Amphibia-infecting members of the genus Trypanosoma represent its most divergent lineage and exhibit remarkable morphological and genetic diversity. However, their detailed study has been hindered by pleomorphism, morphological convergence, multiple infections and scarcity of molecular data. By combining light microscopy with molecular phylogenetics, we investigated trypanosome diversity in a broad sample of frogs from Panama, a climatically stable tropical biodiversity hotspot. The uncovered diversity of trypanosomes parasitizing amphibians was exceptional, exceeding the host species richness twofold. Phylogenetic analyses revealed conspicuous ecological partitioning: distinct lineages were primarily associated with the arboreal hylids, with only rare host switches to understorey species, suggesting that vector feeding preferences structure the parasites' community. Notably, none of the identified haplotypes matched those from South America, underscoring geographic isolation as a driver of diversification. This study revealed a vast, previously undescribed diversity of trypanosomes that reflects the ecological and taxonomic breadth of their amphibian hosts. Given the ongoing global amphibian decline, documenting these parasitic communities is urgent. Our findings highlight how host ecology and biogeography shape parasite evolution, and offer a framework for future research in threatened tropical ecosystems.

Department of Biology University of Oxford Oxford UK

Department of Parasitology Faculty of Science Charles University Prague Czech Republic

Faculty of Science University of South Bohemia České Budějovice Czech Republic

Institute of Parasitology Biology Centre Czech Academy of Sciences České Budějovice Czech Republic

Life Science Research Centre Faculty of Science University of Ostrava Ostrava Czech Republic

Military Health Institute Military Medical Agency Prague Czech Republic

Smithsonian Tropical Research Institute Panama City Panama

Zoological Institute Russian Academy of Sciences Saint Petersburg Russia

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Trypanosomes in Neotropical frogs: unveiling hidden megadiversity and complex host-parasite patterns