Detection and population genomics of sea turtle species via noninvasive environmental DNA analysis of nesting beach sand tracks and oceanic water.

Elusive aquatic wildlife, such as endangered sea turtles, are difficult to monitor and conserve. As novel molecular and genetic technologies develop, it is possible to adapt and optimize them for wildlife conservation. One such technology is environmental (e)DNA - the detection of DNA shed from organisms into their surrounding environments. We developed species-specific green (Chelonia mydas) and loggerhead (Caretta caretta) sea turtle probe-based qPCR assays, which can detect and quantify sea turtle eDNA in controlled (captive tank water and sand samples) and free ranging (oceanic water samples and nesting beach sand) settings. eDNA detection complemented traditional in-water sea turtle monitoring by enabling detection even when turtles were not visually observed. Furthermore, we report that high throughput shotgun sequencing of eDNA sand samples enabled sea turtle population genetic studies and pathogen monitoring, demonstrating that noninvasive eDNA techniques are viable and efficient alternatives to biological sampling (e.g., biopsies and blood draws). Genetic information was obtained from sand many hours after nesting events, without having to observe or interact with the target individual. This greatly reduces the sampling stress experienced by nesting mothers and emerging hatchlings, and avoids sacrificing viable eggs for genetic analysis. The detection of pathogens from sand indicates significant potential for increased wildlife disease monitoring capacity and viral variant surveillance. Together, these results demonstrate the potential of eDNA approaches to ultimately help understand and conserve threatened species such as sea turtles.

Environmental DNA monitoring of oncogenic viral shedding and genomic profiling of sea turtle fibropapillomatosis reveals unusual viral dynamics.

Pathogen-induced cancers account for 15% of human tumors and are a growing concern for endangered wildlife. Fibropapillomatosis is an expanding virally and environmentally co-induced sea turtle tumor epizootic. Chelonid herpesvirus 5 (ChHV5) is implicated as a causative virus, but its transmission method and specific role in oncogenesis and progression is unclear. We applied environmental (e)DNA-based viral monitoring to assess viral shedding as a direct means of transmission, and the relationship between tumor burden, surgical resection and ChHV5 shedding. To elucidate the abundance and transcriptional status of ChHV5 across early, established, regrowth and internal tumors we conducted genomics and transcriptomics. We determined that ChHV5 is shed into the water column, representing a likely transmission route, and revealed novel temporal shedding dynamics and tumor burden correlations. ChHV5 was more abundant in the water column than in marine leeches. We also revealed that ChHV5 is latent in fibropapillomatosis, including early stage, regrowth and internal tumors; higher viral transcription is not indicative of poor patient outcome, and high ChHV5 loads predominantly arise from latent virus. These results expand our knowledge of the cellular and shedding dynamics of ChHV5 and can provide insights into temporal transmission dynamics and viral oncogenesis not readily investigable in tumors of terrestrial species.

Molecular characterization of a marine turtle tumor epizootic, profiling external, internal and postsurgical regrowth tumors.

Sea turtle populations are under threat from an epizootic tumor disease (animal epidemic) known as fibropapillomatosis. Fibropapillomatosis continues to spread geographically, with prevalence of the disease also growing at many longer-affected sites globally. However, we do not yet understand the precise environmental, mutational and viral events driving fibropapillomatosis tumor formation and progression.Here we perform transcriptomic and immunohistochemical profiling of five fibropapillomatosis tumor types: external new, established and postsurgical regrowth tumors, and internal lung and kidney tumors. We reveal that internal tumors are molecularly distinct from the more common external tumors. However, they have a small number of conserved potentially therapeutically targetable molecular vulnerabilities in common, such as the MAPK, Wnt, TGFß and TNF oncogenic signaling pathways. These conserved oncogenic drivers recapitulate remarkably well the core pan-cancer drivers responsible for human cancers. Fibropapillomatosis has been considered benign, but metastatic-related transcriptional signatures are strongly activated in kidney and established external tumors. Tumors in turtles with poor outcomes (died/euthanized) have genes associated with apoptosis and immune function suppressed, with these genes providing putative predictive biomarkers.Together, these results offer an improved understanding of fibropapillomatosis tumorigenesis and provide insights into the origins, inter-tumor relationships, and therapeutic treatment for this wildlife epizootic.