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An 8-week feeding trial was conducted to evaluate the effects of dietary yeast culture (YC) supplementation on growth performance, antioxidant activity, nonspecific immunity, and disease resistance of Pseudobagrus ussuriensis (average initial weight 6.01 ± 0.01 g ). Four isonitrogenous and isolipidic diets were formulated to contain 0 (Y0), 10 (Y1), 20 (Y2), and 30 (Y3) g/kg YC, respectively. After the feeding experiment, the challenge test of injecting Aeromonas hydrophila was executed. Results showed that appropriate YC supplementation level in the diet could improve growth performance, digestive enzyme activities, nonspecific immunity capacity, antioxidant capacity, and disease resistance of P. ussuriensis. And the highest weight gain, feed intake, specific growth rate, and IGF-1 gene expression level were observed in fish fed the Y2 diet. The activities of protease and amylase in intestine in fish fed the Y2 diet were enhanced compared with that in fish fed the Y0 diet significantly ( P < 0.05 ). Simultaneously, fish fed the Y2 diet had significantly higher serum lysozyme activity and significantly lower serum alanine amino transferase activity ( P < 0.05 ). Dietary 20 g/kg YC supplementation increased the activity of catalase and total antioxidant capacity in liver and reduced malondialdehyde content in the liver and intestine of P. ussuriensis significantly ( P < 0.05 ). Fish fed the Y2 diet had the highest disease resistance under the condition of A. hydrophila challenge ( P < 0.05 ). The quadratic regression analysis based on weight gain against dietary YC levels indicated that the appropriate dietary YC supplementation level is 13.4 g/kg diet.