All A. stritum strains, isolated from grain sorghum plants showing Acremonium wilt disease symptoms, varied in their virulence on both tested grain sorghum genotype with the superiority of strain No. 2.  Among 5 different recommended media, King's medium was the most favorable one for propagation and highest antagonistic activity of Bacillus subtilis, previously isolated from rhizosphere of resistant grain sorghum cultivar (Dorado) against A. strictum. The optimum environmental conditions needed for B. subtilis to give maximum antagonistic efficiency against A. strictum were 48h incubation period, 30-35^oC incubation temperature and pH 7.0. in shake-flask submerged culture. Sucrose (9.0 g.l^-1) and KNO₃(20 g.l^-1) were the best carbon and nitrogen sources, respectively. Utilization of agro-industrial wastes for B. subtilis  propagation in modified King's medium showed that glucose  syrup and corn steep liquor as cheap carbon and nitrogen sources, respectively supported the highest antagonistic potential of B. subtilis  using shake-flask submerged culture.