Shotgun sequencing of airborne eDNA achieves rapid assessment of whole biomes, population genetics and genomic variation.

Biodiversity and its associated genetic diversity are being lost at an unprecedented rate. Simultaneously, the distributions of flora, fauna, fungi, microbes and pathogens are rapidly changing. Novel technology can help to capture and record genetic diversity before it is lost and to measure population shifts and pathogen distributions. Here we report the rapid application of shotgun long-read environmental DNA (eDNA) analysis for non-invasive biodiversity, genetic diversity and pathogen assessments from air. We also compared air eDNA with water and soil eDNA. Coupling long-read sequencing with established cloud-based biodiversity pipelines enabled a 2-day turnaround from airborne sample collection to completed analysis by a single investigator. To determine the full utility of airborne eDNA, we also conducted a local bioinformatic analysis and deep short-read shotgun sequencing. From outdoor air eDNA alone, comprehensive genetic analysis was performed, including population genetics (phylogenetic placement) of a charismatic mammal (bobcat, Lynx rufus) and a venomous spider (golden silk orb weaver, Trichonephila clavipes), and haplotyping humans (Homo sapiens) from natural complex community settings, such as subtropical forests and temperate locations. The rich datasets also enabled deeper analysis of specific species and genomic regions of interest, including viral variant calling, human variant analysis and antimicrobial resistance gene surveillance from airborne DNA. Our results highlight the speed, versatility and specificity of pan-biodiversity monitoring via non-invasive eDNA sampling using current benchtop/portable and cloud-based approaches. Furthermore, they reveal the future feasibility of scaling down (equipment and temporally) these approaches for near real-time analysis. Together these approaches can enable rapid simultaneous detection of all life and its genetic diversity from air, water and sediment samples for unbiased non-targeted information-rich genomics-empowered (1) biodiversity monitoring, (2) population genetics, (3) pathogen and disease-vector genomic surveillance, (4) allergen and narcotic surveillance, (5) antimicrobial resistance surveillance and (6) bioprospecting.