Study On The Effect Of Rhodococcus Erythropolis KB1 In Remediation Of Oil-contaminated Soil

Petroleum pollution has becoming one of the most serious environment problems in the oil exploitation and application.Bioremediation is considered as one of the most environmentally friendly and low-cost remediation technologies,which has broad application prospects.In this study,highly efficient oil degrading bacteria Rhodococcus erythropolis KB1 and Bacillus thuringiensis FD1 isolated from local area were used to study their bioremediation effect of natural degradation,bioaugmentation,biostimulation on the contaminated oil in simulated polluted desert soils,actual oilfield polluted soils and simulated contaminated farmland soil.The microbial diversities in petroleum-contaminated desert soil during bioremediation process were analyzed by high-throughput sequencing technique.The main results were as follows:The oil degradation effect of the four oil degrading bacteria was studied.It was found that the crude oil degradation rates of Rhodococcus erythropolis KB1,Bacillus mojavensis JC1,Bacillus thuringiensis FD1 and Microbacterium sp.FD4 were57.55%,20.20%,31.26%and 31.20%,respectively when they were cultured in the minimal medium containing crude oil for sevevn days.However,the degradation rates were not increased significantly in the groups with the combination of different strains.Bioremediation of the actual oilfield contaminated soil was carried out for 115days,the results showed that the degradation rates of the total petroleum hydrocarbon(TPHs)in the natural degradation,KB1 bioaugmentation,biostimulation and bioaugmentation-biostimulation groups were 45.96%,47.81%,47.23%and 49.94%,respectively.The changes of the various components of petroleum hydrocarbons were analyzed by gas chromatography-mass spectrometer(GC-MS).It was found that the degradation rates of residual oil fraction in the groups of the KB1 bioaugmentation and the combination of bioaugmentation and biostimulation were 100.00%.The degradation rates of diesel component were more than 40.00%and the degradation rates of the gasoline component were less than 50.00%in four treatments.The degradation effect of pristane and phytane was the best in the natural degradation group.The culturable bacterial numbers were measured by plate count method.It was found that the number of cultivable bacteria in the soil of the actual oilfield had no significant difference in the changes of each remediation(P?0.05).For the simulated polluted farmland soils,the degradation rates of the TPHs in the natural degradation treatment,FD1 bioaugmentation,KB1 bioaugmentation,biostimulation and bioaugmentation-biostimulation groups were 51.84%,54.51%,53.12%,39.93%and 53.28%,respectively.The degradation rate of gasoline was100.00%in the natural remediation group,the degradation rates of phytan,diesel and residual oil components in the FD1 bioaugmentation group were 100.00%,each component has the worst degradation effect in the biostimulation group.The counts of culturable bacteria in the unpolluted farmland soil were maintained at 3.89×10⁵CFU·g^-1-10.0×10⁵ CFU·g^-1 and maintained at 2.0×10⁶ CFU·g^-1 in the natural degradation and biostimulation treatments.The numbers of culturable bacteria in the bioaugmentation and bioaugmentation-biostimulation groups increased significantly(P<0.05).For the simulated polluted desert soils,the degradation rates of the TPHs in the natural degradation treatment,FD1 bioaugmentation,KB1 bioaugmentation,biostimulation and bioaugmentation-biostimulation groups were 20.20%,38.86%,36.02%,40.10%and 43.89%,respectively.The degradation rate of gasoline components in the natural degradation group was 100.00%,the degradation rates of residual oil components in the KB1 bioaugmentation and the combined of bioaugmentation and biostimulation groups were 100.00%,but each component has the worst degradation effect in the biostimulation group.The culturable bacterial numbers in the unpolluted desert soil were maintained at 3.75×10⁴ CFU·g^-1-7.33×10⁵CFU·g^-1.The number of the culturable bacteria in the bioaugmentation,biostimulation and bioaugmentation-biostimulation groups were higher than that in the natural degradation group(P<0.05).High-throughput sequencing technology analyzed the bacterial community diversity of the simulated polluted desert soils and found that the main dominant bacterial communities in uncontaminated desert soil were Proteobacteria,Patescibacteria,Actinobacteria,Acidobacteria,Planctomycetes,Chloroflexi and Bacteroidetes.The dominant genera were RB41,Gemmatimonas,Lysobacter and Dietzia.After oil pollution,there was no significant changes in the composition of the microorganisms at phylum and genus level,but the significant difference in abundances were observed.Actinobacteria became the most dominant phylum,while the abundances of Patescibacteria,Gemmatimonadetes and Bacteroidetes decreased significantly.On the genus level,the abundance of Nocardioides increased,while the abundances of Erythrobacter and Lysobacter,RB41,Sphingomonas and Gemmatimonas decreased.Bacillus was the most dominant bacteria in the FD1treatment group,and the abundances of Rhodococcus increased significantly in the groups with KB1 and the combined group.After 115 days of remediation,the abundance and uniformity of microorganisms in the soil were reduced.The main dominant genera of the bioaumentation group with FD1 treatment were Erythrobacter,Dietzia,Nocardioides,Croceicoccus,Lysobacter,Alcanivorax,Streptomyces,Acinetobacter.The dominant bacteria genera of KB1treatment group were Erythrobacter,Dietzia,Nocardioides,Alcanivorax,Rhodococcus,Croceicoccus,Lysobacter,Streptomyces.The dominant bacteria genera of the biostimulation group were Dietzia,Virgibacillus,Streptomyces,Nocardioides,Rhodococcus,Pseudogracilibacillu,Erythrobacter.The main dominant bacteria genera of the combined group of bioaugmentation-biostimulation were Dietzia,Virgibacillus,Pseudogracilibacillus,Rhodococcus,Streptomyces,Lysobacter,Nocardioides.