Today scientists, researchers, consultants, and other professionals can take advantage of a simple truth to gather data and make more informed decisions. That truth: as living organisms move through their environment, they shed genetic material in the form of DNA. This material lingers, providing insight into the past and present of the creature that left it behind. The DNA that is found in the environment is called “environmental DNA” or eDNA (e-DNA) for short, and it is formally defined as “genetic material obtained directly from environmental samples without any obvious signs of biological source material” (Thomsen and Willerslev, 2015).
Think about it:
A simple sneeze is a shower of genetic material into the surrounding environment. Bodily functions are not the only way that genetic information is released into the wider world. As an organism moves through its environment it leaves behind a treasure trove of data waiting to be unlocked and harnessed. Recently, eDNA has been harnessed to detect rare or invasive species and pathogens in a broad range of environments. eDNA is improving accuracy, reducing cost, and streamlining work-flows across many fields and industries including paleontology, agriculture, public health, and ecology. (Collins et al., 2015; Jerde et al., 2011)
The ability to rapidly and sensitively detect the presence of a target species through eDNA analysis has enabled a wide range of scientific discoveries and technical advancements. For example, isolation of eDNA from ice cores revealed that Greenland was forested almost 2 million years more recently than previously estimated (Thomsen and Willerslev, 2015; Willerslev et al., 2007). Analysis of Mitochondrial DNA (mtDNA) targets in surface water enabled researchers to distinguish the source of fecal contamination (Martellini et al., 2005). Environmental testing of cooling towers and other water systems enabled more sensitive detection of Legionella, a pathogen that can cause severe illness in the elderly (Collins et al., 2015).
eDNA can be analyzed via the following steps:
Samples are typically collected in the form of water, soil, sediment, or surface swabs. The DNA must then be extracted and purified to remove chemicals such as humic acid that are abundant in soil and sediment and strongly inhibit the PCR reaction. The final step, detection via qPCR, relies on selection of a suitable eDNA target. The ideal eDNA qPCR target is species specific and highly abundant. Mitochondrial DNA (mtDNA) is a popular target as it checks both of these boxes: mtDNA has significant divergence across species and there are thousands of copies of mtDNA per cell. The target sequence is then detected via quantitative polymerase chain reaction (qPCR). In this process, billions of copies of a target sequence are synthesized from template DNA (the purified eDNA sample, which can be present at very low levels) and then detected in real-time via fluorescent signal amplification. At the end of the reaction, if significant amplification of fluorescent signal is detected, the environmental sample is considered positive for the species of interest.
Information is power. eDNA detection has several advantages over traditional methods of species detection. For example, traditional methods in aquatic ecology (the study of ecosystem relationships in aquatic environments) involve observation and/or trapping of the species of interest combined with extensive documentation. This can be difficult and labor intensive, especially if the species of interest is rare or very small. One study found that it took 93 days of human effort to identify a single example of a rare species of fish using electrofishing, whereas it took only 4 hours to detect the species using eDNA (Jerde et al., 2011). eDNA detection has also been shown to be more sensitive than traditional methods such as casting-nets or fishing (Takahara, et al., 2013), which offers a critical advantage in the early detection of invasive species and monitoring of endangered species.
It gets better:
Determining the presence or absence of a target organism is only one of the benefits of eDNA. The “q” in real-time qPCR stands for “quantitative” and, therefore, this technique can also be used to determine the prevalence of a species in the environment. Without having to trap or visually count an animal – conservationists can determine if an endangered species’ population is declining. By measuring the relative quantity of DNA, a fish and wildlife agent can determine if an invasive species population is growing or dwindling.
Environmental DNA techniques can also be used to improve pathogen detection over traditional methods. In public health settings, pathogen detection often involves first growing (or culturing) the organism of interest, which is labor-intensive, and introduces the risk of pathogen spread. Conventional culture-based quantification methods also require transport to a centralized laboratory containing specialized equipment, highly trained lab technicians, which increases their cost and limits their viability for routine monitoring on site and especially in low-resource settings. Additionally, these methods require an overnight incubation step, which creates a significant lag in reporting results.
Other culture based methods have been developed in recent years, including TECTA [Bramburger 2015] and CBTs (Compartment Bag Tests; Brooks 2017; Wang 2017). However, they still require processing times of at least 4 hours and up to 48 hours to obtain results. Additionally, only a limited subset (e.g. CBTs) are appropriate for on-site deployment without access to a centralized laboratory. By directly analyzing the DNA in the environment, accurate results can be achieved in less than 24 hours.
While there are many advantages to eDNA testing, there are a few important considerations when working with eDNA. Different organisms, and even different life stages of a given organism, shed DNA at different rates. Additionally, the rate of DNA degradation is highly dependent on environmental variables such as temperature and microbial activity. These factors can make it difficult to accurately estimate species density and how recently a species was present in the environment. However, careful investigation of environment-specific factors that influence the degradation rate and lab-based studies to quantify rates of DNA shedding can overcome these challenges.
