In contrast to mice or zebrafish, trout exhibits post-larval muscle growth through hypertrophy and formation of new myofibers (hyperplasia). The muscle fibers are formed by the fusion of mononucleated cells (myoblasts) regulated by several muscle-specific proteins such as myomaker or myomixer. In this work, we identified a unique gene encoding a myomixer protein of 77 amino acids (aa) in the trout genome. Sequence analysis and phylogenetic tree, showed moderate conservation of the overall protein sequence across teleost fish (61% of aa identity between trout and zebrafish myomixer sequences). Nevertheless, the functionally essential motif, AxLyCxL is perfectly conserved in all studied sequences of vertebrates. Using in situ hybridization, we observed that myomixer was highly expressed in the embryonic myotome, particularly in the hyperplasic area. Moreover, myomixer remained readily expressed in white muscle of juvenile (1 and 20 g) although its expression decreased in mature fish. We also showed that myomixer is up-regulated during muscle regeneration and in vitro myoblasts fusion. Together, these data indicate that myomixer expression is consistently associated with the formation of new myofibers during somitogenesis, post-larval growth and muscle regeneration in trout.