Common root rot (CRR) caused by Cochliobolus sativus (Ito & Kuribayashi) Drechs. ex Dastur is a serious disease of barley causing significant economic losses worldwide. An in vitro technique was developed for assessing directly the antagonistic potential of Bacillus atrophaeus isolate SY15b against CRR on barley. Seeds of the susceptible barely cv. WI 2291 were inoculated by mixing them thoroughly with C. sativus-SY15bsuspension and placed onto 1.5% water agar in sterile Petri dishes for 10 days. Our results demonstrated that B. atrophaeus had significant antagonistic capacities against the CRR by decreasing the percentage of barley infected area of sub-crown internodes (SCIs) as compared to controls after 10 days post incubation. Highly significant correlation coefficients (r = 0.85, P = 0.001) were found between in vitro and pot methods, indicating that in vitro co-culture testing procedure is reliable. The developed method is both simple and precise and could be used directly for a rapid assessment of the antagonistic activity of Bacillus on barley plants under uniform conditions for selecting suitable biocontrol agent against fungal soil-borne pathogens.