A moderately thermophilic iron-oxidizing bacterium strain TI-1 has been shown to have a unique enzyme system that produces hydrogen sulfide (H2S) extracellularly under acidic conditions. The optimal culture conditions for H2S production were determined to apply the bacterium to the treatment of acidic wastewater containing heavy metals. The production of H2S by the cells was dependent on L-glutamic acid, ferrous sulfate and elemental sulfur in the medium. The optimal culture medium for H2S production contained 0.3% L-glutamic acid, 1.7% ferrous sulfate, and 1.0% elemental sulfur and the optimal pH and temperature were 4.0 and 55 degrees C, respectively. Under the optimal culture conditions, strain TI-1 produced H2S continuously at a rate of 0.26 mu mol/ml/d for 14 d.