During a parasitological survey of Galaxias maculatus (Jenyns, 1842) in the Maullín Basin (Chilean Patagonia), specimens of a new species of Monogenea were collected from the gills. This species is described as the only member of a proposed new genus, Inserotrema n. gen. (Dactylogyridae, Ancyrocephalinae), characterized by similar hooks with 2 subunits, overlapping gonads, coiled cirrus with counterclockwise rings, articulated accessory piece formed by 2 parts, a needlelike sclerite threading the distal part of the MCO, and a sclerotized midventral vagina. This new genus is proposed for dactylogyrids from gills of galaxiids (Galaxiidae). Inserotrema puyei n. sp. infects gills of G. maculatus from Llanquihue Lake, Maullín River, and Maullín Estuary. This is the first species of Ancyrocephalinae described from gills of a galaxiid.