| The Petroleum Hydrocarbon(PH) is a rare resource in nowadays, and it takes part a very important role in the world including industry, aviation, sea transportation. With the development of the society, during the exploitation, smelting, utilizition, transportion, omitting of the oil, and the discharged mud disposals, sewage irrigation, some substance of oiliness of volatilizing and burning uncompleted, more and more soil, water, air has been polluted. TPH has contained so many composition, such as hydrocarbon, alkene, naphthenic, aromatic hydrocarbon.The way of physics or chemistry is not very good to degrade the petroleum in the environment. And the biological method is very effective to degrade the petroleum in the environment.This issue focus on that the isolation and idenfication of petroleum-degrading strains from the petroleum-polluted soil. After acclimation, enrichment, streaking, two strains which can degrade petroleum were obtained. The two strains were added to petroleum-polluted soil and the efficiency rate of TPH were determined using infrared oil analyzer. In order to study degradation mechanism, the degrading genes and the number of degrading strains were determined using PCR-agarose electrophoresis analysis and most probable number (MPN), respectively. After11week degradation experiment, we get the following conclusion:(1) Two bacterium were screened from the petroleum-pollution soil. According to the results of molecular biology identification, two bacterium belong to Pseuomonas aeruginosa and Bacillus, respectively. Two strains can grow on TPH which acted as only carbon source.(2) In order to determine the degradation efficiency of two strains toward petroleum. Four treatments were excuted:petroleum soil (1#; control experiment); petroleum soil+nutrition N、P (2#); petroleum soil+nutrition N、P+Pseuomonas aeruginosa strain(3#); petroleum soil+nutrition N、P+Bacillus strain(4#). The results shown that when adding nutriention N、P with two strains to petroleum-polluted soil, two strains has stronger degradation effects towards TPH than the controlled experiment. After7wks, the degradation efficiency of TPH reached78.4%and82.7%in treatment3#and treatment4#; the TPH degradation efficiency was54.9%in treatment2#; in the control, TPH degradation efficiency was42.8%. At llwk, the TPH degradation efficiency reached93.3%and90.8%in treatment3#and treatment4#; and the TPH degradation efficiency were88.9%and63.6%in treatment2#and treatment1#, respectively.(3) DNA was extracted and diluted from different treatment petroleum soil. Using DNA as template, polymerase chain reaction (PCR) were executed using8gene-controlled primers. And agarose gel electrophoresis are used to determine genes. According the results, In treatment3#,4#and1#, the chain alkane monooxygenase (primer AlkB2F/AlkB2R) gene, alkane hydroxylase alkB (III)(alk-3F/alk-3R) gene, the alpha subunit of naphthalene dioxygenase Fe-S protein(primer nahAcfor/nahAcrev) genens increased significantly. However, in the treatment1#(control experiment) these genes was not too much. And the numbers of genes are positively correlated with TPH degradation efficiency.(4) Soil are sampling in lwk,2wks,3wks,5wks,7wks,9wks,11wks in different treatment soil. The number of TPH-degrading strains, the number of alkane-degrading strain, and polycyclic aromatic hydrocarbon degrading strains were determined using most probable number (MPN). The results shown that in treatment3#, treatment4#, and treatment2#, the number of degrading strains are increase significantly. but the number of degrading strains are not positively correlated with TPH degradation efficiency. So the number of genes is critical factor towards TPH biodegradation. |
PDF Full Text Request