Isolation,Identification Of Phenanthrene-Degradading Bacteria And Its Application In Remedyin Phenanthrene Polluted Soil

Polycyclic aromatic hydrocarbons (PAHs), a class of POPs, which consist of two or more fused aromatic rings, are widely distributed in the environment and persist over long periods of time.Most of PAHs are toxic, mutagenic, and carcinogenic and threrfore are health hazards.Phenanthrene is one of three-aromatic PAHs and distributes in environments widely, which cause diseases of human respiration system and skin, and induces mutation, aberrance and carcinogen of fish and other animals. Phenanthrene has the special structure K-region and Bay-region which can arise carcinogens. Therefore, it has been taken as a model compound for studies on PAHs. Work on phenanthrene biodegradation not only is beneficial to eliminate the contamination of phenanthrene from environments, but also can be valuable to understand the biodegradation mechanism and feasibility of other PAHs, providing theorically and practically knowledge and control measurements for bioremediation of soil and other environment contaminated by PAHs.As the industry is developing, PAHs pollution becomes worse. Soil contamination with PAHs poses a great threat worldwide to the agricultural food quality and human health, and calls for an immediate action to remediate the contaminated sites. Microbial-Phytoremediation has long been recognized as a cost-effective method for removal of PAHs pollutants from soil.In this paper, a microbial consortium which could degrade phenanthrene and take phenanthrene as the sole carbon and energy source were obtained from an aged PAHs-contaminated soil by selective enrichment liquid culture. The microbial consortium could metabolise99%of phenanthrene within7days in a mineral medium containing100mg·L-1phenanthrene. There phenanthrene-degrading microorganisms, B1、J1and L2were isolated from the consortium. Based on Physiological properties and16S rDNA, those three strains were identified as Bordetella petrii, Pseudoxanthomonas mexicana and Achromobacter xylosoxidans, respectively. The degradation rate of phenanthrene in the mineral medium was about80%by Bordetella petrii Bl within7days and99%by Pseudoxanthomonas Mexicana L2within9days. It proved that the strains were high effectient degradation strain to phenanthrene. Phenanthrene concentration is determined using HPLC.The microbial consortium also can utilize fluorine, acenaphthylene, pyrene, naphthanlene and indole as carbon source besides phenanthrene. Meanwhile, the microbial consortium can degradate part of pyrene, fluoranthene and phenol at the same time of metabolizing phenanthrene. While methylbenzene did harm to its growth. As co-metabolism substrates, salicylic acid, potassium acid phthalate, glucose and yeast extract promoted phenanthrene degradation, while vegetable oil did the opposite thing.The results indicated that the ability of phenanthrene-degrading increase greatly when the three strains were used as a mixture. Basd on the results of plate culture and microscopy, comparing the degradation rate between the microbial consortium and mixing strains, it showed that srains B1J1and L2were the dominant bacterias in the microbial consortium, playing the key role in degradating phrenanthrene.The characteristics of strain B1and L2growth and phenanthrene degradation were investigated. The optimal conditions for both growth and degradation process is as follows:temperature28℃. pH7.0.The bacterial growth was inhibited when pH was4or temperature was12℃. Cr3+(20mg-L-1) was toxic to the both strains. Cu2+(50mg·L-1) could partially inhibit the growth of strain B1, Zn2+(50mg·L-1) and Pb2+(200mg·L-1) had no significant influence on the bacterial growth.Effects of microbial consortium on the growth of ryegrass and phenanthrene degradation in PAHs contaminated soils were investigated by pot culture experiment. Results showed that in phenanthrene contaminated soil, both ryegrass and the microbial consortium can cause phenanthrene decrease in the soil. But the biomass of ryegrass with microbial consortium inoculated shows no siganificant difference from that of non-inoculated treatments. By using the technology of microbial-phytoremediation, the residual concentration of phenanthrene in phenanthrene contaminated soil (initial concentration were8.22mg-kg-1and39.6mg-kg-1, respectively) was0.32mg-kg-1and0.59mg-kg-1, while the residual concentration was0.88mg-kg-1and1.11mg·kg-1by using phytoremediation. After88days of soil remediation by the microbial consortium, the degradation rate of phenanthrene reached up to95.74%for the soil containing8.22mg·kg-1of phenanthrene and98.06%for the soil containing39.6mg-kg-1of phenanthrene.With the plate count method for monitoring the remediation process of bacteria, fungi and actinomycetes number and types of changes in micro-organisms, it was found in inverse proportion to the concertration of phenanthrene. In the repair process of artificial contaminated soil, microorganism of two different periods of changes in the structure was monitored in the DGGE patterns, the bacteria B1、J1and L2of the characteristic bands were throughout the repair process. Of different treatments, the microbial community structure differences are significant, agreed to the trends and plate count results.While using microbial consortium to remediation natural PAHs polluted soil, the phenanthrene concentration fell down quickly from4.74mg·kg-1to0.26mg·kg-1within9days. The results showed that microbial remediation was operable to PAHs contaminated soil.