Linking the epidemiology of coccidioidomycosis and environmental exposure through targeted genomic enrichment of Coccidioides posadasii

Valley fever, a disease caused by Coccidioides spp., is a fungal respiratory disease with an expanding range. Methods to culture the pathogen from soil, especially Coccidioides posadasii, are very challenging, limiting the genomics knowledge of environmental strains. In this study, we designed and tested a targeted DNA capture and enrichment system for the characterization of Coccidioides genomes without the need to culture. In this system, RNA probes are hybridized to Coccidioides DNA in a complex sample, followed by DNA amplification, sequencing, and analysis. Our enrichment system was targeted toward coding region sequences in C. posadasii str. Silveira and tested on control DNA spiked into soil; DNA hybridized to probes was then sequenced and correctly placed into a reference phylogeny, based on the known placement of the whole-genome sequence. We then applied the enrichment system to a range of sample types (soil, air filters, rodent tissue) from a site in Mesa, Arizona, USA. The enriched samples were sequenced and placed into the C. posadasii phylogeny to understand the phylogenetic diversity within the Mesa site over time. The results demonstrate that low DNA signal in most sample types was boosted after enrichment. Enriched sequences from air filters collected at multiple time points from the Mesa site linked two different isolates collected from fatal cases of Coccidioidomycosis in a pig-tailed macaque colony housed at the Mesa site. This represents the first time that environmental C. posadasii DNA was directly linked to Coccidioidomycosis and demonstrates the power of this approach for genomic epidemiology. IMPORTANCE All human cases of Valley fever are acquired through environmental exposure, so surveillance and characterization of the pathogen in soil are critical for risk mitigation efforts. Current databases are biased toward human clinical isolates, and little is known about the genomics of environmental strains of Coccidioides posadasii. In this study, we designed, tested, and validated a probe enrichment system that amplifies trace DNA in a complex sample. Sequenced DNA can be used to link environmental exposure with human cases, directing public health agencies to interventions that limit human exposure. This use case was demonstrated in this study, as trace DNA trapped on air filters was linked to a fatal case of primate Coccidioidomycosis at a site in Arizona. The probe enrichment system described in this study represents a powerful tool to better understand the genomic composition of environmental C. posadasii strains, which can aid in public health investigations.