title: Detecting an elusive invasive species: a diagnostic PCR to detect Burmese python in Florida waters and an assessment of persistence of environmental DNA.
abstract: Recent studies have demonstrated that detection of environmental DNA (eDNA) from aquatic vertebrates in water bodies is possible. The Burmese python, Python bivittatus, is a semi-aquatic, invasive species in Florida where its elusive nature and cryptic coloration make its detection difficult. Our goal was to develop a diagnostic PCR to detect P. bivittatus from water-borne eDNA, which could assist managers in monitoring this invasive species. First, we used captive P. bivittatus to determine whether reptilian DNA could be isolated and amplified from water samples. We also evaluated the efficacy of two DNA isolation methods and two DNA extraction kits commonly used in eDNA preparation. A fragment of the mitochondrial cytochrome b gene from P. bivittatus was detected in all water samples isolated with the sodium acetate precipitate and the QIAamp DNA Micro Kit. Next, we designed P. bivittatus-specific primers and assessed the degradation rate of eDNA in water. Our primers did not amplify DNA from closely related species, and we found that P. bivittatus DNA was consistently detectable up to 96 h. Finally, we sampled water from six field sites in south Florida. Samples from five sites, where P. bivittatus has been observed, tested positive for eDNA. The final site was negative and had no prior documented evidence of P. bivittatus. This study shows P. bivittatus eDNA can be isolated from water samples; thus, this method is a new and promising technique for the management of invasive reptiles.
authors.affiliation: USDA, Wildlife Services, National Wildlife Research Center, Wildlife Genetics Lab, 4101 LaPorte Avenue, Fort Collins, CO, 80521, USA.
authors.forename: Antoinette J
authors.initials: AJ
authors.lastname: Piaggio
authors.forename: Richard M
authors.initials: RM
authors.lastname: Engeman
authors.forename: Matthew W
authors.initials: MW
authors.lastname: Hopken
authors.forename: John S
authors.initials: JS
authors.lastname: Humphrey
authors.forename: Kandy L
authors.initials: KL
authors.lastname: Keacher
authors.forename: William E
authors.initials: WE
authors.lastname: Bruce
authors.forename: Michael L
authors.initials: ML
authors.lastname: Avery
mesh_terms: D000818:Animals; D017821:Boidae / Q000145:classification* / Q000235:genetics / Q000254:growth & development*; D045303:Cytochromes b / Q000235:genetics; D004247:DNA / Q000235:genetics / Q000302:isolation & purification*; D017931:DNA Primers / Q000235:genetics; D004272:DNA, Mitochondrial / Q000235:genetics; D005431:Florida; D058865:Introduced Species; D016133:Polymerase Chain Reaction / Q000379:methods*; D014867:Water / Q000737:chemistry
chemical_list: D017931:DNA Primers; D004272:DNA, Mitochondrial; D014867:Water; D004247:DNA; D045303:Cytochromes b
keywords: Burmese python; DNA persistence; PCR; Python bivittatus; environmental DNA; reptile