AbstractIn this study it is reported the operation of a horizontal‐flow anaerobic immobilized biomass (HAIB) reactor under sulfate‐reducing condition which was also exposed to different amounts of ethanol and toluene. The system was inoculated with sludge taken from up‐flow anaerobic sludge blanket (UASB) reactors treating refuses from a poultry slaughterhouse. The HAIB reactor comprised of an immobilized biomass on polyurethane foam and ferrous and sodium sulfate solutions were used (91 and 550 mg/L, respectively), to promote a sulfate‐reducing environment. Toluene was added at an initial concentration of 2.0 mg/L followed by an increased range of different amendments (5, 7, and 9 mg/L). Ethanol was added at an initial concentration of 170 mg/L followed by an increased range of 960 mg/L. The reactor was operated at 30(±2)°C with hydraulic detention time of 12 h. Organic matter removal efficiency was close to 90% with a maximum toluene degradation rate of 0.06 mgtoluene/mgvss/d. Sulfate reduction was close to 99.9% for all‐nutritional amendments. Biofilm microscopic characterization revealed a diversity of microbial morphologies and DGGE‐profiling showed a variation of bacterial and sulfate reducing bacteria (SRB) populations, which were significantly associated with toluene amendments. Diversity of archaea remained unaltered during the different phases of this experiment. Thus, this study demonstrates that compact units of HAIB reactors, under sulfate reducing conditions, are a potential alternative for in situ aromatics bioremediation. © 2005 Wiley Periodicals, Inc.