Scott, Laura C. (ORCID: 0000-0003-0303-5340) Ahlstrom, Christina A. (ORCID: 0000-0001-5414-8076) Woksepp, Hanna (ORCID: 0000-0002-1707-2655) Bonnedahl, Jonas (ORCID: 0000-0002-3182-389X) McKeeman, Cherie M. (ORCID: 0000-0001-9868-2502) Miller, Mark E. (ORCID: 0000-0002-4691-7469) Shirokauer, Dave Ramey, Andrew M. (ORCID: 0000-0002-3601-8400) 20250508 tabular digital data Anchorage, Alaska U.S. Geological Survey, Alaska Science Center Suggested Citation: Scott, L.C., Ahlstrom, C.A., Woksepp, H., Bonnedahl, J., McKeeman, C.M., Miller, M.E., Shirokauer, D., Ramey, A.M., 2025, Data for characterization of antimicrobial resistance genes in surface waters in two national parks in Alaska: U.S. Geological Survey data release, https://doi.org/10.5066/P14BOXET https://doi.org/10.5066/P14BOXET This data set describes the assessment of surface waters in Denali National Park and Preserve and Wrangell St. Elias National Park and Preserve for antimicrobial resistance genes during four summer months (May - August). Data were collected to understand the prevalence and abundance of antimicrobial resistance genes in surface waters in national parks in Alaska. Data can be used for epidemiological estimations of antimicrobial resistance genes in the presented samples. 20240501 20240831 observed Complete None planned Denali National Park and Preserve and Wrangell St. Elias National Park and Preserve -155.3906 -137.1094 64.5484 58.6083 USGS Metadata Identifier USGS:68097850d4be02112351888b ISO 19115 Topic Category Biota Environment Health NASA GCMD Earth Science Keyword Thesaurus Bacteria/archaea Freshwater ecosystems Aquatic ecosystems Ecological dynamics Public health Environmental health factors USGS CSA Biocomplexity Thesaurus Disease reservoirs Disease detection Disease spread Disease control Antimicrobial activity USGS Thesaurus Health and disease Bacteria Environmental DNA NIH MeSH Thesaurus Drug resistance, microbial None Surface water Antimicrobial resistance USGS Geographic Names Information System (GNIS) Alaska USGS Protected Areas Database of the United States (PAD-US) Denali National Park and Preserve Wrangell-St. Elias National Park and Preserve No access constraints. No use constraints. These data are marked with a Creative Common CC0 1.0 Universal License and are in the public domain. It is requested that this USGS data release be cited for any subsequent publications that reference or utilize these data. Users are advised to read the dataset's metadata thoroughly to understand appropriate use and data limitations. U.S. Geological Survey, Alaska Science Center Mailing and Physical

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Anchorage Alaska 99508 USA 907-786-7000 gs-ak_asc_datamanagers@usgs.gov Quantitative PCR (qPCR) markers were checked using NCBI Blast and the Comprehensive Antibiotic Resistance Database to confirm primers amplified the correct target. Data have been checked for completeness. Attribute values fall within expected ranges. Null values are described in the Entities and Attributes section. Associated data represent values for only the surface waters tested at the date and time of collection. Quantitative PCR (qPCR): Each qPCR reaction was done in 20µl volumes with the following inclusions: 10µl of 2x PowerUp SYBR Mastermix (Applied Biosystems), 6µl of nanopure water, 2µl of template DNA, and 1µl of each forward and reverse primer. Final primer concentrations were 0.5µM and are listed for each target in as a table in this release. Thermal cycling conditions for the qPCR reaction were as follows: uracil-DNA glycosylase deactivation at 50°C for 2 minutes, polymerase activation at 95°C for 2 minutes, 40 cycles of 95°C for 15 seconds followed by 60°C for 30 seconds, and a melt curve analysis of 95°C for 1 minute, 60°C for 5 minutes, and gradual increase of temperature from 60°C to 95°C at 0.1°C/second. For every sample, every target was run in duplicate, and every plate included duplicate non-template control wells. Unknown Sampling locations are described without coordinates. Water body names are used to describe general sampling locations. Point 0.00001 0.00001 Decimal degrees World Geodetic System of 1984 (WGS84) World Geodetic System of 1984 (WGS84) 6378137 298.257223563 antimicrobialResistance_surfaceWater_nationalParksAK_2024_primersTemplates.csv Table includes genomic information regarding each target assessed with quantitative PCR in this study. Presented in Comma Separated Value (CSV) format. Author defined GeneID The name of each gene target assessed by qPCR in this study. Author defined Forty-one antimicrobial resistance gene targets were assessed in this study: aminoglycoside resistance genes (aac3-IV, aac(6’)-IIc, aacA43, APH, rmtB), beta-lactam resistance genes (blaCMY, blaGES, blaIMP, blaKPC, blaNDM, blaOXA, blaOXY, blaSHV, blaTEM, blaVIM, blaCTXM), class one integrons (dfrA1, int1), multidrug efflux pumps (qacEdelta, mepR, mexF), glycopeptide resistance