Temperature Exposure and Collection Substrate Choice Govern Post‐Capture Persistence of Airborne eDNA

ABSTRACT Airborne environmental DNA (eDNA) offers a powerful, noninvasive approach to monitor terrestrial biodiversity. However, its broader implementation is constrained by limited understanding of DNA persistence across the eDNA lifecycle, including biological shedding, environmental transport, capture efficiency, and post‐capture decay. While aquatic and soil studies provide valuable insight into eDNA degradation processes, airborne eDNA presents distinct challenges, and isolating individual lifecycle stages remains necessary for accurate signal interpretation. Here, we focus on one critical and underexplored stage: post‐capture DNA persistence on collection substrates. We conducted a 180‐day controlled laboratory trial to quantify how environmental exposure and material choice influence DNA signal retention following capture. Using a standardized vertebrate DNA source proxy (pork meat meal), we evaluated seven candidate collection materials, including five dry substrates and two liquid media, exposed to two temperature regimes (23°C and 40°C) and three light exposure levels. By directly applying DNA to substrates, we decoupled post‐capture DNA decay from variability in airborne transport and capture efficiency. We modeled DNA signal loss using exponential decay functions, revealing temperature as the dominant driver of post‐capture degradation across materials. Light exposure had no measurable effect under the low UV irradiance tested. DNA declined most rapidly within the first 2 weeks following capture, and decay constants ( k ) varied widely among substrates. Dry substrates retained DNA comparably to positive controls over extended periods, whereas liquid media exhibited accelerated signal loss and higher PCR inhibition. Pairwise statistical comparisons confirmed significant differences in both persistence and inhibition among materials. Together, these results demonstrate that collection substrate choice strongly shapes post‐capture airborne eDNA signal stability and downstream analytical performance. By quantifying material‐ and temperature‐dependent decay trajectories, this study provides empirical guidance for selecting sampling substrates and defining deployment duration, supporting more reliable interpretation of airborne eDNA detections in terrestrial biodiversity monitoring.