Using digital droplet PCR to improve the detection of helminth infections of ruminants in Northern Ireland

肝片吸虫，通常被称为“温带肝吸虫”，在世界各地都有发现，并引起一种称为肝片吸虫病的疾病。这些寄生虫主要是对生产反刍动物的一种破坏性感染。在欧洲，英国、爱尔兰、法国、葡萄牙、西班牙、瑞士、意大利、荷兰、德国和波兰都报告了这种疾病。作为一种食源性疾病，肝片吸虫病是由反刍动物摄取感染幼虫引起的。这些蠕虫极大地降低了全球农业能力和发展中国家和发达社区的产量，导致世界各地数百万反刍动物的生产效率低下。片吸虫病也是一种重要的人畜共患病，被世界卫生组织(WHO)认可。所有的生物都将DNA释放到他们的环境中，这种物质被称为Edna。这种EDNA的检测能够对环境中的物种多样性和丰度进行分类。当存在片状寄生虫时，在它们生命周期的一部分，为了感染蜗牛或动物宿主，它们会暴露在环境中，它们也会留下埃德娜。这一EDNA的改进检测是本项目的基础。虽然这种物质在水、土壤或植物中的含量预计很低，但将使用新的分子方法来克服这些困难。这些方法包括使用第三代技术液滴数字聚合酶链式反应(DdPCR)，它极大地提高了DNA检测的灵敏度和重复性。DdPCR的独特价值在于它是一种完全定量的方法，具有高度的灵敏度和重复性。我们的建议将推动EDNA分析方法的一步改变，以识别蠕虫寄生虫感染风险。这将涉及检查一系列样本类型，包括水、土壤、草和粪便。这些具有挑战性的样本将以高度敏感的ddPCR技术为特征，在收集的样本上提供寄生虫/病媒Edna的全面定量测量。北爱尔兰的绵羊养殖场将在一年多的时间里进行采样，并收集这些新的样本，以寻找疾病的分子指标。这代表了一种侵入性更小、劳动密集度更低的疾病监测方法。在更早地检测寄生虫感染时，可以通过智能牧场管理减少或避免药物(驱虫剂)的使用-根据牧场的年龄、物种和季节轮换动物，而不是感知这些牧场的感染风险。如果农民能够从他们自己的农场获得相关数据，他们将能够更有效地做出这些决定。

Fasciola hepatica, usually termed "the temperate liver fluke", is found worldwide and causes a disease known as fasciolosis. These helminth parasites is primarily a damaging infection of production ruminants. In Europe, the disease has been reported in UK, Ireland, France, Portugal, Spain, Switzerland, Italy, The Netherlands, Germany and Poland. As a food-borne disease, fasciolosis results from consumption of the infective larvae by ruminant animals. These helminths greatly reduce global agricultural capacity and outputs of both developing and developed communities, causing production inefficiency in millions of ruminants worldwide. Fasciolosis is also an important zoonosis, and is acknowledged by the World Health Organization (WHO).All organisms shed DNA into their environment, this material is known as eDNA. The detection of this eDNA enables the classification of species diversity and abundance in an environment. When Fasciola parasites are present, during part of their lifecycle they are exposed in the environment in order to be infectious to the snail or animal host, they also leave eDNA behind. The improved detection of this eDNA is the basis of this project. While low levels of this materials is expected in water or soil, or on plants, new molecular methods will be used to overcome these difficulties.These methods include the use of third generation technology droplet digital PCR (ddPCR) which greatly increases the sensitivity and reproducibility of DNA detection. ddPCR has the distinct value of being a fully quantitative method, and being highly sensitive and reproducible. Our proposal will drive a step-change in the way eDNA analysis is performed for the identification of helminth parasite infection risk. This will be involve examining a range of sample types, including water, soil, grass and faeces. These challenging samples will be characterised by the highly sensitive ddPCR technology to provide comprehensive quantitative measurements of parasite/vector eDNA on collected samples. Sheep farms in Northern Ireland will sampled over one year and these new sample collected to find molecular indicators of disease. This represents a less invasive and potentially less labor intensive approach to disease monitoring.In detecting parasitic infections earlier, the use of drugs (anthelmintics) can be reduced or avoided through intelligent pasture management - rotating animals around pastures according to their age, species and season, relative to the perception of infection risk in those pastures. Farmers would be able to make these decisions more effectively if they had access to relevant data from their own farm.