Abstract: Gymnotus tiquie, new species, is described from the Rio Tiquié, a tributary of the Uaupés (Vaupés) in the upper Negro basin, Amazonas, Brazil. The new species was collected in non-floodplain (terra firme) streams, where it occurs sympatrically and syntopically with two geographically widespread congeners, the type species of the genus, G. carapo, and G. coropinae. The new species is diagnosed by a unique combination of morphometric, meristic, and osteological traits, and by a characteristic color pattern in which the dark oblique pigment bands, diverse in shape and design, are divided into band-pairs along the length of the body, in which the band-pairs are often recurved (dorsally concave), more variable, and often reticulated in the abdominal region, and in which the pale inter-bands meet at the dorsal mid-line along most of the length of the body. Gymnotus tiquie is a member of the G. pantherinus species group, with which it shares the presence of one (vs. two) pore in the dorsolateral portion of the preopercle (except in G. pantanal and G. anguillaris), needle-shaped (vs. conical or arrowhead-shaped) teeth on the dentary and premaxilla, and a slender body (BD 5.6–10.6% HL vs. deep 8.7–13.5%, except G. chaviro, G. curupira, G. varzea, G. chimarrao, G. maculosus, G. henni, and G. inaequilabiatus that also have a slender body). Gymnotus tiquie is most similar in overall appearance to G. cataniapo of the upper Orinoco. These two species share three unique features within the G. pantherinus group: dark band-pairs with wavy irregular margins along the length of the body, a long body cavity with 45 or more pre-caudal vertebrae, and a darkly pigmented membrane in the caudal region of the anal fin.