The phylogeography of trypanosomes from South American alligatorids and African crocodilids is consistent with the geological history of South American river basins and the transoceanic dispersal of Crocodylus at the Miocene

Abstract Background Little is known about the diversity, phylogenetic relationships, and biogeography of trypanosomes infecting non-mammalian hosts. In this study, we investigated the influence of host species and biogeography on shaping the genetic diversity, phylogenetic relationship, and distribution of trypanosomes from South American alligatorids and African crocodilids. Methods Small Subunit rRNA (SSU rRNA) and glycosomal Glyceraldehyde Phosphate Dehydrogenase (gGAPDH) genes were employed for phylogenetic inferences. Trypanosomes from crocodilians were obtained by haemoculturing. Growth behaviour, morphology, and ultrastructural features complement the molecular description of two new species strongly supported by phylogenetic analyses. Results The inferred phylogenies disclosed a strongly supported crocodilian-restricted clade comprising three subclades. The subclade T. grayi comprised the African Trypanosoma grayi from Crocodylus niloticus and tsetse flies. The subclade T. ralphi comprised alligatorid trypanosomes represented by Trypanosoma ralphi n. sp. from Melanosuchus niger, Caiman crocodilus and Caiman yacare from Brazilian river basins. T. grayi and T. ralphi were sister subclades. The basal subclade T. terena comprised alligatorid trypanosomes represented by Trypanosoma terena n. sp. from Ca. yacare sharing hosts and basins with the distantly genetic related T. ralphi. This subclade also included the trypanosome from Ca. crocodilus from the Orinoco basin in Venezuela and, unexpectedly, a trypanosome from the African crocodilian Osteolaemus tetraspis. Conclusion The close relationship between South American and African trypanosomes is consistent with paleontological evidence of recent transoceanic dispersal of Crocodylus at the Miocene/Pliocene boundaries (4–5 mya), and host-switching of trypanosomes throughout the geological configuration of South American hydrographical basins shaping the evolutionary histories of the crocodilians and their trypanosomes.

We are grateful to many people who kindly helped us in crocodilian capture and sample collection in Brazil, Venezuela, and Guinea Bissau. We would like to thank Dr. Miguel U. Trefault Rodrigues for the animal identifications. We acknowledge the Brazilian Ministry of Science, Technology and Innovation (MCTI) for support through the Mamirauá Institute for Sustainable Development (IDSM). We thank Cristina Schwartz for the coordination of the work in Guinea Bissau. We also thank Marcio C. Valentin from the Laboratory of Electron Microscopy, Institute of Biosciences, USP, and Carlos E. Jared and Marta M. Antoniazzi from the Institute Butantan, São Paulo, SP, Brazil, for their kindly permission to use their electron microscopic facilities. This work was supported by CNPq (PROAFRICA and PROSUL) and CAPES (Programa Nacional de Incentivo à Pesquisa em Parasitologia Básica). LBV was postdoctoral fellow sponsored by CNPq-MCTI (PROTAX – National Program of Taxonomy) and CAPES (PNPD). BRF is recipient of a scholarship from CNPq (PROTAX).