Developing and evaluating a point-of-use environmental DNA test for rapid field detection of highly invasive brown treesnakes
New terrestrial environmental DNA (eDNA) sampling techniques are sensitive tools for early detection of the highly invasive brown treesnakes (BTS). In a new collaborative research effort involving four USGS science centers, scientists have initiated the development of an innovative point-of-use (POU) assay for BTS. POU assays are tests that can be run in the field with minimal equipment and no expertise needed. These improvements overcome the issue of time lags associated with traditional eDNA assays and allow field biologists to assess the presence of BTS in real time.
About the invader
Brown treesnakes (BTS) are highly invasive snakes that were accidentally introduced onto the island of Guam after World War II. With no natural predators in Guam, these snakes have flourished and now occur at high densities. They threaten military infrastructure in Guam and pose threats to humans through their venomous bites and propensity to cause power outages. Further, since their introduction in Guam, BTS have either suppressed or caused the local extinction of most native forest birds on the island. Because Guam is a transportation hub for the Western Pacific, the accidental transport of BTS to other locations, such as Hawaii or the Northern Mariana Islands, is a significant and ongoing threat.
Detecting BTS using DNA left in the environment
While there are numerous ways to detect cryptic or invasive species, recent methods have focused on using DNA left behind in the environment (known as environmental DNA or eDNA) for such purposes. Most eDNA applications have been conducted in aquatic environments where a large water sample is taken and filtered to concentrate the DNA before testing. This line of research has expanded to include terrestrial environments, which poses additional challenges. On land, sampling consists of swabbing targeted terrestrial substrates, which typically produces less DNA than aquatic sampling. USGS researchers have developed and tested methods for sampling terrestrial eDNA and analyzing samples with a lab-based assay designed specifically to detect BTS DNA. While this assay holds much promise, a critical limitation in invasive species applications like early detection is the substantial amount of time required to process samples and get results back (on the order of weeks to months).
Photos: USGS researchers testing brown treesnake eDNA sampling methods. By Corinna Pinzari (USGS).
Point-of-use (POU) assay for BTS
POU assays are rapid, field-ready tests that field biologists can conduct with no expertise in laboratory work and without specialized equipment. These tests can be completed in minutes, overcoming the issue of time lags associated with traditional eDNA assays and allowing field biologists to assess the presence of BTS in real time.
Innovative combination of methods
The implementation of POU assays is relatively new for the detection of invasive species. Most current efforts make use of a method called Loop-mediated isothermal amplification (LAMP) for detection, yet recently CRISPR-Cas (short for clustered regularly interspaced short palindromic repeats-CRISPR-associated protein) technology has been developed as an alternative method. Combinations of the two methods (LAMP and CRISPR-Cas) have been explored to increase specificity and sensitivity of the POU assay. USGS researchers are testing both methods and a combined POU assay for BTS. Once developed and optimized, the efficacy of the POU assays will be compared with the standard lab-based assay to determine which is most likely to identify eDNA sampled from terrestrial environments.
Background image: Exposure with a UV-B light of CRISPR-Cas12a-processed samples after a reaction time of 30 minutes. Those that are illuminated are positive for brown treesnake DNA, those that are not illuminated contain negative controls (no brown treesnake DNA). Photo by Adam Perez (USGS).
Molecular Ecology Lab (MEL)
Invasive Species We Study: Brown Treesnake
Biology, Impacts and Control of Invasive Reptiles in the Pacific
New terrestrial environmental DNA (eDNA) sampling techniques are sensitive tools for early detection of the highly invasive brown treesnakes (BTS). In a new collaborative research effort involving four USGS science centers, scientists have initiated the development of an innovative point-of-use (POU) assay for BTS. POU assays are tests that can be run in the field with minimal equipment and no expertise needed. These improvements overcome the issue of time lags associated with traditional eDNA assays and allow field biologists to assess the presence of BTS in real time.
About the invader
Brown treesnakes (BTS) are highly invasive snakes that were accidentally introduced onto the island of Guam after World War II. With no natural predators in Guam, these snakes have flourished and now occur at high densities. They threaten military infrastructure in Guam and pose threats to humans through their venomous bites and propensity to cause power outages. Further, since their introduction in Guam, BTS have either suppressed or caused the local extinction of most native forest birds on the island. Because Guam is a transportation hub for the Western Pacific, the accidental transport of BTS to other locations, such as Hawaii or the Northern Mariana Islands, is a significant and ongoing threat.
Detecting BTS using DNA left in the environment
While there are numerous ways to detect cryptic or invasive species, recent methods have focused on using DNA left behind in the environment (known as environmental DNA or eDNA) for such purposes. Most eDNA applications have been conducted in aquatic environments where a large water sample is taken and filtered to concentrate the DNA before testing. This line of research has expanded to include terrestrial environments, which poses additional challenges. On land, sampling consists of swabbing targeted terrestrial substrates, which typically produces less DNA than aquatic sampling. USGS researchers have developed and tested methods for sampling terrestrial eDNA and analyzing samples with a lab-based assay designed specifically to detect BTS DNA. While this assay holds much promise, a critical limitation in invasive species applications like early detection is the substantial amount of time required to process samples and get results back (on the order of weeks to months).
Photos: USGS researchers testing brown treesnake eDNA sampling methods. By Corinna Pinzari (USGS).
Point-of-use (POU) assay for BTS
POU assays are rapid, field-ready tests that field biologists can conduct with no expertise in laboratory work and without specialized equipment. These tests can be completed in minutes, overcoming the issue of time lags associated with traditional eDNA assays and allowing field biologists to assess the presence of BTS in real time.
Innovative combination of methods
The implementation of POU assays is relatively new for the detection of invasive species. Most current efforts make use of a method called Loop-mediated isothermal amplification (LAMP) for detection, yet recently CRISPR-Cas (short for clustered regularly interspaced short palindromic repeats-CRISPR-associated protein) technology has been developed as an alternative method. Combinations of the two methods (LAMP and CRISPR-Cas) have been explored to increase specificity and sensitivity of the POU assay. USGS researchers are testing both methods and a combined POU assay for BTS. Once developed and optimized, the efficacy of the POU assays will be compared with the standard lab-based assay to determine which is most likely to identify eDNA sampled from terrestrial environments.
Background image: Exposure with a UV-B light of CRISPR-Cas12a-processed samples after a reaction time of 30 minutes. Those that are illuminated are positive for brown treesnake DNA, those that are not illuminated contain negative controls (no brown treesnake DNA). Photo by Adam Perez (USGS).