Fusarium root rot is an important disease of soybean in Ontario, Canada. This study is to select antagonistic bacterial agents as effective alternatives to chemical pesticides for the control of root rots caused by Fusarium oxysporum and F. graminearum. Twenty-two Bacillus subtilis strains from soybean and corn roots were tested in dual cultures for inhibition of mycelial growth of F. oxysporum and F. graminearum. All strains significantly reduced mycelial growth of F. oxysporum by approximately 17 to 48% and of F. graminearum by 10 to 32%. Ten B. subtilis strains selected based on their larger fungal inhibition zones were evaluated against macroconidial germination. These strains inhibited the spore germination of F. oxysporum by 20 to 48% and of F. graminearum by 14 to 32% in cell-free filtrates. Under greenhouse conditions, the efficacy of seed and soil treatments with B. subtilis strains against the two Fusarium root rot pathogens was evaluated based on root rot severity, seedling emergence, plant height, and root dry weight. Six B. subtilis strains (SB01, SB04, SB23, SB24, SB28, and SB33) from soybean roots and two strains (CB01 and CH22) from corn roots significantly reduced the severity of the two Fusarium root rots in seed or soil treatments. Strains SB01, SB04, SB23, and SB24 were the most effective treatments against both pathogens in either seed or soil treatment. When applied as seed treatments, these four strains reduced root rot severity by 43 to 63% and increased emergence by 13 to 17%, plant height by 9 to 18%, and root dry weight by 8.4 to 19%. When used as soil treatments, they reduced root rot severity by 68 to 74% and increased emergence by 14 to 18%, plant height by 11 to 23%, and root dry weight by 16 to 24%. These results suggest that the novel strains of B. subtilis identified in this research can be effective alternatives to fungicides in managing Fusarium root rots of soybean, and a greater level of efficacy may be achieved when they were used as soil treatments than seed treatments.