In order to study the effect of three heterotrophic nitrifying bacteria on water quality of tilapia pond with bottom aeration, 600 Oreochromis niloticus with initial body weight of (2.66±0.94) g were randomly divided into 20 ponds. Five treatments included control group 1 (with air stone), control group 2 (with bottom microporous aerator), Rhodococcus pyridinivorans (P1-2) group (with bottom microporous aerator), Alcaligenes faecalis (P3-1) group (with bottom microporous aerator), and Bacillus megaterium (P5-2) group (with bottom microporous aerator), and each group had four replicates. Heterotrophic nitrifying bacteria with a final concentration of 5 x 10⁵ CFU/mL were added into the ponds every week. Water quality, growth and antioxidative abilities of fish were determined. The concentration of total nitrogen in water of all groups showed a continuous increasing trend during the experiment, while the accumulation of all indexes in the experiment groups were always lower than that in the control groups. The effect of P5-2 on improving the water quality of O. niloticus pond was the best. The accumulation of total nitrogen in pond water of P5-2 group was lower than that of control group 1 and control group 2 during the whole experiment. The accumulation of total nitrogen in pond water of P3-1 group was always lower than that of control group 2 during the whole experiment. The accumulation of total nitrogen in pond water of the P1-2 group was significantly lower than that of control group 1 and control group 2 at the 8th and 10th weeks. The accumulation of ammonia nitrogen in pond water of P5-2, P3-1 and P1-2 groups was lower than that of control group 1 and control group 2 during the whole experiment. The accumulation of nitrite nitrogen content in pond water of P5-2 group was significantly lower than that of control group 2 at the 2nd, 6th and 8th weeks. Adding heterotrophic nitrifying bacteria in pond water improved the antioxidative abilities of O. niloticus. Compared with control group 1, total superoxide dismutase (SOD) activity in serum of tilapia in P3-1 and P5-2 groups increased by 12.10% and 8.05%, respectively. Compared with control group 2, SOD activity in serum of O. niloticus in P3-1 and P5-2 groups increased by 21.25% and 16.87%, respectively. Compared with the control group 1, total antioxidant capacity (T-AOC) activity in serum of O. niloticus in P1-2, P3-1 and P5-2 groups increased by 82.78%, 98.68% and 245.70% respectively. Compared with control group 2, T-AOC activity in serum of O. niloticus in P3-1 and P5-2 groups increased by 1.69% and 76.95%, respectively. The results showed that adding heterotrophic nitrifying bacteria into pond could effectively improve the water quality and the antioxidative abilities of O. niloticus, but had no effect on the growth of O. niloticus.