This project will develop a novel and practical nano-scale tracer for estimating advective transport of environmental DNA (eDNA), develop a related nano-scale method for sensitive multi-species detection of eDNA, and apply both methods to determine the spatio-temporal relationships between the occurrence and abundance of eDNA and biodiversity at two hierarchical levels: (1) community composition of fishes and water birds and (2) intra-specific (genetic) diversity/variation in one fish species. In doing so, the project will address important technological support for the maturation of eDNA science, fundamental research on eDNA transport, and the generation of accurate estimates of genetic diversity, and richness and abundance of species useful to management.

The project's nanotechnology methods are rooted in the point-of-care testing used in medical diagnostics. The specific nano-scale methods will be designed to a) serve as eDNA-like tracers to characterize hydrodynamic and associated transport-flow paths that are critical to interpreting eDNA data, and b) provide real-time, high sensitivity and low cost species detection and quantification in eDNA samples. As a proof of principle study, much of the project will be conducted in Cayuga Lake, New York because the fish and bird fauna are exceptionally well characterized, and because a verified, high resolution three dimensional hydrodynamic model for this lake can be used to reduce the uncertainty around the geographic origin and time of release of detected eDNA. To achieve our objectives, five tasks will be completed:

• Design and validate engineered eDNA-nanotags (eDNA-NTs) with similar transport characteristics to that of natural eDNA sources;
• Combine eDNA-NTs with a 3D-hydrodynamic model to spatially and temporally quantify the plume of detectable eDNA relative to the location and timing of the release of eDNA;
• Develop fluorescent barcodes, a novel nanotechnology method for multiplex eDNA detection that provides faster and cheaper results than existing methods;
• Apply the new nanotechnology method to study spatio-temporal variation in well-described fish and bird communities; and
• For a population of one fish species of management concern, use high throughput sequencing to estimate genetic structure and variability from eDNA samples for comparison with traditional methods for estimating genetic diversity and structure.

The novel methods developed to detect eDNA and model its location of origin within a waterbody will be generally applicable to freshwater and coastal water bodies including those of relevance to the Department of Defense’s mission. The eDNA-NTs would make possible much more rigorous studies of eDNA transport in a variety of habitats, the fluorescent barcodes will provide faster and cheaper results than existing detection methods, and the high-throughput sequencing of microsatellite eDNA will provide useful estimates of population size, allele frequencies and genetic differentiation of relevant species in conservation management.