Title

The evaluation of PAH, Phenanthrene and Pyrene compounds removal efficiancy using genetically engineered dioxygenas producing pseudomonas putida in Vadose zone pilot

Principal Investigator

Dr. Amir Hossin Mahvi

Abstract

چکیده لاتین One of the most common groups of petroleum pollutants are polycyclic aromatic hydrocarbons which pollute the air, soil, surface and underground waters and sea environments. The purpose of the present study was to investigate the efficiency of aromatic hydrocarbons removal including phenanthrene and pyrene using genetically engineered Pseudomonas putida (P. putida). In this study, the pUC18 vector containing the nahH gene encoding dioxygenase enzyme was built and transferred into E. coli for amplification. After vector purification the recombinant plasmid was transferred again into Pseudomonas putida and bacteria were amplified for transmission to the pilot. Then, the engineered bacteria and normal strains were separately added to pilots containing soil which had adsorbed 500mg/kg phenanthrene and pyrene. Finally, the pilot samples were analyzed once every five days for 3 months and were measured triplicate by HPLC-Agilent1200 device. In this study soil was contaminated to 500mg/kg of oil compounds including phenanthrene and pyrene. The numbers of 11 unglazed clay pots were prepared, each containing 500 grams of soil. Evaporation percentage for phenanthrene and pyrene was 16.43 and 13.25%, respectively. The removal of phenanthrene in autoclaved soil containing engineered Pseudomonas was 94.34%. In the autoclaved soil spiked with genetically engineered microorganisms (GEMs), the pyrene removal rate was 91.89%. The phenanthrene and pyrene removal rate in autoclaved soil containing GEMs were 93.52 and 90.32%, respectively. In autoclaved soil containing phenanthrene and pyrene and GEMs and enriched with chemical fertilizers nitrogen and phosphorus, removal rates were 95.18 and 92.31%, (respectively), that the highest removal rate happened in this situation. In soil containing natural microbial flora and GEMs, phenanthrene and pyrene removal rates were 92.33 and 70.93%, (respectively). In the autoclaved soil containing normal P. putida, phenanthrene and pyrene removal rates were 24.30 and 18.17%, respectively. In soil containing natural flora removal of phenanthrene and pyrene were 22.94 and 29.14%, (respectively). In the autoclaved soil containing kerosene and GEMs, removal rate of phenanthrene and pyrene were 93.53 and 92.15%, (respectively). In the autoclaved soil spiked with normal P. putida and kerosene the removal of phenanthrene and pyrene were 19.98 and 16.78%, (respectively). In soil spiked with natural flora and kerosene and GEMs, removal of phenanthrene and pyrene were 85.85 and 88.47%, respectively. Statistical analysis was conducted by SPSS software (version 20) and One Way ANOVA–Tukey analysis. Statistical analysis results indicated that degradation of phenanthrene and pyrene in soils containing engineered bacteria in comparison with soils with natural flora and normal P. putida was significant (P<0.05) and this analysis represents that the removal of aromatic petroleum hydrocarbon compounds using genetically engineered P. putida was high. The results indicated that engineered P. putida had high efficiency in the removal of PAHs compounds compared with microflora of soil and normal bacteria.

Year

2015