Environmental DNA (eDNA) sampling improves occurrence and detection estimates of invasive burmese pythons

Abstract

Environmental DNA (eDNA) methods are used to detect DNA that is shed into the aquatic environment by cryptic or low density species. Applied in eDNA studies, occupancy models can be used to estimate occurrence and detection probabilities and thereby account for imperfect detection. However, occupancy terminology has been applied inconsistently in eDNA studies, and many have calculated occurrence probabilities while not considering the effects of imperfect detection. Low detection of invasive giant constrictors using visual surveys and traps has hampered the estimation of occupancy and detection estimates needed for population management in southern Florida, USA. Giant constrictor snakes pose a threat to native species and the ecological restoration of the Florida Everglades. To assist with detection, we developed species-specific eDNA assays using quantitative PCR (qPCR) for the Burmese python (Python molurus bivittatus), Northern African python (P. sebae), boa constrictor (Boa constrictor), and the green (Eunectes murinus) and yellow anaconda (E. notaeus). Burmese pythons, Northern African pythons, and boa constrictors are established and reproducing, while the green and yellow anaconda have the potential to become established. We validated the python and boa constrictor assays using laboratory trials and tested all species in 21 field locations distributed in eight southern Florida regions. Burmese python eDNA was detected in 37 of 63 field sampling events; however, the other species were not detected. Although eDNA was heterogeneously distributed in the environment, occupancy models were able to provide the first estimates of detection probabilities, which were greater than 91%. Burmese python eDNA was detected along the leading northern edge of the known population boundary. The development of informative detection tools and eDNA occupancy models can improve conservation efforts in southern Florida and support more extensive studies of invasive constrictors. Generic sampling design and terminology are proposed to standardize and clarify interpretations of eDNA-based occupancy models.

Fig 1. Map of study area with environmental DNA sample collection locations (N = 21) and Burmese python occupancy.

Locations with present and absent environmental DNA detections are indicated with black and white dots, respectively. Some sample locations overlap due to the scale. BDB, Bird Drive Basin; DE, Deering Estates; ENP, Everglades National Park; HLWM, Holey Lands Wildlife Management Area; STA, Stormwater Treatment Area 5. Southwest Florida samples (SWFL) include three radiotagged Burmese pythons: ELV, Elvis; NOS, Noosa; and SWP, Sweet Pea.

Fig 2. Estimated concentration of Burmese python environmental DNA (mean and 95% confidence interval) per sample.

a. Higher concentration regions, y-axis scaled from 0 to 10 molecules/mL. b. Lower concentration regions, y-axis scaled from 0.0 to 1.4 molecules/mL. From each region, one to three locations were targeted, with triplicate field samples collected at each location. BDB, Bird Drive Basin; DE, Deering Estates; ENP, Everglades National Park; HLWM, Holey Lands Wildlife Management Area; SWP, Sweet Pea; NOS, Noosa; ELV, Elvis; STA, Stormwater Treatment Area 5.

Fig 3. The effect of sample size on estimates of the cumulative probability of occurrence of Burmese python environmental DNA (θ*) associated with samples taken from the four field locations analyzed by the three-level occupancy model.

BDB, Bird Drive Basin; DE, Deering Estates; ENP, Everglades National Park; STA, Stormwater Treatment Area 5.