Abstract

not-yet-known not-yet-known not-yet-known unknown Global loss of biodiversity prioritizes the need for comprehensive and effective biomonitoring methods. Airborne eDNA has shown promise for monitoring terrestrial vertebrates but has not yet been rigorously compared to established biomonitoring methods. This study aims to compare species detection from airborne eDNA (eDNA) with observer-based monitoring (OBM), camera trapping (CT) and passive acoustic monitoring (PAM), focusing on birds and mammals. Monitoring was performed over the course of four weeks within an agroforestry ecosystem in the Netherlands. Birds were monitored using eDNA, OBM and PAM, while eDNA and CT were used for mammals. Four mammal species were detected by both CT and eDNA, while eDNA identified an additional 17 species, primarily small-bodied, including two invasive species. A total of 78 bird species was detected, with 28 identified by all three methods. In terms of species uniquely identified per method, eDNA detected the most (19 species), followed by PAM (10, but including 4 misidentifications) and OBM (2). All but three bird and four mammal species detected were consistent with known occurrences near the study site . Rarefaction shows that eDNA has the highest potential species diversity, but OBM is most efficient when limited time is available. Unique species can largely be explained by method characteristics or limitations; PAM detections are limited to species that make sound, while eDNA requires further research on detection range and sensitivity. This is the first study to compare airborne eDNA with OBM and acoustic data, further confirming the high potential of airborne eDNA for biodiversity monitoring.