Many of the advancements enabled by eDNA analysis have been in aquatic ecology. For example, Asian carp are invading Chicago’s waterways. Asian carp has the potential to severely disrupt the Great Lakes’ $7-billion fishing industry.
Environmental DNA is also used by the U.S. Department of the Interior to monitor water for Coliforms, Escherichia coli, and enterococci, all of which are used as indicators of contamination. The agency has found that molecular methods, like eDNA testing, can overcome issues associated with pathogen detection in water: low pathogen densities, large volumes of water, detection of multiple organisms at the same time, and the detection of live organisms (Keele, J., 2016).
More recently, the Wildlife Conservation Society (WCS) used eDNA to find the world’s rarest turtle, Rafetus swinhoei. Check out the video below where the team takes you through their process, results, and what they have planned next!
Another example includes Ruth Richardson, Associate Professor of Civil and Environmental Engineering at Cornell, working with her team to keep beaches open in New York state parks by testing for fecal indicator bacteria.
Lastly, see how the Norwegian Veterinary Institute is fighting the Crayfish Plague using eDNA!
Thanks to recent developments in molecular biology, it has become possible to test for environmental DNA in the field in less than an hour. This opens up a host of new applications. For example, eDNA can be used to monitor recreational waters like lakes, ponds, pools, and rivers for biological contaminants. In Michigan, organizations are using eDNA testing techniques to monitor lakes for the presence of the parasite that causes Swimmer’s Itch. The parasite, called a schistosome, burrows into the skin producing an itchy rash. The ability to detect schistosomes quickly allows for not only effective monitoring, but also quick remediation. For communities and businesses that depend on clean safe water, this kind of technology is a welcome addition to their portfolio of water management tools.
Until recently, the ability to perform analysis of bacteria and viruses in water has been limited to highly trained personnel located in specialized laboratories. Biomeme’s innovation is to put the power of biological water analysis into the hands of the common person and decision makers on the ground, enabling them to make evaluations in real time and at the point of need. Such an innovation will fundamentally change the way we monitor and report water quality.
Biomeme has created a complete end-to-end platform for eDNA testing. This system can be operated by almost anyone, is field portable, and has a long shelf-life without refrigeration. It is a qPCR testing platform consisting of three major parts:
Part 1: A DNA extraction kit, the M1 Sample Prep, has been shown to efficiently extract DNA from environmental water in a few minutes, can be used in low resource settings, and is so easy to use a seven year old can do it.
Part 2: A handheld, smartphone-operated real-time qPCR thermocycler that is as accurate as lab-bound devices. Biomeme’s two3 and three9 systems are ideal for meeting the challenges of environmental eDNA testing.
Part 3: The final component of this end-to-end platform are the Go-Strips. These contain all the necessary reagents to perform a real-time qPCR assay. In addition, Go-Strips come in a lyophilized (freeze-dried) format that is excellent for field use because there is no need for cold chain (carrying around coolers of dry ice in which to store the test chemistries).
Biggs et al. (2015). Using eDNA to develop a national citizen science-based monitoring programme for the great crested newt (Triturus cristatus). Biological Conservation.
Collins, et al. (2015). Real-time PCR to supplement gold-standard culture-based detection of Legionella in environmental samples. Journal of Applied Microbiology.
Dejean et al. (2012). Improved detection of an alien invasive species through environmental DNA barcoding: the example of the American bullfrog Lithobates catesbeianus. Journal of Applied Ecology.
Goldberg et al. (2013). Environmental DNA as a new method for early detection of New Zealand mudsnails (Potamopyrgus antipodarum). Freshwater Science.
Jerde et al. (2011). “Sight-unseen” detection of rare aquatic species using environmental DNA: eDNA surveillance of rare aquatic species. Conservation Letters.
Keele, J. (2016). Using eDNA to test for pathogens in reused water. U.S. Department of the Interior Bureau of Reclamation Research and Development Office.
Martinelli et al. (2005). Use of eukaryotic mitochondrial DNA to differentiate human, bovine, porcine, and ovine sources in fecally contaminated surface water. Water Research.
Olson et al. (2013). An eDNA approach to detect eastern hellbenders (Cryptobranchus a. alleganiensis) using samples of water. Wildlife Research.
Sigsgaard et al. (2015). Monitoring the near-extinct European weather loach in Denmark based on environmental DNA from water samples. Biological Conservation.
Takahara et al. (2013). Using environmental DNA to estimate the distribution of an invasive fish species in ponds. PLoS ONE.
Thomsen, P.F. and Willerslev, E. (2015). Environmental DNA – An emerging tool in conservation for monitoring past and present biodiversity. Biological Conservation.
Willerslev, E. et al. (2007). Ancient biomolecules from deep ice cores reveal a forested Southern Greenland. Science.