Detection and quantification of Geobacter lovleyi strain SZ:: Implications for Bioremediation at tetrachloroethene- and uranium-impacted sites
Detection and quantification of Geobacter lovleyi strain SZ:: Implications for Bioremediation at tetrachloroethene- and uranium-impacted sites
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DOI：
10.1128/aem.01218-07
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发表时间：
2007-11-01
影响因子：
4.4
通讯作者：
Loeffler, Frank E.
中科院分区：
文献类型：
--
作者：
Amos, Benjamin K.;Sung, Youlboong;Loeffler, Frank E.
洛夫勒地杆菌SZ是金属还原细菌群中第一个能利用四氯乙烯(PCE)作为生长支持电子受体的菌株。以16S rRNA基因为靶点的直接和套式聚合酶链式反应将菌株SZ与其他已知的氯化乙烯脱氯细菌和密切相关的地质杆菌分离株区分开来，包括其最近的培养亲戚G.thigenes。直接聚合酶链式反应和套式聚合酶链式反应的检测下限分别约为1×106和1×10(4)16S rRNA基因拷贝数。一种定量实时聚合酶链式反应(QPCR)方法提高了对每亩模板DNA中只有30个16S rRNA基因拷贝的敏感性，但特异性较低。熔解曲线分析和扩增图谱形状的比较鉴定了假阳性信号，并在单独分析时区分了菌株SZ和硫代革兰氏菌。当目标(菌株SZ)DNA与序列相似性较高的非目标(G.thigenes)DNA混合在一起时，这些指标的可靠性较低，这表明qPCR方法的发展不仅要评估特异性，还应探索非目标DNA对定量准确性的影响。特定工具在四氯乙烯脱氯联合体(包括生物增强联合体KB-1和两个受氯化乙烯影响的地下水样本)中检测到菌株SZ样扩增的应用。在田纳西州橡树岭的铀和氯化溶剂污染的综合野外规模地下研究挑战(IFC)场地进行生物刺激后，在22个地下水样本中也检测到了SZ菌株样扩增片段。每升地下水中类SZ菌的数量从未检测到的增加到2.3×10(7)+/-0.1×10(7)，表明类SZ菌对污染物的转化起到了促进作用。G.lovleyi菌株SZ专用工具将有助于监测受铀和/或氯化溶剂影响的地点的生物修复工作，例如橡树岭IFC地点。
Geobacter lovleyi strain SZ reduces hexavalent uranium, U(VI), to U(IV) and is the first member of the metal-reducing Geobacter group capable of using tetrachloroethene (PCE) as a growth-supporting electron acceptor. Direct and nested PCR with specific 16S rRNA gene-targeted primer pairs distinguished strain SZ from other known chlorinated ethene-dechlorinating bacteria and closely related Geobacter isolates, including its closest cultured relative, G. thiogenes. Detection limits for direct and nested PCR were approximately 1 x 106 and 1 x 10(4) 16S rRNA gene copies per mu I of template DNA, respectively. A quantitative real-time PCR (qPCR) approach increased the sensitivity to as few as 30 16S rRNA gene copies per mu I of template DNA but was less specific. Melting curve analysis and comparison of the shapes of amplification plots identified false-positive signals and distinguished strain SZ from G. thiogenes when analyzed separately. These indicators were less reliable when target (strain SZ) DNA and nontarget (G. thiogenes) DNA with high sequence similarity were mixed, indicating that the development of qPCR protocols should not only evaluate specificity but also explore the effects of nontarget DNA on the accuracy of quantification. Application of specific tools detected strain SZ-like amplicons in PCE-dechlorinating consortia, including the bioaugmentation consortium KB-1, and two chlorinated ethene-impacted groundwater samples. Strain SZ-like amplicons were also detected in 13 of 22 groundwater samples following biostimulation at the uranium- and chlorinated solvent-contaminated Integrated Field-Scale Subsurface Research Challenge (IFC) site in Oak Ridge, TN. The numbers of strain SZ-like cells increased from below detection to 2.3 x 10(7) +/- 0.1 x 10(7) per liter groundwater, suggesting that strain SZ-like organisms contribute to contaminant transformation. The G. lovleyi strain SZ-specific tools will be useful for monitoring bioremediation efforts at uranium- and/or chlorinated solvent-impacted sites such as the Oak Ridge IFC site.