eDNA barcoding: Using next-generation sequencing of environmental DNA for detection nad identification of cetacean species
Abstract
ABSTRACT We propose to develop next-generation sequencing methodology for detection and species identification of cetaceans using environmental (e)DNA collected from seawater. Referred to here as ‘(e)DNA barcoding’, this new methodology can be used to [a] detect and identify the presence of (non-vocal) cetaceans in the field and [b] identify the species of unknown cetacean vocalizations to improve the accuracy of ongoing U.S. Navy passive acoustic monitoring efforts. This development involves optimizing and modifying a number of standard laboratory protocols, as well as initial field-testing under a relatively controlled environment. We will recover (e)DNA from seawater using filtering and binding reagents and optimize the amplification of mitochondrial (mt)DNA for species identification using a suite of PCR primers designed from a comprehensive reference database of sequences from cetacean species (Baker et al. 2003). Nextgeneration sequencing of the pooled amplicons will provide 100,000s of short reads (up to 250 base pairs in length) from each (e)DNA sampling experiment, improving the reliability and sensitivity of species identification, even when multiple species are present. The unique amplicon sequences (haplotypes) can be matched back to the reference database or searched against GenBank for species identification. By using individually indexed sequencing adaptors, we can multiplex many experiments in a single next-generation sequencing run, greatly reducing the cost of species monitoring. In the first year, we will optimize laboratory methodology and conduct a series of (e)DNA sampling experiments in the vicinity of killer whales Orcinus orca near San Juan Island in Puget Sound. The regular habits of the southern residents in these semi-enclosed waters and their well characterized vocalizations will allow us to detect pods and sample (e)DNA at various distances and time periods. The known differences between (mt)DNA lineages of killer whales will be used to demonstrate the potential for identification of ecotypes, as well as species. In the second year, we will move to an open-ocean environment, supported by a fixed or towed acoustic detection array to locate cetaceans. The location of these open-ocean tests will be decided dependent on the (e)DNA thresholds estimated in the first year and in consultation with the Navy. If successful under open ocean conditions, routine (e)DNA barcoding could complement the interpretation of acoustic and visual surveys now routinely used to monitor cetacean habitat.
Document Details
- Document Type
- DoD Grant Award
- Publication Date
- Aug 12, 2016
- Source ID
- N000141512297
Entities
People
- Scott Baker
Organizations
- Office of Naval Research
- Oregon State University
- United States Navy