The unknown unknown: A framework for assessing environmental DNA assay specificity against unsampled taxa.

Taxon-specific quantitative PCR (qPCR) assays are commonly used for environmental DNA sampling-based inference of animal presence. These assays require thorough validation to ensure that amplification truly indicates detection of the target taxon, but a thorough validation is difficult when there are potentially many non-target taxa, some of which may have incomplete taxonomies. Here, we use a previously published, quantitative model of cross-amplification risk to describe a framework for assessing qPCR assay specificity when there is missing information and it is not possible to assess assay specificity for each individual non-target confamilial.

Broad-scale eDNA sampling for describing aquatic species distributions in running waters: Pacific lamprey Entosphenus tridentatus in the upper Snake River, USA.

One of the most fundamental yet challenging tasks for aquatic ecologists is to precisely delineate the range of species, particularly those that are broadly distributed, require specialized sampling methods, and may be simultaneously declining and increasing in different portions of their range. An exemplar is the Pacific lamprey Entosphenus tridentatus, a jawless anadromous fish of conservation concern that is actively managed in many coastal basins in western North America. To efficiently determine its distribution across the accessible 56,168 km of the upper Snake River basin in the north-western United States, we first delimited potential habitat by using predictions from a species distribution model based on conventionally collected historical data and from the distribution of a potential surrogate, Chinook salmon Oncorhynchus tshawytscha, which yielded a potential habitat network of 10,615 km.

eDNAssay: A machine learning tool that accurately predicts qPCR cross-amplification.

Environmental DNA (eDNA) sampling is a highly sensitive and cost-effective technique for wildlife monitoring, notably through the use of qPCR assays. However, it can be difficult to ensure assay specificity when many closely related species co-occur. In theory, specificity may be assessed in silico by determining whether assay oligonucleotides have enough base-pair mismatches with nontarget sequences to preclude amplification.

qPCR detection of Sturgeon chub (Macrhybopsis gelida) DNA in environmental samples.

The Sturgeon chub (Macrhybopsis gelida) is a cyprinid fish native to the Missouri and Mississippi River basins of the U.S. Suspected long-term declines in the size of its distribution have prompted a review of its conservation status by the U.

An improved environmental DNA assay for bull trout (Salvelinus confluentus) based on the ribosomal internal transcribed spacer I.

The majority of environmental DNA (eDNA) assays for vertebrate species are based on commonly analyzed regions of the mitochondrial genome. However, the high degree of mitochondrial similarity between two species of charr (Salvelinus spp.), southern Dolly Varden (S.

Repurposing environmental DNA samples-detecting the western pearlshell () as a proof of concept.

Information on the distribution of multiple species in a common landscape is fundamental to effective conservation and management. However, distribution data are expensive to obtain and often limited to high-profile species in a system. A recently developed technique, environmental DNA (eDNA) sampling, has been shown to be more sensitive than traditional detection methods for many aquatic species.