Toluene uptake by a benthic biofilm community was previously shown to vary seasonally from 0.03 m hr?1 in winter to 0.2 m hr?1 in summer in a solvent-contaminated stream of the Aberjona watershed. We used quantitative PCR to estimate the population dynamics of previously isolated species of toluene-degrading Xanthobacter autotrophicus and Mycobacterium sp. in both toluene-contaminated and uncontaminated reaches of the stream, and to estimate their relative roles in overall biodegradation rate. Quantification using specific 16S rDNA primers forX. autotrophicus and Mycobacterium sp. showed that populations of both species were much larger in the toluene-contaminated than the toluene-free reach, in agreement with earlier culture-based investigations. A relatively brief bloom of X. autotrophicus occurred in the contaminated reach in the summer, while Mycobacterium sp. populations occurred at elevated densities for more than 5 months. Calculations showed that Mycobacterium, previously thought to be less important than Xanthobacter in annual toluene degradation based on single time-point CFU estimates, appears actually more important because of this longer persistence.