Can environmental DNA be used to detect first arrivals of the cane toad, Rhinella marina, into novel locations?

Eradicating invasive species is difficult, but success is more likely when populations are small after arrival. The cane toad, Rhinella marina, is an invasive pest species that threatens native fauna worldwide. Increasingly, environmental DNA (eDNA) is used as a technique to monitor the presence of invasive species given its power to detect low numbers of individuals. We aimed to investigate eDNA persistence in freshwater at three different temperatures (25, 30 and 35°C) and eDNA detection thresholds for R. marina using controlled experiments in aquaria. For the latter, two water volumes and two cane toad exposure times were used (800 or 200 L volume with 5 or 30 min exposure). A 15-ml water sample was collected from each replicated aquaria and preserved with 5 ml Longmire's buffer. Environmental DNA was extracted and four technical quantitative PCR replicates were analyzed targeting the cane toad 16S rDNA mitochondrial gene. Environmental DNA decayed rapidly in water and was reliably detected for up to 3 days after cane toad removal, regardless of the temperature treatment. Also, cane toad eDNA was detected in the water after a 5-min initial exposure of a single individual in 800 L of water. Under the physical parameters tested here, a positive detection means that a cane toad has been in contact with the water body between 1 and 3 days prior to the sampling event. The results of the present study show the importance of eDNA for determining the presence of a species that occurs at low abundance in a small water body, such as at the onset of a cane toad invasion.