genes (vanA, vanD), macrolide resistance genes (ereA, ermB, ermD, ermF), phenicol resistance genes (catII, catP), polymyxin resistance genes (mcr-1, mcr-3), quinolone resistance genes (oqxA, qnrB, qnrS), sulfonamide resistance genes (sul1, sul3), and tetracycline resistance genes (tetA, tetD, tetM, tetL, tetX). Specific taxonomic markers for E.coli and Salmonella were used. Microbial source trackers were also selected for humans (HF183), birds (GFD), and ruminants (Rum2Bac) and the universal bacteria marker (16S rRNA). Forward_Primer The oligonucleotide sequence used to match to the five-prime end of the gene target sequence. Author defined The oligonucleotide sequence used to match to the five-prime end of the gene target sequence. Reverse_Primer The oligonucleotide sequence used to match to the three-prime end of the gene target sequence. Author defined The oligonucleotide sequence used to match to the three-prime end of the gene target sequence. Template_Sequence The entire nucleotide sequence of the gene target expected to be amplified by the forward and reverse primers during qPCR. Author defined The entire nucleotide sequence of the gene target expected to be amplified by the forward and reverse primers during qPCR. antimicrobialResistance_surfaceWater_nationalParksAK_2024_sampleDescriptions.csv Table describing the collection date, time, location, and the volume of water filtered for each sample. Presented in Comma Separated Value (CSV) format. Author defined SampleID A unique alphanumeric code to identify each surface water sample. Author defined Codes were designated with the following scheme: national park starting letter and the sample site number (D1 = Denali National Park and Preserve sample site 1), the letter "R" and the field replicate number, and the letter "T" and the timepoint number of the sample. Grab samples from the Denali National Park wastewater treatment plant have the code "DWTTP" followed by a replicate number. The sample "DWTTPU" was taken from the Denali National Park wastewater treatment plant with an ultrafilter rather than a grab sample and has no replicate. Park The name of the national park where the surface water sampling occurred. Author defined Wrangell St. Elias National Park Wrangell St. Elias National Park and Preserve Author defined Denali National Park Denali National Park and Preserve Author defined Site_Number A number given to designate each sampling site within each national park. Author defined NA There is no site number for wastewater treatment plant samples. Author defined 1 6 integer Site_Name The common place name of the body of water where each sample was collected. Author defined Ruth Lake Ruth Lake, Wrangell St. Elias National Park (approximately 61.367N 143.447W). Author defined Swimming Hole Swimming Hole, Wrangell St. Elias National Park (approximately 61.435N 142.932W). Author defined Jumbo Creek Jumbo Creek, Wrangell St. Elias National Park (approximately 61.503N 142.898W). Author defined National Creek National Creek, Wrangell St. Elias National Park (approximately 61.484N 142.887W). Author defined Roadside Pond Roadside Pond, Denali National Park (approximately 63.701N 148.879W). Author defined Upper Savage River Upper Savage River, Denali National Park (approximately 63.711N 149.263W). Author defined Lower Savage River Lower Savage River, Denali National Park (approximately 63.739N 149.292W). Author defined Upper Rock Creek Upper Rock Creek, Denali National Park (approximately 63.724N 148.965W). Author defined Lower Rock Creek Lower Rock Creek, Denali National Park (approximately 63.722N 148.962W). Author defined Horseshoe Lake Horseshoe Lake, Denali National Park (approximately 63.740N 148.912W). Author defined Wastewater Treatment Plant Wastewater Treatment Plant, Denali National Park (approximately 63.732N 148.886W). Author defined Replicate The field replicate number for each surface water sample collected. Author defined 1 2 integer Timepoint The timepoint value when each surface water sample was collected. Samples were collected from the same locations at four timepoints throughout the summer months. Author defined NA Wastewater treatment samples were not taken at multiple timepoints. Author defined 1 4 integer Latitude The approximate latitude of the surface water sample collection location. Author defined 61.36515 63.74027 decimal degrees Longitude The approximate longitude of the surface water sample collection location. Author defined -149.29196 -142.8874 decimal degrees Volume_Filtered The volume of water filtered for each sample, measured in liters. Author defined NA For samples designated as "NA" the meter measuring sample flow malfunctioned and sampled volume could not be measured. Author defined 1.0 101.438 liters Sample_Date The date the surface water sample was collected (YYYY-MM-DD). Author defined 2024-05-28 2024-08-27 date (YYYY-MM-DD) antimicrobialResistance_surfaceWater_nationalParksAK_2024_cycleThresholdValues.csv