Abstract: We developed a Chinese herbal compound to improve the immunity of Chinese soft-shelled turtle (Pelodiscus sinensis) and invetigated its effect on the regulation of intestinal microflora, so as to provide references for its disease control. In this study, we divided 100 individuals of P. sinensis with an average body mass of (45.53±3.89) g into two groups. The control group (CG) was fed with a basal diet and the experimental group (CPG) was fed with a diet supplemented with 2% of compound herbs for 42 d. The results show that the dominant phylum was the same in both groups, but the proportion of each dominant bacterial phylum differed: the beneficial flora of Tenericutes, Planctomycetes and Lactobacillus in the intestine of the CPG group was 32, 2.25 and 4.12 times higher than that of the CG group, respectively. After being challenged by 10⁷ CFU·mL⁻¹ of Aeromonas hydrophila, the activity of lysozyme (LZM), total superoxide dismutase (T-SOD), alkaline phosphatase (AKP) and acid phosphatase (ACP) in serum of P. sinensis were determined. The results show that the activities of T-SOD, LZM, ACP and AKP in the CG and CPG groups reached the peak values at 72^nd hour, and activities of the four enzymes in the serum of CPG group were significantly higher than those in CG group under the normal conditions and after the challenge , except for LZM, which was not significantly different when the two groups were challenged for 12 h. In conclusion, this Chinese herbal compound can effectively optimize the intestinal microflora of P. sinensis and has certain immune enhancing effects, which shows a application prospect.