Introduction:The increasing practice of ecotourism and rural tourism in the State of Minas Gerais, Brazil, highlights the importance of studies concerning the occurrence of potential intermediate hosts of Schistosoma mansoni. This study aimed to identify species of Biomphalaria snails in municipalities along the Estrada Real, an important Brazilian tourism project. Methods: The specimens were collected in different water collections of 36 municipalities along the Estrada Real in the southeast of the State of Minas Gerais. Biomphalaria species were characterized using both morphological and molecular approaches. The research was conducted between August 2005 and September 2009 and all the sites visited were georeferenced using GPS. Results: Six Biomphalaria species were found in 30 of the 36 municipalities studied: glabrata, tenagophila, straminea, peregrina, occidentalis and schrammi. The first three species of Biomphalaria, recognized as intermediate hosts of S. mansoni, were present in 33.3%, 47.2% and 8.3% of the municipalities studied, respectively. The mollusks were found in different types of water collections and no infection by S. mansoni was detected. The highest occurrence of Biomphalaria concentration was verified in the area covered by the Caminho Novo route (Diamantina/MG to Rio de Janeiro/RJ). Conclusions: Considering the occurrence of schistosomiasis in the State of Minas Gerais and the socioeconomic repercussions involved in the Estrada Real Project, this work focuses on the vulnerability of water collections due to the presence of Biomphalaria mollusks and emphasizes the need for epidemiological surveillance and sanitary and educational measures integrated with the local community and tourism sectors.