Developing terrestrial environmental DNA sampling methods for detecting arboreal invasive reptiles: a case study of the green anole in the Ogasawara Island, Japan

Early detection of invasive alien species is essential for preventing establishment and mitigating ecological impacts, particularly in island ecosystems harbouring evolutionarily isolated endemic species. Recently, despite increasing reptile introductions and their suggested widespread impacts, methods for monitoring arboreal invasive reptiles remain limited. This study addressed the need for practical detection tools by developing terrestrial environmental DNA (eDNA) sampling methods that collect DNA from leaf surfaces to detect the arboreal invasive green anole ( Anolis carolinensis ) in the Ogasawara Islands, Japan. Two methods, wiping leaf surfaces with gauze and rinsing with sprayed water, were tested. At an invaded site, green anoles were successfully detected in eight out of 10 samples in both methods. Although no significant difference in eDNA concentration was observed, the wiping method was selected for its greater simplicity. Subsequently, the relationship between green anole population density and eDNA concentrations detected using the wiping method was investigated, suggesting a significant positive relationship. This is the first report demonstrating that terrestrial eDNA concentrations can reflect arboreal reptile population density, suggesting potential applications in quantitative terrestrial biodiversity assessments. Furthermore, the successful detection of eDNA even in the extremely low-density habitat of the green anole demonstrates the usefulness of eDNA-based surveys for early detection of invasions. The method developed here may be broadly applicable to terrestrial biodiversity monitoring, especially in tree-dwelling taxa. Given accelerating biological invasions and biodiversity loss, this approach is expected to benefit managers, conservationists, and researchers concerned with terrestrial ecosystems.