A Direct DNA Extraction Workflow for Metabarcoding Fungal Bioaerosols from Adhesive Samplers

Monitoring fungal bioaerosols is essential for understanding their impacts on human health and crop productivity. However, current monitoring approaches rely on culture-dependent techniques, which underestimate the diversity of fungal exposures. Adhesive-coated films can effectively capture airborne particles but have been regarded as unsuitable for DNA downstream analysis, owing to the lack of standardised DNA extraction methods for the adhesive capture surface. Here, we present an optimised DNA extraction protocol for adhesive film, which we developed and validated to enable direct and culture-independent ITS2-based metabarcoding of fungal bioaerosols collected on a 96 well plate sealing film ( Sticky sampler). This validated approach was applied to field samples collected from four agricultural farms and from a domestic garden to evaluate seasonal and spatial changes in fungal bioaerosols. Our analysis uncovered pronounced site-specific contrasts linked to geography and land management, as well as clear seasonal shifts in garden fungal communities. This study bridges a long-standing methodological gap, demonstrating that adhesive samplers, combined with optimised DNA extraction, provide a practical and inexpensive tool for fungal bioaerosol research by resolving ecologically meaningful spatiotemporal dynamics of airborne fungi. This protocol opens new opportunities for highly scalable surveillance of environmental bioaerosols.