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16S rRNA Gene-Based Detection of Tetrachloroethene-Dechlorinating Desulfuromonas and Dehalococcoides Species

机译:基于16S rRNA基因的四氯乙烯脱氯脱硫尿单胞菌和Dehalococcoides物种的检测。

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摘要

Members of the genera Desulfuromonas and Dehalococcoides reductively dechlorinate tetrachloroethene (PCE) and trichloroethene. Two primer pairs specific to hypervariable regions of the 16S rRNA genes of the Dehalococcoides group (comprising Dehalococcoides ethenogenes and Dehalococcoides sp. strain FL2) and the acetate-oxidizing, PCE-dechlorinating Desulfuromonas group (comprising Desulfuromonas sp. strain BB1 and Desulfuromonas chloroethenica) were designed. The detection threshold of a nested PCR approach using universal bacterial primers followed by a second PCR with the Desulfuromonas dechlorinator-targeted primer pair was 1 × 103 BB1 cells added per gram (wet weight) of sandy aquifer material. Total community DNA isolated from sediments of three Michigan rivers and six different chloroethene-contaminated aquifer samples was used as template in nested PCR. All river sediment samples yielded positive signals with the BB1- and the Dehalococcoides-targeted primers. One chloroethene-contaminated aquifer tested positive with the Dehalococcoides-targeted primers, and another contaminated aquifer tested positive with the Desulfuromonas dechlorinator-targeted primer pair. Restriction fragment analysis of the amplicons could discriminate strain BB1 from other known Desulfuromonas species. Microcosm studies confirmed the presence of PCE-dechlorinating, acetate-oxidizing Desulfuromonas and hydrogenotrophic Dehalococcoides species in samples yielding positive PCR signals with the specific primers.
机译:Desulfuromonas和Dehalococcoides属的成员还原性地将四氯乙烯(PCE)和三氯乙烯脱氯。特异于Dehalococcoides组的16S rRNA基因的高变区的两个引物对(包括Dehalococcoides ethenogenes和Dehalococcoides sp。菌株FL2)和醋酸盐氧化,PCE脱氯的Desulfuromonas组(包含Desulfuromonas sp。BB1和Desulfuroenas)。设计。使用通用细菌引物的嵌套PCR方法的检测阈值,然后使用脱硫磺单胞菌脱氯剂靶向的引物对进行的第二次PCR,是每克(湿重)含沙含水层材料添加1×103 BB1细胞。在巢式PCR中,将从三条密歇根河的沉积物和六种不同的氯乙烯污染的含水层样品中分离出的总群落DNA用作模板。所有的河底沉积物样品均以BB1-和Dehalococcoides靶向引物产生阳性信号。一个被氯乙烯污染的含水层用靶向Dehalococcoides的引物检测为阳性,而另一个被污染的含水层则用脱硫脲类脱氯剂目标的引物对检测为阳性。扩增子的限制性片段分析可以将菌株BB1与其他已知的脱硫门孢菌种区分开。缩影研究证实了样品中存在PCE脱氯,乙酸盐氧化的Desurouromonas和氢营养型Dehalococcoides物种,这些样品可通过特定引物产生阳性PCR信号。

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