This study aimed to investigate the synergetic effects of dietary emodin and Clostridium butyricum on antioxidant capacities and immune gene expression of freshwater prawn (Macrobrachium rosenbergii) under ammonia stress. Nine experimental diets were formulated with three emodin levels (0, 25 and 50 mg/kg) and three C. butyricum levels (0, 250 and 500 mg/kg, 2 × 107 CFU g⁻¹) arranged in a 3 × 3 factorial design. Emodin dictated the relatively lower superoxide dismutase (SOD) activity after stress compared to before. Glutathione peroxidase (GPx) exhibited a significant difference among treatments fed emodin after ammonia exposure. Emodin and C. butyricum sparked the relatively lower malondialdehyde (MDA) levels before and after stress compared with the control. The expression of heat shock protein (HSP70) significantly augmented by emodin alone (p < 0.001) and in the interaction with C. butyricum (p < 0.01), while C. butyricum showed insignificant result before stress (p > 0.05) independently. After the ammonia challenge, the expression of HSP70 was significantly down‐regulated by the individual and synergetic effects of emodin and C. butyricum (p < 0.001). Toll gene expression was significantly increased by both emodin and C. butyricum before and after ammonia antagonism (p < 0.001). Similarly, dorsal gene expression was remarkably enhanced by emodin and C. butyricum before and after ammonia challenge (p < 0.01), and synergetic effects were observed before stress exposure (p < 0.05). The present study indicated that the synergetic effects of C. butyricum and emodin could significantly improve the antioxidant capacity, immune‐related gene expression and survival rate of M. rosenbergii under ammonia stress. The optimum combination of these components was 250 mg/kg (2x 107 CFUg⁻¹) and 50 mg/ kg.