Widespread amphibian Perkinsea infections associated with Ranidae hosts, cooler months and Ranavirus co-infection

Amphibians suffer from large-scale population declines globally, and emerging infectious diseases contribute heavily to these declines. Amphibian Perkinsea (Pr) is a worldwide anuran pathogen associated with mass mortality events, yet little is known about its epidemiological patterns, especially in comparison to the body of literature on amphibian chytridiomycosis and ranavirosis. Here, we establish Pr infection patterns in natural anuran populations and identify important covariates including climate, host attributes and co-infection with Ranavirus (Rv). We used quantitative (q)PCR to determine the presence and intensity of Pr and Rv across 1234 individuals sampled throughout central Florida in 2017-2019. We then implemented random forest ensemble learning models to predict infection with both pathogens based on physiological and environmental characteristics. Perkinsea infected 32% of all sampled anurans, and Pr prevalence was significantly elevated in Ranidae frogs, cooler months, metamorphosed individuals and frogs co-infected with Rv, while Pr intensity was significantly higher in ranid frogs and individuals collected dead. Ranavirus prevalence was 17% overall and was significantly higher in Ranidae frogs, metamorphosed individuals, locations with higher average temperatures, and individuals co-infected with Pr. Perkinsea prevalence was significantly higher than Rv prevalence across months, regions, life stages and species. Among locations, Pr prevalence was negatively associated with crayfish prevalence and positively associated with relative abundance of microhylids, but Rv prevalence did not associate with any tested co-variates. Co-infections were significantly more common than single infections for both pathogens, and we propose that Pr infections may propel Rv infections because seasonal Rv infection peaks followed Pr infection peaks and random forest models found Pr intensity was a leading factor explaining Rv infections. Our study elucidates epidemiological patterns of Pr in Florida and suggests that Pr may be under-recognized as a cause of anuran declines, especially in the context of pathogen co-infection.