A pot experiment, designed to have five treatments, was carried out in greenhouse to investigate effects of bio-organic fertilizer(BOF) and BOF plus soil amendments on tobacco bacterial wilt and on microflora in rhizosphere soil. The five treatments were set as follows: T1, the control; T2, the pot soil was applied with common organic fertilizer; T3, the pot soil was applied with the BOF; T4, the pot soil was treated with lime before the application of the BOF; T5, the pot soil was treated with lime and bicarbonate before the application of the BOF. DGGE and plate counting were used to investigate microflora in the rhizosphere soils. Relusults show that T2 not only had no control effect against tobacco bacterial wilt, but also increased the disease index, while the control effects against tobacco bacterial wilt of T3, T4 and T5 varied in the range of 66.7%~87.9%. All the organic fertilizer treatments had some influence on microflora in the rhizosphere, increasing the number of bacteria by 3.5 and 6.1 times and the number of actinomycetes by 3.7 and 3.5 times, but decreasing the number of fungi by 66.2% and 70.1%, respectively in T3 and T4, and increasing the number of bacteria and actinomycetes by 13.6 times and 5.1 times, respectively, but decreasing the number of fungi by 75.0% in T5. The preliminary study found that a combined treatment of the soil with lime and bicarbonate before application of the BOF could control tobacco bacterial wilt effectively by altering composition of the microflora in tobacco rhizosphere, and decreasing the number of pathogens.