The microbial community response during oxygen biostimulation process of aged oil-polluted soils is poorly documented and there is no reference for long-term monitoring of unsaturated zone. Two treatments (0 and 0.056 mol×h−1 molar flow rate of oxygen) were performed in fixed bed reactors containing oil-polluted soil in order to assess the potential effect of air-supplying on hydrocarbon fate and microbial community structure. Microbial activity was continuously monitored during two years throughout the oxygen biostimulation process. Microbial community structure before and after 12 and 24 months treatment was determined by a dual rRNA/rRNA gene approach allowing us to characterize bacteria which were presumably metabolically active and therefore responsible for the functionality of the community in this polluted soil. Clone library analysis revealed that the microbial community contained many rare phylotypes. These were never observed in other studied ecosystems. The bacterial community shifted from Gammaproteobacteria to Actinobacteria during the treatment. Without air supplying, the samples were dominated by a phylotype linked to the Streptomyces. Members belonging to eight dominant phylotypes were well adapted to air supplying process. The air supplying stimulated an Actinobacteria phylotype which might be involved in the restoring of the studied ecosystem. Phylogenetic analyses suggested that this phylotype is a novel, deep-branching member of the Actinobacteria related to the well studied genus Acidimicrobium.