Table of cycle threshold values from qPCR analysis for surface water samples collected in two national parks in Alaska at four defined timepoints. Presented as a Comma Separated Value (CSV) file. Author defined GeneID_Replicate The gene identification name that was tested, and the technical replicate number separated by an underscore. Author defined Forty-one antimicrobial resistance gene targets were assessed in this study: aminoglycoside resistance genes (aac3-IV, aac(6’)-IIc, aacA43, APH, rmtB), beta-lactam resistance genes (blaCMY, blaGES, blaIMP, blaKPC, blaNDM, blaOXA, blaOXY, blaSHV, blaTEM, blaVIM, blaCTXM), class one integrons (dfrA1, int1), multidrug efflux pumps (qacEdelta, mepR, mexF), glycopeptide resistance genes (vanA, vanD), macrolide resistance genes (ereA, ermB, ermD, ermF), phenicol resistance genes (catII, catP), polymyxin resistance genes (mcr-1, mcr-3), quinolone resistance genes (oqxA, qnrB, qnrS), sulfonamide resistance genes (sul1, sul3), and tetracycline resistance genes (tetA, tetD, tetM, tetL, tetX). Specific taxonomic markers for E.coli and Salmonella were used. Microbial source trackers were also selected for humans (HF183), birds (GFD), and ruminants (Rum2Bac) and the universal bacteria marker (16S rRNA). Non template controls (NTC) were assessed with every sample. columns "D1R1T1" through "DWTTPU" Cycle threshold values for antimicrobial resistance genes measured with qPCR for water samples with the code indicated in the column header. Author defined Numerical values indicate at which cycle the fluorescence crossed a pre-determined threshold for detection. Gene targets listed as "Undetermined" did not amplify across the threshold for positive detection. antimicrobialResistance_surfaceWater_nationalParksAK_2024_meltTemp.csv Table of melt temperatures for gene targets measured by qPCR from surface water samples collected from two national parks in Alaska at four defined timepoints. Presented in Comma Separated Value (CSV) format. Author defined GeneID_Replicate The name of the gene target measured in samples using qPCR, and the technical replicate separated by an underscore. Author defined Forty-one antimicrobial resistance gene targets were assessed in this study: aminoglycoside resistance genes (aac3-IV, aac(6’)-IIc, aacA43, APH, rmtB), beta-lactam resistance genes (blaCMY, blaGES, blaIMP, blaKPC, blaNDM, blaOXA, blaOXY, blaSHV, blaTEM, blaVIM, blaCTXM), class one integrons (dfrA1, int1), multidrug efflux pumps (qacEdelta, mepR, mexF), glycopeptide resistance genes (vanA, vanD), macrolide resistance genes (ereA, ermB, ermD, ermF), phenicol resistance genes (catII, catP), polymyxin resistance genes (mcr-1, mcr-3), quinolone resistance genes (oqxA, qnrB, qnrS), sulfonamide resistance genes (sul1, sul3), and tetracycline resistance genes (tetA, tetD, tetM, tetL, tetX). Specific taxonomic markers for E.coli and Salmonella were used. Microbial source trackers were also selected for humans (HF183), birds (GFD), and ruminants (Rum2Bac) and the universal bacteria marker (16S rRNA). Non template controls (NTC) were assessed with every sample. columns "D1R1T1_Tm1" through "DWTTPU_Tm2" Melt temperatures for antimicrobial resistance genes measured with qPCR for water samples with the code indicated by the column header. The most prominent melt temperature is designated by "Tm1" and the second most prominent with "Tm2". Blank cells indicate that no melting temperature was observed. Author defined Melt temperature equals the temperature, in degrees Celsius, at which the PCR amplicon dissociates. Multiple melting temperatures can be detected per sample, and the top two are recorded here as Tm1 and Tm2 for each replicate. Empty cells indicate that no melting temperature was able to be measured for that gene target, replicate combination. U.S. Geological Survey USGS ScienceBase Team Mailing and Physical

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Denver Colorado 80225 USA 1-888-275-8747 sciencebase@usgs.gov The U.S. Geological Survey, Alaska Science Center is the authoritative source of these data, distributed by ScienceBase (a USGS Trusted Digital Repository). 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, no warranty expressed or implied is made regarding the display or utility of the data for other purposes or on all computer systems, nor shall the act of distribution constitute any such warranty. Any use of trade, firm, or product names is for descriptive purposes only and does not imply endorsement by the U.S. Government. CSV Tabular data in CSV format; FGDC metadata in XML and HTML formats. https://doi.org/10.5066/P14BOXET None 20250508 U.S. Geological Survey, Alaska Science Center Mailing and Physical

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Anchorage Alaska 99508 USA 907-786-7000 gs-ak_asc_datamanagers@usgs.gov FGDC Biological Data Profile of the Content Standard for Digital Geospatial Metadata (CSDGM) FGDC-STD-001.1-1999