{
"USGS Publications Warehouse": {
"@context": "https://schema.org",
"@type": "Article",
"additionalType": "Journal Article",
"name": "Isothermal recombinant polymerase amplification and CRIPSR (CAS12A) assay detection of Renibacterium salmoninarum as an example for wildlife pathogen detection in environmental DNA samples",
"identifier": [
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"propertyID": "USGS Publications Warehouse IndexID",
"value": "70250151",
"url": "https://pubs.usgs.gov/publication/70250151"
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"value": 70250151
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"propertyID": "DOI",
"value": "10.7589/JWD-D-22-00128",
"url": "https://doi.org/10.7589/JWD-D-22-00128"
}
],
"journal": {
"@type": "Periodical",
"name": "Journal of Wildlife Diseases",
"volumeNumber": "59",
"issueNumber": "4"
},
"inLanguage": "en",
"isPartOf": [
{
"@type": "CreativeWorkSeries",
"name": "Journal of Wildlife Diseases"
}
],
"datePublished": "2023",
"dateModified": "2023-11-22",
"abstract": "Improving rapid detection methods for pathogens is important for research as we collectively aim to improve the health of ecosystems globally. In the northern hemisphere, the success of salmon (Oncorhynchus\u00a0spp.) populations is vitally important to the larger marine, aquatic, and terrestrial ecosystems they inhabit. This has led to managers cultivating salmon in hatcheries and aquaculture to bolster their populations, but young salmon face many challenges, including diseases such as bacterial kidney disease (BKD). Early detection of the BKD causative agent,\u00a0Renibacterium salmoninarum, is useful for managers to avoid outbreaks in hatcheries and aquaculture stocks to enable rapid treatment with targeted antibiotics. Isothermal amplification and CRIPSR-Cas12a systems may enable sensitive, relatively rapid, detection of target DNA molecules from environmental samples compared to quantitative PCR (qPCR) and culture methods. We used these technologies to develop a sensitive and specific rapid assay to detect\u00a0R. salmoninarum\u00a0from water samples using isothermal recombinase polymerase amplification (RPA) and an AsCas12a RNA-guided nuclease detection. The assay was specific to\u00a0R. salmoninarum\u00a0(0/10 co-occurring or closely related bacteria detected) and sensitive to 0.0128 pg/\u00b5L of DNA (approximately 20\u201340 copies/\u00b5L) within 10 min of Cas activity. This assay successfully detected\u00a0R. salmoninarum\u00a0environmental DNA in 14/20 water samples from hatcheries with known quantification for the pathogen via previous qPCR (70% of qPCR-positive samples). The RPA-CRISPR/AsCas12a assay had a limit of detection (LOD) of >10 copies/\u00b5L in the hatchery water samples and stochastic detection below 10 copies/\u00b5L, similar to but slightly higher than the qPCR assay. This LOD enables 37 C isothermal detection, potentially in the field, of biologically relevant levels of\u00a0R. salmoninarum\u00a0in water. Further research is needed to develop easy-to-use, cost-effective, sensitive RPA/CRISPR-AsCas12a assays for rapidly detecting low concentrations of wildlife pathogens in environmental samples.",
"description": "12 p.",
"publisher": {
"@type": "Organization",
"name": "Wildlife Disease Association"
},
"author": [
{
"@type": "Person",
"name": "D\u2019Agnese, Erin",
"givenName": "Erin",
"familyName": "D\u2019Agnese",
"affiliation": [
{
"@type": "Organization",
"name": "University of Washington, School of Marine and Environmental Affairs, 3737 Brooklyn Ave NE, Seattle, WA 98105, USA"
}
]
},
{
"@type": "Person",
"name": "Chase, Dorothy M.",
"givenName": "Dorothy M.",
"familyName": "Chase",
"identifier": {
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"propertyID": "ORCID",
"value": "0000-0002-7759-2687",
"url": "https://orcid.org/0000-0002-7759-2687"
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"affiliation": [
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"@type": "Organization",
"name": "Western Fisheries Research Center",
"url": "https://www.usgs.gov/centers/western-fisheries-research-center"
}
]
},
{
"@type": "Person",
"name": "Andruszkiewicz-Allan, Elizabeth",
"givenName": "Elizabeth",
"familyName": "Andruszkiewicz-Allan",
"affiliation": [
{
"@type": "Organization",
"name": "Wild EcoHealth LLC, Seattle, WA 98117"
}
]
}
],
"funder": [
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"@type": "Organization",
"name": "Western Fisheries Research Center",
"url": "https://www.usgs.gov/centers/western-fisheries-research-center"
}
]
}
}