Cultivation-Dependent and –Independent Approaches for Determining Microbial Community in Petroleum-Contaminated Soil

• Main Authors: Hsien-Ming Chiu, 邱憲明
• Other Authors: I-cheng Tseng
• Format: Others
• Language: zh-TW
• Published: 2004
• Online Access: http://ndltd.ncl.edu.tw/handle/68262721939291210276

碩士 === 國立成功大學 === 生物學系碩博士班 === 92 === 　　Petroleum hydrocarbons are the most widespread contaminants in the environment. Several of petroleum aromatic hydrocarbons (PAHs) are known mutagens or carcinogens for human and other organisms and they are difficult to be degraded in the natural environment. Bioremediation is considered an useful method to degrade the PAHs using the PAH-degrading bacteria. In this study, polyphasic approach combined cultivation-dependent with cultivation-independent methods was used to study the microbial communities in petroleum-contaminated soil. A total of 7 strains were isolated and 71% (5 strains) of which were oil-degrading bacteria that belong to Pseudomonas (2 strain)、Bacillus (1 strain)、Mycobacterium (1 strain)、Acinetobacter (1 strain). Phylogenetic analysis of a bacterial 16S rDNA clone library from the petroleum-contaminated soil treated with BTEX as carbon source showed that 9 of 24 OTUs (operational taxonomic units) were the oil-degrading bacteria which belong to Pseudomonas (30.5%)、Burkholderia (24.5%)、Sphingomononas (6.0%)、Bacillus (2.5%) and Nocardioides (0.5%). The results of combined cultivation-dependent with cultivation-independent methods revealed the majority of the oil-degrading bacteria in soil were Pseudomonas. Functional gene (xylE gene；catechol 2,3-dioxygenase) primer was designed in this study could get better and more PCR product from pure bacterium strain and environmental samples than the DEG primers used by Mesarch et al.(2000). The change of the microbial community in petroleum-contaminated soil was investigated by denaturing gel electrophoresis (DGGE) approach. The results indicated that different microbial community structure depended on the soil source and depth. After adding BTEX as carbon source into the petroleum-contaminated soil through one to three weeks, the bacterial number became less and community structure changed. It indicated that BTEX have adverse effects to microbial community structure.