Background Hydrocarbon-degrading biofilms in the treatment of contaminated groundwaters have received increasing attention due to the role played in the so-called "biobarriers". These are bioremediation systems in which a microbial consortium adherent to a solid support is placed across the flow of a contaminated plume, thus promoting biodegradation of the pollutant. Results A microbial consortium adherent to pumice granules (biofilm) developed from a toluene-enriched microflora in a mini-scale system, following continuous supply of a mineral medium containing toluene, over a 12-month period. Observation by scanning electron microscopy, together with quantification of the biomass attached to pumice, evidenced the presence of abundant exopolymeric material surrounding the cells in the biofilm. Toluene removal monitored during 12-month operation, reached 99%. Identification of the species, based on comparative 16S ribosomal DNA (rDNA) sequence analysis, revealed that Rhodococcus erythropolis and Pseudomonas marginalis were the predominant bacterial species in the microbial consortium. Conclusion A structurally complex toluene-degrading biofilm, mainly formed by Rhodococcus erythropolis and Pseudomonas marginalis, developed on pumice granules, in a mini-scale apparatus continuously fed with toluene.