Enhancement of repeated inoculation strategy with a domesticated bacterial consortium on the biodegradation of high-level crude oil in soil

Repeated inoculation of hydrocarbon degrading microbes should be powerful to improve the survival of inoculant, which is vital to achieve efficient remediation of petroleum contaminated soil. This paper aims to study the repeated inoculation (with different inoculum size and time interval) enhanced bioremediation of high-level petroleum contaminated soil with a domesticated bacterial consortium. The copy number of bacterium and alkB gene, soil enzyme activities and microbial community structure during the remediation were systematically analyzed to preliminarily reveal the mechanism of repeated inoculation affecting remediation for the first time. The results revealed that repeated inoculation remarkably improved the total petroleum hydrocarbon (TPH) removal in soil (86.5 % in HC120) compared with a single inoculation (68.9 % in HA120). The TPH removal of repeated inoculation with high inoculum size (HC) on the 60th day was close to that of once inoculation (HA) on the 120th day, suggesting that repeated inoculation led to faster degradation. Interestingly, the effect of inoculation with low dose and more times (LC120, 78.5 %) was equal to that with high dose and less times (HB120, 78.0 %), even much better than that with high dose and once inoculation (HA120). Treatment HC had a significant impact on the soil bacterial diversity and community structure, and the dominant species in the inoculants, such as Stenotrophomonas and Pseudomonas, which was low abundance in the blank group (CK), still maintained high abundance during the remediation process. The soil catalase activities and the number of alkB gene were the highest in HC. Correlation analysis implied that repeated inoculation of hydrocarbon degrading bacteria did improve the survival of inoculant, soil enzyme activities and maintain the number of degrading bacteria, thus promoting the TPH removal. These findings will facilitate the practical application of bioremediation technology to contaminated environment, which has important environmental and economic benefits.