An annotated checklist of the ichthyofauna from the upper Pirai river drainage is provided. The Pirai river was a major right-bank tributary of the Paraiba do Sul river, but it has been artificially diverted to the coastal Guandu river system in southeastern Brazil to generate electric power and water for the metropolitan area of Rio de Janeiro. Based on our field sampling of 23 sites, 32 species belonging to 24 genera and 12 families were collected in 6 headwater tributaries of the Pirai river between 2009 and 2016. Phalloceros harpagos (Lucinda, 2008), Astyanax intermedius (Eigenmann, 1908), and Neoplecostomus microps (Steindachner, 1877b) were the most abundant and most widely distributed species in the samples. The sampled ichthyofauna is mostly composed by species from the Paraiba do Sul basin. Eight species are reported for the first time in the upper Pirai drainage, showing the importance of continuous ichthyofaunal surveys of fish in remaining areas of Atlantic Forest. Key words Atlantic Forest, coastal streams, ichthyological survey, Neotropical fishes, Paraiba do Sul. Academic editor: Mariangeles Arce H. | Received 19 September 2018 | Accepted 8 January 2019 | Published 22 February 2019 Citation: de Brito V, Buckup PA (2019) The fish fauna of the upper Pirai drainage, a transposed mountain river system in southeastern, Brazil. Check List 15 (1): 235-247. https://doi.org/10.15560/15.L235