A packed-bed bioreactor inoculated with a mixed culture obtained from a contaminated site was used to continuously treat a saturated solution of nonylphenol. The reactor was operated at feeding rates of 13-112 ml h-1 and temperatures of 5.5, 10, and 15 degrees C. Optimal bioreactor performance was achieved at 10 degrees C and at a feeding rate of 84 ml h-1 (with a removal rate of 43 mg l-1 day-1 of nonylphenol). No endocrine activity was observed in the effluent of the bioreactor at any of the temperatures tested, and the only metabolic products found were branched carboxylic acids and alkanes (lacking an aromatic ring). The study of the microbial populations in the biofilm at the three temperatures tested using fluorescence in situ hybridization showed that all the bacterial species that could be identified belonged to the phylum Proteobacteria. The most abundant class identified at all three temperatures was beta-Proteobacteria. The proportions of bacteria that bound to the specific probes among the total population, identified with the bacterial probe EUB338MIX, were 60, 43, and 24% at 15, 10, and 5.5 degrees C, respectively.