What is Environmental DNA?
DNA, short for deoxyribonucleic acid, is the hereditary material in organisms that contains the biological instructions for building and maintaining them. The chemical structure of DNA is the same for all organisms, but differences exist in the order of the DNA building blocks, known as base pairs. Unique sequences of base pairs, particularly repeating patterns, provide a means to identify species, populations, and even individuals.
Environmental DNA (eDNA) is nuclear or mitochondrial DNA that is released from an organism into the environment. Sources of eDNA include secreted feces, mucous, and gametes; shed skin and hair; and carcasses. eDNA can be detected in cellular or extracellular (dissolved DNA) form.
In aquatic environments, eDNA is diluted and distributed by currents and other hydrological processes (fig. 1), but it only lasts about 7–21 days, depending on environmental conditions (Dejean and others, 2011). Exposure to UVB radiation, acidity, heat, and endo- and exonucleases can degrade eDNA.
Use of eDNA for Inventory and Monitoring
Improved Detection of Native Species
Protocols using eDNA may allow for rapid, cost-effective, and standardized collection of data about species distribution and relative abundance. For small, rare, secretive, and other species that are difficult to detect, eDNA provides an attractive alternative for aquatic inventory and monitoring programs. Increasing evidence demonstrates improved species detection and catch-per-unit effort compared with electrofishing, snorkeling, and other current field methods. Thus, detection of species using eDNA may improve biodiversity assessments and provide information about status, distribution, and habitat requirements for lesser-known species.
Early Detection of Invasive Species
eDNA may also be an effective tool for early detection of aquatic invasive species. Recent studies have focused on Asian carp (Jerde and others, 2011) and American bullfrogs (Dejean and others, 2012), but protocols are being developed for New Zealand mudsnails, zebra mussels, quagga mussels, and others. Application of eDNA methods for invasive species monitoring may include periodically collecting water samples and screening them for several invasive species at once. Boat-ballast water, a source of introduction for many invasive species including mollusks, also could be sampled. Some intensive eradication programs for invasive species fail when a few surviving individuals recolonize the ecosystem. eDNA methods may provide a means of confirming eradication of all invaders.
Developing eDNA Protocols for Species Monitoring
Primer and Probe Design
Conventional Polymerase Chain Reaction (PCR) methods can be used for analyzing eDNA, but they may cross-amplify and provide false-positive results. Quantitative PCR (qPCR) methods are preferable to conventional PCR because they are likely more sensitive. In qPCR, primers are used to amplify a region of DNA that is specific to a target organism, and a probe is used to provide additional specificity and quantitative information. Several eDNA primer and probe sequences for individual species have been published. Other primer/probe sets must be designed for new species or specific needs.
Laboratory Optimization
The primer and probe design must be tested to ensure that the qPCR reaction always results in a positive detection in the presence of target-organism DNA and that amplification of non-target DNA does not occur. DNA from the target species should be screened prior to analyzing environmental samples to ensure test sensitivity. Preferably, this will involve 10 or more samples collected across the range of the species where the test will be applied. Closely related, co-occurring species also should be screened to ensure specificity prior to analyzing environmental samples, preferably involving five or more samples of each.
DNA can be extracted from preserved tissue samples using readily available kits. It is critical that no cross contamination occurs between species during the tissue-storage or extraction processes.
