DEVELOPMENT OF A PCR-BASED ASSAY FOR THE DETECTION OF CHIKEN FECES

本子项目是利用由NIH/NCRR资助的中心赠款提供的资源的众多研究子项目之一。子项目和研究者（PI）可能已经从另一个NIH来源获得了主要资金，因此可以在其他CRISP条目中表示。列出的机构是中心的，不一定是研究者的机构。水质和安全是主要的公共卫生问题。我们家庭和娱乐场所的水应该是安全的，可以饮用或游泳，并且相对不含包括粪便污染物在内的污染物。许多人类疾病，如霍乱、伤寒、沙门氏菌病和痢疾，可通过粪便污染的水传播。本研究项目的目的是利用拟杆菌目粪便厌氧细菌开发一种基于pcr的鸡粪便污染检测方法。先前的研究表明，存在于不同物种消化道或具有共同特征的一组物种（例如反刍动物）中的拟杆菌属菌株在遗传上是不同的。基于16S rRNA基因序列差异的检测方法已经用于鉴定人类和反刍动物粪便污染，但是还没有一种将鸡粪便污染与其他类型的粪便污染区分开来的检测方法。因此，本项目的重点是确定鸡粪便特异性的遗传标记，并建立基于pcr的鸡粪便污染检测方法。该试验将使用其他动物的粪便DNA样本进行特异性测试。将对含有已知粪便污染物的水样进行分析，并对从当地流域采集的水样进行测试，从而进一步检验该化验方法。最后，将测试该测定法对不同程度粪便污染的定量能力和灵敏度极限（上限和下限）。本研究的长期目标是开发和实施快速、特异、敏感和廉价的分子方法来确定水样中粪便污染的来源。

This subproject is one of many research subprojects utilizing the resources provided by a Center grant funded by NIH/NCRR. The subproject and investigator (PI) may have received primary funding from another NIH source, and thus could be represented in other CRISP entries. The institution listed is for the Center, which is not necessarily the institution for the investigator. Water quality and safety are major public health issues. The water in our homes and recreational areas is expected to be safe to drink or swim in and to be relatively free of pollutants including fecal pollutants. Many human diseases such as cholera, typhoid, salmonellosis, and dysentery can be transmitted through fecal contamination of water. The goal of this research project is to develop a PCR-based assay for chicken fecal contamination using fecal anaerobic bacteria from the order Bacteroidales. Previous studies have shown that Bacteroidales strains present in the digestive tracts of different species, or in a group of species sharing a common characteristic (e.g. ruminants), are genetically distinct. Assays based on differences in 16S rRNA gene sequences have already been developed for the identification of human and ruminant fecal contamination, but an assay to distinguish chicken fecal contamination from other types of fecal pollution is not yet available. Therefore, the focus of this project is to identify a genetic marker specific for chicken feces and develop a PCR-based assay for chicken fecal contamination. The assay will be tested for specificity using fecal DNA samples from other animals. The assay will be further tested by analysis of water samples containing known fecal contaminants and by testing water samples taken from local watersheds. Finally, the assay will be tested for its ability to quantitate different levels of fecal contamination and for limits of sensitivity (upper and lower). The long-term objective of this research is to develop and implement rapid, specific, sensitive, and inexpensive molecular methods for the determination of the source or sources of fecal contamination in water samples.