Mitochondrial DNA in the environment: towards the development of better animal-specific markers for fecal source tracking

许多来源可能会导致水和土壤的粪便污染。由于许多水生致病微生物都与粪便污染有关，因此在水质评估中监测此类污染对公众健康具有重要意义。事实上，地表水和地下水的粪便污染会对公众健康(饮用水)和当地经济(农业和娱乐活动)产生重大影响。水的微生物质量主要是使用粪便指示性生物，如大肠菌群进行监测。然而，这些指标并不能确定粪便物质的来源。已经开发了几种粪便来源追踪(FST)方法来解决宿主对粪便污染的特定贡献的问题。用聚合酶链式反应方法检测的宿主特异性遗传标记表明存在来自特定动物物种的粪便污染。这些遗传标记大多依赖于肠道(粪便)物质中与特定动物宿主相关的微生物的存在。尽管数量众多，但这些标记物都不是完全、足够特异和敏感的。因此，开发新的遗传标记和创新的FST方法是非常必要的。 最近开发的FST遗传标记依赖于肠腔内结肠上皮细胞的重要释放。这些细胞含有大量的线粒体及其基因组(MtDNA)。在本世纪初，我的团队是第一个开发基于线粒体DNA的FST标记的团队；这种标记现在被其他团队使用。线粒体DNA序列具有高度的物种特异性；在亲缘关系密切的物种之间可以发现线粒体DNA的显著差异。在这些序列变异的基础上，我们设计了可用于聚合酶链式反应的物种特异性引物。我们和其他人已经证明，来自人、牛和猪的线粒体DNA存在于未经处理的生活污水、农业土壤径流、养猪场污水和来自已知污染源的河流的水中，并可检测到。尽管在环境样本中成功地检测到线粒体DNA，但在环境水和土壤中检测这种标记可能是具有挑战性的。这个问题源于我们对环境中线粒体DNA动态的缺乏了解。 这项研究计划的总体目标是表征线粒体DNA在不同水和土壤环境中的命运，最终目标是开发一种改进的线粒体DNA检测方法作为一种可靠的FST方法。该研究计划的目标将解决动物特有的mtDNA标记在各种环境情况下的命运和运输问题，这些环境情况在实验室环境中以及在试验田中得到很好的控制。开发经改进的动物特异性标记以确定排泄物的确切来源，对于实施有效的预防或补救策略至关重要。

Numerous sources can contribute to fecal contamination of water and soil. As many waterborne pathogenic microorganisms are associated with fecal pollution, monitoring such contamination in water quality assessments is of great importance for public health. Indeed, fecal pollution of surface water and groundwater can have a great impact on public health (drinking water) and local economy (agriculture and recreational activities). The microbiological quality of water is primarily monitored using fecal indicator organisms, such as coliforms. However, these indicators do not identify the source of the fecal matter. Several fecal source tracking (FST) methods have been developed to address the issue of host-specific contributions to fecal contamination. Host-specific genetic markers detected by PCR methods indicate the presence of fecal contamination from specific animal species. Most of these genetic markers rely on the presence of microorganisms in the intestinal (fecal) matter that are associated with specific animal host. Despite their number, none of these markers are completely and sufficiently specific and sensitive. Therefore, it is essential to develop novel genetic markers and innovative methods for FST. Recently developed FST genetic markers rely on the important release of colonic epithelium cells in the intestinal lumen. These cells contain numerous copies of mitochondria with their genome (mtDNA). In the early 2000s, my team has been the first group to develop mtDNA-based FST markers; such markers are now used by other groups. mtDNA sequences are highly species-specific; substantial differences in mtDNA can be found between closely related species. Building on these sequence variations, we designed species-specific primers that can be used for PCR. We and others have demonstrated that mtDNA from human, bovine and porcine are present and detectable in untreated domestic sewage, agricultural soil runoff, swine farm effluents and water from rivers with known pollution sources. Despite the successful detection of mtDNA in environmental samples, the detection of such markers can be challenging in environmental water and soil. This problem stems from our lack of knowledge on the dynamics of mtDNAs in the environment. The overall aim of this research program is to characterize the fate of mtDNAs in various water and soil environments with the ultimate goal of developing improved mtDNA detection as a robust FST method. The aims of the research program will address the fate and transport of animal-specific mtDNA markers in various environmental situations well controlled in laboratory settings as well as in experimental field plots. The development of improved animal-specific markers identifying the exact sources of fecal matter is essential for implementing effective preventive or remedial strategies.