by Sanjana Kulkarni
figures by Swathy Karamchedu

Forensic DNA testing has become crucial in criminal investigations and legal proceedings. DNA has linked people to crime scenes using hair or blood and exonerated wrongfully convicted individuals. This type of DNA is called environmental DNA (eDNA) because it is collected from the environment, rather than from a person. Scientists have also begun analyzing eDNA from non-human organisms. All organisms are constantly shedding bits of themselves into the environment, and the DNA extracted from these materials is helping scientists identify organisms that live or lived in an environment even if the organism itself cannot be seen. eDNA technology is advancing public health surveillance and environmental conservation by allowing scientists to gather much more information about the natural world.

How is eDNA Identified/Sequenced?

Aided by the falling cost of DNA sequencing and better methods to collect and analyze DNA, scientists today can learn more about the natural world from DNA remnants than ever before. Because environmental samples contain DNA from many organisms, scientists use unique species identifiers, called barcode sequences, to identify the individual species. Barcode sequences are like human fingerprints because every species has a unique one. Scientists compare the barcodes in a sample to known barcodes in a database of all species. Any novel barcode sequence is placed near its closest relative, creating a family tree of barcode sequences that matches the family tree of organisms (Figure 1).

Figure 1. Sequencing eDNA from a pool of samples: A single environmental sample (e.g. soil or water) contains DNA from many species. Scientists use unique DNA barcodes to identify the species found in the sample. This technique is called metabarcoding because there are many different species’ barcodes present in the sample (in DNA-based analyses, the prefix “meta” denotes analyses done across species, as opposed to on a single one).

Applications of eDNA Technology

Disease Surveillance

Wastewater has long been tested for the DNA (or RNA, a related compound) of disease-causing microorganisms called pathogens that spread via contaminated food and water. The COVID-19 pandemic galvanized interest in wastewater testing more broadly as many other pathogens can be found in wastewater. Wastewater surveillance is an important tool for monitoring the spread of asymptomatic (infections without symptoms) and presymptomatic (infections before symptoms develop) COVID-19 cases, which are responsible for much of the virus’ spread. When public health officials are alerted of higher-than-normal levels of viral genetic material in wastewater, they can respond efficiently and proactively with targeted testing, contact tracing, and localized containment measures (Figure 2).

Figure 2. Overview of how public health officials can use wastewater testing to reduce outbreak spread: Continuous wastewater testing provides a baseline of the pathogens that are present in a community. Deviations from this baseline (e.g. a new pathogen begins circulating) alert public health officials to begin proactive measures like containment and testing individual people for the disease. This can then limit the spread of the disease, reducing the number of people who fall ill and become hospitalized.

The COVID-19 pandemic provided the impetus to set up additional large-scale wastewater disease surveillance systems for other pathogens like hepatitis A virus and norovirus. Many pathogens are always circulating, so early outbreak detection systems can help public health officials respond swiftly when an outbreak occurs to avoid overburdening hospitals. Future disease tracking will benefit from such expanded surveillance systems.

Cataloging Species from Different Habitats

To aid in conservation efforts, scientists catalog species from various habitats to identify those that are present. Traditional methods of cataloging species, such as visual surveys and trapping, are stressful on animals, time- and labor-intensive, and can miss very rare or very small species. Collecting eDNA samples is a cheap, non-invasive alternative that is safer for both wildlife and people. Collection methods are sensitive enough to capture DNA from soil, water, air, and snow (Figure 3). By gathering DNA from a habitat, scientists can rapidly identify the plant, animal, and microbial species that live there without needing to see and count them individually.

Figure 3. eDNA can be collected from many different sources: Soil and underground rocks can preserve DNA for thousands of years, so they are particularly good for studying natural history. DNA collected from bodies of water provides a snapshot of the organisms that currently live in those habitats. Air sensors can collect information about terrestrial habitats (e.g. dense forests) more efficiently than visual surveys.

The oceans are difficult to explore because of high water pressure, lack of oxygen, and low temperatures. eDNA can be collected from seawater with remotely controlled equipment, alleviating many of these difficulties. Even so, collecting this DNA is difficult because of the vastness of the ocean. Scientists have been helped by an unlikely source: sponges. Sponges gather food (microorganisms and other debris) by filtering it from the water. As they filter out their food, eDNA from the water is also captured and concentrated in the body of the sponge. Scientists sampled pieces of sponge tissue and were able to find DNA from dozens of fish species. Even in the Antarctic, where biodiversity is lower than in waters closer to the equator, DNA was found from penguins, seals, and sea stars.

eDNA collection from the air began by chance when air pollution sensors trapped DNA from various birds, trees, and fungi. These sensors can easily be adapted to also monitor wildlife through their DNA, making it a promising technique that can be rapidly deployed for environmental conservation. Air filters are stored long after they have been removed from the sensors. One study retrospectively analyzed 30 years of stored filters designed to measure nuclear fallout and found enough DNA to measure changes in species diversity over time.

DNA typically degrades very quickly in the environment, but if preserved properly, it can be intact enough for scientists to sequence. Ideal sites for preserving DNA have low temperatures, little sunlight (because ultraviolet rays break down DNA), and are shielded from major weather events. Researchers have found plant and animal DNA from thousands of years ago in cave stalagmites and Siberian permafrost.

eDNA has given scientists a better idea of what ancient habitats looked like by supplementing information from fossils. Only a tiny fraction of all the organisms that ever lived became fossils, and it is very likely that some species, such as soft-bodied invertebrates, never left behind any. A 2022 study of eDNA collected from frozen soil showed that 2 million years ago, northern Greenland was full of plant and animal life. Researchers also found DNA from mastodons, animals related to mammoths, providing the first non-fossil evidence of them.

Challenges and Looking Forward

eDNA technology can be likened to a medical test, with similar development challenges. eDNA tests must be rigorously validated and standardized so everyone agrees on the tests’ ability to detect different species. If eDNA that matches a particular species is present, how confident are we that that species is actually present? Could it actually have come from another similar species? It is also not straightforward to infer species abundance from the amount of DNA in a sample. Some animals shed more DNA than others, so more DNA does not necessarily mean more individuals. Once DNA is collected, it needs to be matched against a database to identify the source organism, which requires comprehensive, well-curated databases.

But these challenges of a new growing field can be overcome with sufficient time, investment, and research. Aided by advances in computational methods and electronic data storage, eDNA technology is already making strides in disease surveillance for public health and monitoring plant and animal populations amidst great biodiversity loss. eDNA collection is both cost-effective and scalable, and in some cases, it can be added onto existing environmental sensors. Insights from DNA will allow us to remotely monitor both environmental and human health and peer into the natural history of the Earth.

Sanjana Kulkarni is a 3rd-year PhD student studying antibiotic resistance in tuberculosis.

Swathy Karamchedu is a graduate student in the Media, Medicine, and Health program at Harvard Medical School who is developing visual narratives for sleep health education.

Cover image by RoadLight from pixabay.

For more information:

  • To learn about how scientists are using eDNA to track invasive species, check out these articles about European green crabs in Washington and invasive ants in Australia. 
  • Scientists are analyzing rodent feces to understand how climate and environmental changes affect plant pathogens. 
  • Wastewater and sewage are also being tested for antibiotic resistance, which can spread easily and quickly between different bacteria when water sources are mixed. 
  • The U.S. Centers for Disease Control and Prevention (CDC) maintains interactive dashboards of COVID-19 detection in wastewater.
  • Ancient DNA is starting to be incorporated into archaeological studies, though not without significant debate about its merits.

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