Abstract: The statistical experimental design (Plackett-Burman and Box-Behnken design) was applied to optimize the culture medium of nitrite oxidizing bacteria for improving the nitrite oxidizing rate. Estimated optimum medium composition of the nitrite oxidizing rate was as follows: NaHCO3 2.0 gL-1; NaNO2 2.36 gL-1; Na2CO3 0.37 gL-1; NaCl 0.34 gL-1; KH2PO4 0.05 gL-1; MgSO47H2O 0.05 gL-1; and FeSO47H2O 0.03 gL-1. The nitrite oxidizing rate reached a maximum at 905.0 mg NO2-N(g MLSSd)-1 (mixed liquor suspended solids, MLSS). In the field trial, 50 L of nitrite oxidizing bacteria concentrate (1.99 g VSSL-1)(volatile solid, VSS) with 850 mg NO2-N(g MLSSd)-1 were added to 0.6 hectares of the aquaculture water. Nitrite level in all treated ponds remained very low compared to the steady increase observed in all of the control ponds during 7 days.