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This study compared the nutrient-energy retention, digestive function, growth performance, and welfare of rainbow trout (ibw 54 g) fed isoproteic (42%), isolipidic (24%), fishmeal-free diets (CV) over 13 weeks. The diets consisted of plant-protein replacement with graded levels (10, 30, 60%) of protein from poultry by-product (PBM) and black soldier fly

Simple Summary Sustainability and fish welfare have been receiving increasing attention in the aquaculture sector, with an emphasis on the search for new, sustainable, and healthy aquafeed ingredients. For many years, plant ingredients have been widely used in aquafeed formulation; however, negative side effects on gut welfare have often been reported in several carnivorous fish species. From this perspective, alternative ingredients such as poultry by-products and insect meal are receiving attention due to their low ecological footprint and high nutritional value. In the present study, these two ingredients were used, singly or in combination, to formulate practical diets for gilthead seabream (Sparus aurata). After a twelve-week feeding trial, a multidisciplinary laboratory approach including histological, molecular, and spectroscopic techniques was adopted in order to investigate fish physiological responses to the new test diets. The results obtained showed excellent zootechnical performances and ameliorated gut health in fish fed dietary inclusions of poultry by-products and insect meal compared to those fed a vegetable-based diet. In addition, the modulation of nutrient absorption in relation to the ingredients used was highlighted by means of spectroscopic tools. The results obtained demonstrated that poultry by-products and insect meal can be successfully used to replace plant-derived ingredients in diets for gilthead seabream without negatively affecting fish welfare. Abstract The attempt to replace marine-derived ingredients for aquafeed formulation with plant-derived ones has met some limitations due to their negative side effects on many fish species. In this context, finding new, sustainable ingredients able to promote fish welfare is currently under exploration. In the present study, poultry by-products and Hermetia illucens meal were used to replace the vegetable protein fraction in diets totally deprived of fish meal intended for gilthead seabream (Sparus aurata). After a 12-week feeding trial, a multidisciplinary approach including histological, molecular, and spectroscopic techniques was adopted to investigate intestine and liver responses to the different dietary formulations. Regardless of the alternative ingredient used, the reduction in dietary vegetable proteins resulted in a lower incidence of intestine histological alterations and inflammatory responses. In addition, the dietary inclusion of insect meal positively affected the reduction in the molecular inflammatory markers analyzed. Spectroscopic analyses showed that poultry by-product meal improved lipid absorption in the intestine, while insect meal induced increased liver lipid deposition in fish. The results obtained demonstrated that both poultry by-products and H. illucens meal can successfully be used to replace plant-derived ingredients in diets for gilthead seabream, promoting healthy aquaculture.