Triplicate groups of rainbow trout (Oncorhynchus mykiss) (1.4 ± 0.1 g) were fed experimental diets containing four levels of protein substitution to totally replace fishmeal with poultry by-product meal (0, 33, 67 and 100%). Fishmeal (FM) from tuna fish by-products and poultry by-product meal ‘pet food grade' (PBM) were used. Diets were formulated to be isonitrogenous (43.5% crude protein) and isolipidic (12.5% crude fat). After 80 days of feeding, no significant differences were found in terms of thermal unit growth coefficient (TGC). However, the EPA levels in muscle tissue among treatments 0PBM, 24PBM and 44PBM were found to be significantly higher than those for the 59PBM diet. Nevertheless, a re-feeding period with the control diet showed that 28 days was long enough to regain the FA profile needed. We concluded that PBM used up to 44% in diets for juvenile rainbow trout can be used without a significantly decrease in EPA and DHA under the present conditions. The use of a whole fishmeal diet could be an important strategy to recuperate the fatty acid profile obtained when trout is feed on PBM basis. It will be important to perform longer experiments with larger fish to confirm these results.