Crude oil degradation efficiency of a recombinant Acinetobacter baumannii strain and its survival in crude oil-contaminated soil microcosm

A hydrocarbon degrading Acinetobacter baumannii S30 strain, isolated from crude oil-contaminated soil, was inserted with the lux gene from the luciferase gene cassette luxCDABE. Soil microcosms were designed to study the degradation efficacy for total petroleum hydrocarbon (TPH) of crude oil by lux-tagged A. baumannii S30 pJES. Bioaugmentation of a TPH-contaminated microcosm with A baumannii S30 pJES showed that TPH levels were reduced from 89.3 to 53.9 g/kg soil in 90 days. Biodegradation of TPH by A baumannii S30 pJES was also monitored in shake flask conditions, which showed a reduction of initial TPH levels by over 50% at the end of 120 h. A lux-PCR-based approach along with the standard dilution plating with selective antibiotics was successfully utilized to monitor the survivability of the lux-tagged strain A. baumannii S30 pJES in soil microcosms and stability of the lux insert in the host strain A. baumannii S30. The selective plating technique indicated the population of A. baumannii S30 pJES to be 6.5+/-0.13 x 10(8) CFU/g at day zero (just after bioaugmentation) and 2.09+/-0.08 x 10(8) CFU/g of soil after 90 days of incubation. lux-PCR confirmed the stability of the insert in all the randomly selected colonies of A. baumannii strains from the antibiotic plates. The lux insert was stable after 50 generations in Luria Bertini broth and storage at -70 degrees C as glycerol stocks for over a year. These results revealed that the lux insert was stable and lux-tagged A. baumannii S30 strain could survive in a TPH-contaminated soil microcosm and could degrade TPH in the soil microcosm conditions. It can be used as an effective marker to monitor the survival of augmented strains at a bioremediation site.