Specimens of the Nile puffer, Tetraodon lineatus Linnaeus (Tetraodontiformes: Tetraodontidae), were collected from the Nile River, Qena Governorate, Egypt (26°08′N, 32°42′E), and examined for monogenoidean parasites. A single species, initially identified as Heterobothrium lineatum Morsy, Saady, Abdel-Gaffar, Bashtar, Melhorn, Al Quraishy & Adel, 2012 (Monogenoidea: Diclidophoridae), was recovered from the gill lamellae of six out of seven examined hosts (prevalence = 86%). The taxonomic and phylogenetic status of this helminth was reassessed using an integrative approach combining morphological and molecular data. Comparative morphological analysis revealed that H. lineatum represented a junior subjective synonym of H. fluviatile Euzet & Birgi, 1975. Molecular analysis based on the 28S rDNA gene, representing the first inclusion of H. fluviatile as an ingroup taxon, placed this species within a well-supported clade of Heterobothrium species infecting tetraodontid fishes. Heterobothrium indicum Verma, 2022, a parasite of the goldband goatfish, Upeneus moluccensis (Bleeker) (Mulliformes: Mullidae), is herein transferred to Upenicola Unnithan, 1966 as Upenicola indicus (Verma, 2022) n. comb. The study highlighted the need for further investigation into the taxonomic status of Diclidophora indica Tripathi, 1959, and Neodiclidophora simhai Gupta & Krishna, 1980, as probable members of Heterobothrium. Moreover, Heterobothrium lamothei of Morsy et al. (nec Vidal Martinez & Mendoza-Franco) is regarded as a species inquirenda.