ct rep 2 A value that specifies the Ct of the 2nd PCR replicate of the COCytb assay. Producer Defined Undetermined No detectable qPCR amplification. Producer defined 37.292 42.32 COCytb qPCR Ct rep 3 A value that specifies the Ct of the 3rd PCR replicate of the COCytb assay. Producer Defined Undetermined No detectable qPCR amplification. Producer defined 37.108 44.961 COCytb qPCR assay y-intercept Y-intercept value of the qPCR standard curve for the COCytb assay. Producer Defined NA No Data Producer defined 42.117 43.006 COCYtb qPCR assay R2 R2 value of the qPCR standard curve for the COCytb assay. Producer Defined NA No Data Producer defined 0.99 0.992 COCytb qPCR assay slope Slope value of the qPCR standard curve for the COCytb assay. Producer Defined NA No Data Producer defined -3.158 -3.132 COCytb qPCR assay % efficiency Percent efficiency value of the qPCR standard curve for the COCytb assay. Producer Defined NA No Data Producer defined 107.351 108.566 SECO3 qPCR Ct rep 1 A value that specifies the Ct of the1st PCR replicate of the SECO3 assay. Producer Defined Undetermined No detectable qPCR amplification. Producer defined 37.277 41.526 SECO3 qPCR Ct rep 2 A value that specifies the Ct of the2nd PCR replicate of the SECO3 assay. Producer Defined Undetermined No detectable qPCR amplification. Producer defined 36.954 41.031 SECO3 qPCR Ct rep 3 A value that specifies the Ct of the 3rd PCR replicate of the SECO3 assay. Producer Defined Undetermined No detectable qPCR amplification. Producer defined 37.737 40.677 SECO3 qPCR assay y-intercept Y- intercept value of the qPCR standard curve for the SECO3 assay. Producer Defined 40.081 41.1 SECO3 qPCR assay R2 R2 value of the qPCR standard curve for the SECO3 assay. Producer Defined 0.963 0.992 SECO3 qPCR assay slope Slope value of the qPCR standard curve for the SECO3 assay. Producer Defined -3.166 -2.978 SECO3 qPCR assay % efficiency Percent efficiency value of the qPCR standard curve for the SECO3 assay. Producer Defined 106.926 116.646 eDNA carry over experiment.csv Comma Separated Value (CSV) file containing data. Producer Defined Sample Name Unique identifier code for each eDNA sample collected from the Lake Washington Ship Canal for the eDNA carry over study based on the sequential order in which they were taken. Producer Defined Values represent discrete sample identification codes. Date Date of sample collection. Producer Defined Date of sample collection. Sample type Identifies whether the filter samples was collected form the Lake Washington Ship Canal or from deionized water. Producer Defined Deionized Water A filter sample collected from deionized water. Producer defined Ship Canal A filter sample collected from the Lake Washington Ship Canal. Producer defined CKCO3 qPCR Ct rep 1 A value that specifies the Ct of the1st PCR replicate of the CKCO3 assay. Producer Defined Undetermined No detectable qPCR amplification. Producer defined 39.687 40.922 CKCO3 qPCR Ct rep 2 A value that specifies the Ct of the 2nd PCR replicate of the CKCO3 assay. Producer Defined Undetermined No detectable qPCR amplification. Producer defined 39.456 42.936 CKCO3 qPCR Ct rep 3 A value that specifies the Ct of the 3rd PCR replicate of the CKCO3 assay. Producer Defined Undetermined No detectable qPCR amplification. Producer defined 40.788 42.534 CKCO3 qPCR assay y-intercept Y-intercept value of the qPCR standard curve for the CKCO3 assay. Producer Defined 40.995 40.995 CKCO3 qPCR assay R2 R2 value of the qPCR standard curve for the CKCO3 assay. Producer Defined 0.989 0.989 CKCO3 qPCR assay slope Slope value of the qPCR standard curve for the CKCO3 assay. Producer Defined -2.988 -2.988 CKCO3 qPCR assay % efficiency Percent efficiency value of the qPCR standard curve for the CKCO3 assay. Producer Defined 116.125 116.125 COCytb qPCR Ct rep 1 A value that specifies the Ct of the 1st PCR replicate of the COCytb assay. Producer Defined Undetermined No detectable qPCR amplification. Producer defined 38.249 42.273 COCytb qPCR Ct rep 2 A value that specifies the Ct of the 2nd PCR replicate of the COCytb assay. Producer Defined Undetermined No detectable qPCR amplification. Producer defined 38.144 41.843 COCytb qPCR Ct rep 3 A value that specifies the Ct of the 3rd PCR replicate of the COCytb assay. Producer Defined Undetermined No detectable qPCR amplification. Producer defined 39.543 44.124 COCytb qPCR assay y-intercept Y-intercept value of the qPCR standard curve for the COCytb assay. Producer Defined 43.006 43.006 COCYtb qPCR assay R2 R2 value of the qPCR standard curve for the COCytb assay. Producer Defined 0.992 0.992 COCytb qPCR assay slope Slope value of the qPCR standard curve for the COCytb assay. Producer Defined -3.158 -3.158 COCytb qPCR assay % efficiency Percent efficiency value of the qPCR standard curve for the COCytb assay. Producer Defined 107.351 107.351 SECO3 qPCR Ct rep 1 A value that specifies the Ct of the1st PCR replicate of the SECO3 assay. Producer Defined Undetermined No detectable qPCR amplification. Producer defined SECO3 qPCR Ct rep 2 A value that specifies the Ct of the2nd PCR replicate of the SECO3 assay. Producer Defined Undetermined No detectable qPCR amplification. Producer defined SECO3 qPCR Ct rep 3 A value that specifies the Ct of the 3rd PCR replicate of the SECO3 assay. Producer Defined Undetermined No detectable qPCR amplification. Producer defined SECO3 qPCR assay y-intercept Y- intercept value of the qPCR standard curve for the SECO3 assay. Producer Defined 40.287 40.287 SECO3 qPCR assay R2 R2 value of the qPCR standard curve for the SECO3 assay. Producer Defined 0.963 0.963 SECO3 qPCR assay slope Slope value of the qPCR standard curve for the SECO3 assay. Producer Defined -2.978 -2.978 SECO3 qPCR assay % efficiency Percent efficiency value of the qPCR standard curve for the SECO3 assay. Producer Defined 116.646 116.646 GS ScienceBase U.S. Geological Survey mailing address

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Denver CO 80225 United States 1-888-275-8747 sciencebase@usgs.gov Unless otherwise stated, all data, metadata and related materials are considered to satisfy the quality standards relative to the purpose for which the data were collected. Although these data and associated metadata have been reviewed for accuracy and completeness and approved for release by the U.S. Geological Survey (USGS), no warranty expressed or implied is made regarding the display or utility of the data for other purposes, nor on all computer systems, nor shall the act of distribution constitute any such warranty. Digital Data https://doi.org/10.5066/xxxxxxxx None 20251203 Marshal Hoy USGS Western Fisheries Research Center mailing and physical

6505 NE 65TH ST

Seattle WA 98115 USA 206 526 6282 mhoy@usgs.gov FGDC Biological Data Profile of the Content Standard for Digital Geospatial Metadata FGDC-STD-001.1-1999