The biodegradation of No. 2 diesel fuel under anaerobic conditions was investigated using anaerobic soil slurry reactors. Soil samples were collected from a diesel spill site, where the diesel spill has migrated to vadose zone. The purpose of this study was to find out which of the various groups of anaerobic bacteria present in the study site will degrade the petroleum hydrocarbons in the soil. The various anaerobic groups included sulfate-, nitrate-reducing, methanogenic, and mixed electron acceptor conditions. The results indicated enhanced biodegradation of diesel fuel under all conditions compared to no electron-supplemented reactor (passive bioremediation). However, the rate of diesel degradation was the highest under mixed electron acceptor conditions followed in order by sulfate-, nitrate-reducing, and methanogenic conditions. Under mixed electron acceptor condition, 80.5% removal of diesel fuel was achieved within 290 days. While under sulfate-reducing condition 55% degradation of diesel fuel was observed for the same period. This study showed that the mixed heterogeneous microbial population of the contaminated site has the metabolic capability to degrade diesel fuel. (C) 2003 Elsevier Ltd. All rights reserved.