In this study, we determined the effects of dietary chitosan on the growth, biochemical composition, gut microbiota, and hematological and histological parameters of juvenile Barbonymus gonionotus. Three test diets containing three different concentrations (1, 2, and 3 g kg-1 feed) of dietary chitosan were formulated. A basal diet without dietary chitosan was considered a control, and the fish were reared for 60 days. Comparing the effects of the dietary chitosan-containing diets with those of the control diet, we found that dietary chitosan significantly improved the muscle growth, nutrient and mineral contents, hematological parameters, lactic acid bacterium abundance, and digestive enzyme activities of B. gonionotus. Moreover, dietary chitosan significantly inhibited the growth of pathogenic bacteria in fish. Interestingly, an increase in the dietary chitosan level significantly enhanced the protein contents of the muscles and inversely significantly decreased the lipid contents compared to those with the basal diet. Quantitative study revealed that dietary chitosan significantly enhanced the length of intestinal villi, and qualitative study showed that dietary chitosan considerably reduced the fat content in the liver and improved the morphology of the kidney compared to those with the basal diet. Taken together, our results suggest that the application of dietary chitosan at a dose of 1 g kg-1 feed produced the highest benefit to treated B. gonionotus, indicating its potential for safe use in aquaculture.