A New High Throughput Sequencing Assay for Characterizing the Diversity of Natural Communities and Its Application to a Pacific Oyster Mortality Event.

The genus is notable for including several pathogens of marine animals and humans, yet characterization of diversity using routine 16S rRNA sequencing methods is often constrained by poor resolution beyond the genus level. Here, a new high throughput sequencing approach targeting the heat shock protein () as a phylogenetic marker was developed to more precisely discriminate members of the genus in environmental samples. The utility of this new assay was tested using mock communities constructed from known dilutions of isolates. Relative to standard and -specific 16S rRNA sequencing assays, the assay delivered high levels of fidelity with the mock community composition at the species level, including discrimination of species within the clade. This assay was subsequently applied to characterize community composition in seawater and delivered substantially improved taxonomic resolution of species compared to 16S rRNA analysis. Finally, this assay was applied to examine patterns in the community within oysters during a Pacific oyster mortality event. In these oysters, the assay identified species-level community shifts prior to disease onset, pinpointing as a putative pathogen. Given that shifts in the community can precede, cause, and follow disease onset in numerous marine organisms, there is a need for an accurate high throughput assay for defining community composition in natural samples. This -centric sequencing assay offers the potential for precise high throughput characterization of diversity, providing an enhanced platform for dissecting dynamics in the environment.