Burmese Python Environmental DNA (eDNA) Surveys in the Stormwater Treatment Areas and Corkscrew Regional Ecosystem Watershed
USGS scientists have developed and optimized python-specific eDNA markers to detect Burmese pythons from water samples taken from the Florida Everglades. This development may aid in the identification of possible waterways used as corridors for northward expansion of this invasive species.
The Science Issue and Relevance: The interconnected web of canals and river systems in southern Florida are known to serve as a conduit for the spread of non-native and invasive aquatic and semi-aquatic species, including the Burmese python (Python bivittatus). This network of waterways makes it difficult to determine the distribution of Burmese pythons across the region. Environmental DNA (eDNA) techniques are one way to help improve our understanding of python distribution and range delimitation. Though eDNA sampling design takes into account optimal sample numbers, densities, and locations, without proper statistical power, false negative eDNA detections can occur. Sampling design also needs to consider the species densities and the effects of different types of habitats and land use patterns on those densities. As compared to other aquatic species (e.g., fish, amphibians), detection of eDNA from semi-aquatic reptiles is challenging due to the limited amounts of DNA shed directly into waterbodies. Also, the low densities of pythons often result in lower eDNA concentrations. Additional challenges encountered in southern Florida include large amounts of suspended sediment, the presence of high microbial communities, and environmental inhibitors in water samples. For these reasons, detection of pythons in new areas can be challenging and at times limit the ability to derive conclusive information from negative results. Adaptive sampling design and calculations of occurrence and detection estimates can improve the confidence in eDNA detections and non-detections. This project conducted Burmese python eDNA sampling surveys in Stormwater Treatment Areas (STAs), Corkscrew Regional Ecosystem Watershed (CREW), and adjacent river or canal systems connected to previously sampled southern Florida waterways to fill knowledge gaps of Burmese python occupation in south Florida. Occurrence estimates of Burmese pythons in the region were estimated and helped to identify possible waterways used as corridors for northward expansion.
Methodology for Addressing the Issue: Water samples were collected throughout five Stormwater Treatment Areas in southern Florida in canals (38 sampling sites) and lakes, ponds, or reservoirs (sampling sites). Water filtration concentrated the eDNA, which was then isolated using highly optimized procedures to increase eDNA yield and reduce environmental inhibitors such as tannins or humic acid, compounds characteristic of tea-colored waters. To detect Burmese pythons, three species-specific markers (two primers and a fluorescently labelled probe) were developed and optimized for use on a droplet digital polymerase chain reaction (ddPCR) platform. Compared to quantitative PCR (qPCR) platforms, the ddPCR platform has higher accuracy and precision and can detect a single molecule of DNA from an environmental sample.
Future Steps: Due to the potentially low densities of expanding Burmese python populations and low snake eDNA concentrations, data from previous years will be used to help to refine the sampling methods for improved eDNA detection. Additionally, it can be logistically difficult to obtain accurate sampling of the waterbodies in the region and data collection will likely vary by habitat type. Accurate estimation of Burmese python range delimitation may require additional sampling and/or alternative sampling locations throughout the study area. The limited water flow in these areas can also concentrate inhibitors, which resulted in many of the samples having inhibitors even after the extra removal processes. Ongoing efforts to improve PCR inhibition removal are continuously underway to improve data analysis.
Priority Landscapes: Greater Everglades
Efficacy of eDNA as an Early Detection and Rapid Response Indicator for Burmese Pythons in the Northern Greater Everglades Ecosystem
Genetic Analysis of the Invasive Burmese Python to Aid Management and Population-Control Decision-Making
Using Environmental DNA for Burmese Python Detection Probabilities and Range-Delimitation in Southern Florida
Ecology of and Control Strategies for Invasive Burmese Pythons (Python molurus bivitattus) in the Greater Everglades
USGS scientists have developed and optimized python-specific eDNA markers to detect Burmese pythons from water samples taken from the Florida Everglades. This development may aid in the identification of possible waterways used as corridors for northward expansion of this invasive species.
The Science Issue and Relevance: The interconnected web of canals and river systems in southern Florida are known to serve as a conduit for the spread of non-native and invasive aquatic and semi-aquatic species, including the Burmese python (Python bivittatus). This network of waterways makes it difficult to determine the distribution of Burmese pythons across the region. Environmental DNA (eDNA) techniques are one way to help improve our understanding of python distribution and range delimitation. Though eDNA sampling design takes into account optimal sample numbers, densities, and locations, without proper statistical power, false negative eDNA detections can occur. Sampling design also needs to consider the species densities and the effects of different types of habitats and land use patterns on those densities. As compared to other aquatic species (e.g., fish, amphibians), detection of eDNA from semi-aquatic reptiles is challenging due to the limited amounts of DNA shed directly into waterbodies. Also, the low densities of pythons often result in lower eDNA concentrations. Additional challenges encountered in southern Florida include large amounts of suspended sediment, the presence of high microbial communities, and environmental inhibitors in water samples. For these reasons, detection of pythons in new areas can be challenging and at times limit the ability to derive conclusive information from negative results. Adaptive sampling design and calculations of occurrence and detection estimates can improve the confidence in eDNA detections and non-detections. This project conducted Burmese python eDNA sampling surveys in Stormwater Treatment Areas (STAs), Corkscrew Regional Ecosystem Watershed (CREW), and adjacent river or canal systems connected to previously sampled southern Florida waterways to fill knowledge gaps of Burmese python occupation in south Florida. Occurrence estimates of Burmese pythons in the region were estimated and helped to identify possible waterways used as corridors for northward expansion.
Methodology for Addressing the Issue: Water samples were collected throughout five Stormwater Treatment Areas in southern Florida in canals (38 sampling sites) and lakes, ponds, or reservoirs (sampling sites). Water filtration concentrated the eDNA, which was then isolated using highly optimized procedures to increase eDNA yield and reduce environmental inhibitors such as tannins or humic acid, compounds characteristic of tea-colored waters. To detect Burmese pythons, three species-specific markers (two primers and a fluorescently labelled probe) were developed and optimized for use on a droplet digital polymerase chain reaction (ddPCR) platform. Compared to quantitative PCR (qPCR) platforms, the ddPCR platform has higher accuracy and precision and can detect a single molecule of DNA from an environmental sample.
Future Steps: Due to the potentially low densities of expanding Burmese python populations and low snake eDNA concentrations, data from previous years will be used to help to refine the sampling methods for improved eDNA detection. Additionally, it can be logistically difficult to obtain accurate sampling of the waterbodies in the region and data collection will likely vary by habitat type. Accurate estimation of Burmese python range delimitation may require additional sampling and/or alternative sampling locations throughout the study area. The limited water flow in these areas can also concentrate inhibitors, which resulted in many of the samples having inhibitors even after the extra removal processes. Ongoing efforts to improve PCR inhibition removal are continuously underway to improve data analysis.