A pot experiment in greenhouse was carried out to investigate effects of application of rhamnolipid (RH), a type of biosurfactant on the effect of PAHs-specific degrading bacteria (DB) in bioremediation of soils that have long been contaminated by PAHs. Results indicate that application of RH in addition to DB inoculation significantly increased total PAHs and promoted degradation of PAHs different in number of rings in the soil. After 90 days of incubation, the PAHs degradation ratio of Treatments RH, DB and RH+DB reached 21.3%, 32.6% and 36.0%, respectively, and increased by 333.0%, 563.3% and 633.0% as compared against that (4.9%) of the control. In addition, the average PAHs degradation ratio declined with the number of rings of the 15 PAHs. It was also found that the number of PAHs degrading bacteria, dehydrogenase activity and polyphenol oxidase activity in soils of Treatments DB and RH+DB were much higher than in Treatment RH and in CK, but differed slightly between Treatment RH and CK, suggesting that the mechanism of DB promoting degradation of PAHs in the soil is different from that of RH.