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In addition to its high content in protein and lipid, Black soldier fly (BSF) (Hermetia illucens. L) has antimicrobial peptides (AMP) that could inhibit the growth pathogenic microbes. Therefore BSF extract could be very potential in substituting the use of antibiotics in animal production system. This research was conducted to determine the antibacterial activity of BSF extract in vitro on the growth of Salmonella typhimurium, E. coli and Pseudomonas aureginosa. The experiment was carried out according to the Completely Randomized Design (CRD) consisting of six treatments and three replications for each treatment. The treatments were different concentration levels of BSF extract, i.e. 75, 125, 175, 225, 275 and 325 mg.ml-1. Chloramphenicol with concentration of 30 µg.discs paper-1 was used as a positive control and dimethyl sulfoxide (DMSO) as a negative control. BSF extract was made using maceration extraction method. The antibacterial activity of BSF extract was determined by measuring the diameter of inhibitory zone (mm) formed around the discs paper following the 24 h of the incubation period. The results of this study indicated that the antibacterial activity of BSF extract increased (P<0.05) in line with the increase level of BSF extract concentration. The average diameter of the inhibition zone for Salmonella typhimurium, E. coli and Pseudomonas aureginosa was 11.77 ± 0.03 mm, 11.15 ± 0.05 mm, and 11.15 ± 0.23 mm respectively, which was categorized as strong inhibition zone, even though was still lower compared to a very strong inhibiton zone of chloramphenikol, which was 14.74 ± 0.38 - 27.61 ± 0.35. In conclusion, the concentration of BSF extract of 325 mg.ml-1 is an effective concentration to inhibit the growth of the bacteria Salmonella typhimurium, E. coli and Pseudomonas aureginosa.