Environmental DNA succeeds in detecting a low‐density population of endangered harlequin frogs

Abstract The neotropical harlequin frog genus Atelopus is among the most severely impacted by population declines and extinctions. Environmental DNA (eDNA) offers a valuable, non‐invasive tool to detect low‐density and elusive species. We evaluated its effectiveness in detecting the recently rediscovered, Critically Endangered, Jambato Harlequin Frog ( Atelopus ignescens ) across seasons and habitats in Angamarca, Ecuador, the species' only known locality, where just 71 individuals (including 6 tadpoles) were recorded after a year of intensive monthly surveys. We collected 18 water samples from 14 sites, including rivers, creeks, and an irrigation ditch used by adults, during both wet and dry seasons. eDNA metabarcoding analysis detected the species in 5 of 18 samples, all from the wet season. Most detections ( n = 4) were from rivers, mainly toward the end of the wet season (early May), including three from the only river where tadpoles had been previously observed. The remaining detections came from the least degraded site and the irrigation ditch. Our findings highlight eDNA's value for detecting A. ignescens during reproductive periods and in tadpole habitats. The study underscores the potential of eDNA to inform monitoring and conservation strategies and emphasizes the importance of conserving less disturbed habitats to ensure the survival of critically endangered species.