Baertsch Carolina, Paez-Rubio Tania, Viau Emily, Peccia Jordan
Department of Civil and Environmental Engineering, Arizona State University, Arizona 85287, USA.
Appl Environ Microbiol. 2007 Jul;73(14):4522-31. doi: 10.1128/AEM.02387-06. Epub 2007 May 18.
DNA-based microbial source tracking (MST) methods were developed and used to specifically and sensitively track the unintended aerosolization of land-applied, anaerobically digested sewage sludge (biosolids) during high-wind events. Culture and phylogenetic analyses of bulk biosolids provided a basis for the development of three different MST methods. They included (i) culture- and 16S rRNA gene-based identification of Clostridium bifermentans, (ii) direct PCR amplification and sequencing of the 16S rRNA gene for an uncultured bacterium of the class Chloroflexi that is commonly present in anaerobically digested biosolids, and (iii) direct PCR amplification of a 16S rRNA gene of the phylum Euryarchaeota coupled with terminal restriction fragment length polymorphism to distinguish terminal fragments that are unique to biosolid-specific microorganisms. Each method was first validated with a broad group of bulk biosolids and soil samples to confirm the target's exclusive presence in biosolids and absence in soils. Positive responses were observed in 100% of bulk biosolid samples and in less than 11% of the bulk soils tested. Next, a sampling campaign was conducted in which all three methods were applied to aerosol samples taken upwind and downwind of fields that had recently been land applied with biosolids. When average wind speeds were greater than 5 m/s, source tracking results confirmed the presence of biosolids in 56% of the downwind samples versus 3% of the upwind samples. During these high-wind events, the biosolid concentration in downwind aerosols was between 0.1 and 2 microg/m3. The application of DNA-based source tracking to aerosol samples has confirmed that wind is a possible mechanism for the aerosolization and off-site transport of land-applied biosolids.
基于DNA的微生物源追踪（MST）方法被开发出来并用于在大风事件期间特异性且灵敏地追踪土地施用的厌氧消化污水污泥（生物固体）的意外气溶胶化。对大量生物固体进行培养和系统发育分析为三种不同的MST方法的开发提供了基础。它们包括：（i）基于培养和16S rRNA基因对双发酵梭菌进行鉴定；（ii）对厌氧消化生物固体中常见的未培养绿弯菌门细菌的16S rRNA基因进行直接PCR扩增和测序；（iii）对广古菌门的16S rRNA基因进行直接PCR扩增并结合末端限制性片段长度多态性，以区分生物固体特异性微生物特有的末端片段。每种方法首先用大量生物固体和土壤样本进行验证，以确认目标物在生物固体中独家存在且在土壤中不存在。在100%的大量生物固体样本中观察到阳性反应，而在所测试的大量土壤样本中阳性反应率不到11%。接下来，开展了一项采样活动，将所有三种方法应用于最近施用了生物固体的田地的上风向和下风向采集的气溶胶样本。当平均风速大于5米/秒时，源追踪结果证实56%的下风向样本中存在生物固体，而上风向样本中这一比例为3%。在这些大风事件期间，下风气溶胶中的生物固体浓度在0.1至2微克/立方米之间。将基于DNA的源追踪应用于气溶胶样本已证实，风是土地施用生物固体气溶胶化和场外运输的一种可能机制。
