Avocado is affected by Colletotrichum gloeosporioides causing anthracnose. Antagonistic microorganisms against C. gloeosporioides represent an alternative for biological control, therefore, in this work we focused on the isolation and the characterization of potential antagonist bacteria against a member of the C. gloeosporioides species complex for their possible future application.Samples of avocado rhizospheric soil were obtained from an orchard located in Ocuituco, Morelos, Mexico, in order to obtain bacterial isolates with potential antifungal activity. From the soil samples, 136 bacteria were isolated and then they were challenged against a member of the C. gloeosporioides species complex, only three bacterial isolates A1, A2 and A3 significantly diminished the mycelial fungus growth in 75, 70 and 60%, respectively. Two of these isolates were identified by 16S rRNA, as Bacillus mycoides (A1 and A2) and the third as Bacillus tequilensis (A3). Bacillus mycoides bacterial cell-free supernatant reduced mycelial growth of a member of the C. gloeosporioides species complex isolated from avocado in 65%, whereas Bacillus tequilensis A3 supernatant did so in 25% after 3 days post inoculation. B. tequilensis mycoides A1was a producer of proteases, indolacetic acid and siderophores. Preventive treatment using cell-free supernatant of B. mycoides A1, diminished the severity of anthracnose disease (41.9%) on avocado fruit.These results reveal the possibility of using Bacillus mycoides A1 as a potential biological control agent. This article is protected by copyright. All rights reserved.