TEN PURE bacterial isolates with biosurfactant activity were obtained from autochthonous microflora of oil contaminated soil. Soil samples were selected from bus garage stations in Cairo and from different plots around oil wells of western desert of Egypt. Biosurfactant production was investigated using three parameters (hydrolyses of blood, surface tension reduction, and emulsification index techniques). The biosurfactant accumulation was coincided with growth. By means of standard bacterial identification procedures, the isolates were identified as different strains of Bacillus licheniformis. The phylogenetic relationships between these strains were determined using the RAPDPCR technique. The similarity between the strains was ranging from 78 to 100 %.