Echinostoma paraensei Lie and Basch, 1967 (Echinostomatidae:Platyhelminthes), a 37 collar spine echinostome of the "revolutum group", has been used extensively as a model organism to study the interactions of digenetic trematodes with both their snail and vertebrate hosts. This worm was first isolated from the snail Biomphalaria glabrata from Belo Horizonte (BH isolate), Minas Gerais State, Brazil, by Lie and Basch [J Parasitol (1967) 53:1192-1199]. The natural definitive host for the BH isolate was never determined, and it has been maintained in the laboratory since 1967 in B. glabrata and hamsters. In this study, using light and scanning electron microscopy and molecular analysis, we describe an echinostome recently obtained from its natural vertebrate host, the wild rodent Nectomys squamipes (Rodentia: Sigmodontinae) from Sumidouro, Rio de Janeiro State, Brazil (RJ isolate). This echinostome was also compared to the laboratory-maintained BH isolate of E. paraensei. We observed that adult worms of both BH and RJ isolates could be differentiated from other echinostome species by the relatively small size of the dorsal collar spines relative to lateral and corner collar spines. SEM confirmed the similarity of this morphological character between the two isolates. As additional diagnostic features, the tegumentary spines are scale-like and the region between the genital pore and the acetabulum lacks scales. There is a folded protuberance with a pore just posterior to the genital pore. The tegument of the acetabulum is unspined and radially wrinkled, and there are numerous randomly distributed small, domed, ciliated papillae. The sequences of the internal transcribed spacers of the nuclear rDNA complex of the RJ and BH isolates are identical. Together these shared features provide strong evidence that both isolates are the same and can be referred to as E. paraensei. In conclusion, we have identified, for the first time, one of the natural definitive hosts for E. paraensei, the rodent N. squamipes, and have extended the known geographical distribution of this species to include Sumidouro in Rio de Janeiro State, Brazil.