We describe a new species of Phyllodactylus from a xerophytic forest in the Andes of southern Ecuador. The new species differs from other mainland species of Phyllodactylus from South America by the combination of the following characters: preanal scales similar in size to other ventral scales, well-defined longitudinal rows of tubercles dorsally, a medial longitudinal row of enlarged caudal scales ventrally, tubercles on dorsal surface of tibia present, enlarged postanal scale absent, tubercles on dorsal surface of forearm absent, interorbital scales homogeneous in size, enlarged scales on proximal one-fourth of tail absent, gular scales granular, and maximum snout–vent length of 55 mm. Even though morphological similarity suggests a close relationship between the new species and Phyllodactylus reissii, phylogenetic analyses of 10 nuclear and mitochondrial genes contradict this hypothesis. We also report the first records of Phyllodactylus kofordi for Ecuador.