Hristova K R, Lutenegger C M, Scow K M
Department of Land, Air, and Water Resources, University of California, Davis, California 95616, USA.
Appl Environ Microbiol. 2001 Nov;67(11):5154-60. doi: 10.1128/AEM.67.11.5154-5160.2001.
The fuel oxygenate methyl tert-butyl ether (MTBE), a widely distributed groundwater contaminant, shows potential for treatment by in situ bioremediation. The bacterial strain PM1 rapidly mineralizes and grows on MTBE in laboratory cultures and can degrade the contaminant when inoculated into groundwater or soil microcosms. We applied the TaqMan quantitative PCR method to detect and quantify strain PM1 in laboratory and field samples. Specific primers and probes were designed for the 16S ribosomal DNA region, and specificity of the primers was confirmed with DNA from 15 related bacterial strains. A linear relationship was measured between the threshold fluorescence (C(T)) value and the quantity of PM1 DNA or PM1 cell density. The detection limit for PM1 TaqMan assay was 2 PM1 cells/ml in pure culture or 180 PM1 cells/ml in a mixture of PM1 with Escherichia coli cells. We could measure PM1 densities in solution culture, groundwater, and sediment samples spiked with PM1 as well as in groundwater collected from an MTBE bioaugmentation field study. In a microcosm biodegradation study, increases in the population density of PM1 corresponded to the rate of removal of MTBE.
燃料含氧化合物甲基叔丁基醚（MTBE）是一种广泛分布于地下水中的污染物，显示出原位生物修复的处理潜力。细菌菌株PM1在实验室培养物中能迅速将MTBE矿化并以其为生长底物，接种到地下水或土壤微观体系中时能够降解该污染物。我们应用TaqMan定量PCR方法来检测和定量实验室及现场样品中的菌株PM1。针对16S核糖体DNA区域设计了特异性引物和探针，并用15种相关细菌菌株的DNA确认了引物的特异性。在阈值荧光（C(T)）值与PM1 DNA量或PM1细胞密度之间测定了线性关系。PM1 TaqMan检测法在纯培养中的检测限为2个PM1细胞/毫升，在PM1与大肠杆菌细胞的混合物中的检测限为180个PM1细胞/毫升。我们能够测定接种了PM1的溶液培养物、地下水和沉积物样品以及从MTBE生物强化现场研究采集的地下水中的PM1密度。在一项微观体系生物降解研究中，PM1种群密度的增加与MTBE的去除速率相对应。
