Taxonomia integrativa de nematódeos Oswaldocruzia (Trichostrongyloidea: Molineidae) da Amazônia Oriental

Abstract

Oswaldocruzia is a genus of parasitic nematodes of amphibians (Anura, Caudata) and reptiles (Squamata), represented by about 90 species distributed worldwide, of which 43 occur in the Neotropical region. The species of Oswaldocruzia are mainly characterized by the spicular morphology of the males and are divided into five biogeographic groups (Oriental-Ethiopian, Neo-Ethiopian, Holartic, Neotropical Caribbean and Neotropical Continental) and also three morphological types of copulatory bursa (types I, II and III). However, morphological similarity, absence of updated identification keys and molecular data complicate the systematic of the genus. Thus, the objective of this study was the realization of the integrated taxonomy of nine species of Oswaldocruzia parasitizing eight species of amphibians and one reptile species from different collections in three locations in the state of Pará, Brazil. The hosts were necropsied and the helminths found were rinsed, fixed and stored in 70% ethanol. For the morphological study, the specimens were used for observation by light microscopy and scanning electron microscopy. For the molecular study, we performed the extraction, amplification and sequencing of the coding region of the Cytochrome c Oxidase Subunit I enzyme from the mitochondrial DNA. The resulting sequences were aligned and compared to ten sequences publicly available in GenBank and two phylogenetic reconstructions were performed to observe their relationships, one under the Maximum Likelihood criterion and the other by Bayesian Inference. As results we identified Oswaldocruzia belenensis in Rhinella marina and Rhinella margaritifera, Oswaldocruzia chabaudi in Boana geographica and Boana wavrini, Oswaldocruzia chambrieri in Amazophrynella bokermanni and R. margaritifera, Oswaldocruzia lanfrediae in Leptodactylus paraensis, Oswaldocruzia vitti in Anolis fuscuauratus, Oswaldocruzia sp. nov. 1 in Phyllomedusa vaillantii and Oswaldocruzia sp. nov. 2 in Osteocephalus oophagus. Alignment and comparison of levels of divergence demonstrated significant differences between the new obtained sequences and the sequences from GenBank. Both phylogenetic reconstructions demonstrated two main clades, one including the sequences from Mexico and another clade genetically distinct from the Amazon, highlighting the occurrence of O. chabaudi in B. wavrini and B. geographica, the close relationship between the sequences from parasites of Bufonidae and new records of hosts and localities for O. chambrieri and O. belenensis. This study adds information about the diversity of helminth parasites of amphibians and reptiles of Amazon, and shows that the combination of morphological and molecular methods presents satisfactory potential for species delimitation and characterization of the genus.