Open cast coal mining operations involve the use of the explosive Ammonium Nitrate Fuel Oil (ANFO) for detonation processes. Five bacterial strains belonged to Pseudomonas sp and Pseudomonas stutzeri were isolated from an open cast coal mine located in La Guajira, Colombia. Degradation of the second component of ANFO, which is diesel, by the five isolates and by a consortium was evaluated. The biodegradation of diesel was determined by gas chromatography with a flame ionization detector. Biodegradation Efficiency (BE) was 96% for the consortium; individual strains had levels between 10-95% of BE. Analysis for the presence and expression of the alkane monooxygenase gene involved in the degradation of diesel was evidenced in two strains showing band size products between 500-600 bp. Results suggest that these bacteria are candidates for diesel bioremediation.