Paper-based platform for onsite, rapid and multiplexed pathogen detection in shrimp farms

南亚的水产养殖业，特别是印度的养虾业，一直在迅速增长，现在具有减轻低收入地区和社区贫穷的巨大潜力。在这些环境中，养殖场通常很小（印度90%的虾农使用不到2公顷的土地），由贫困家庭经营。然而，它们可以通过改善就业、增加收入（来自与该行业及其供应链有关的工作，以及通过出口等），为很大一部分当地人口带来好处。这些利益可以流向更广泛的社区，在当地减轻贫困。然而，这些利益通常无法实现，因为这些农场的生产不可持续，主要是由于传染病爆发的频率和影响，这些传染病破坏作物并导致生长缓慢，对农场本身以及更广泛的社区和经济造成重大影响。在国家一级，这意味着。50，000公吨（MT）损失（约在这个项目中，我们将翻译一种新颖的、低成本的、电池供电的手持平台，用于在农场或孵化场对病原体进行早期、多重检测，使当地农民和社区能够控制和减少疾病爆发的发生和影响。我们将与我们的合作伙伴阿难（一家历史悠久的海产品公司，总部设在安得拉邦的印度虾养殖中心），ICAR-中央渔业技术研究所，（in Cochin），Mologic（快速诊断测试提供商），Epigem（一家低成本设备制造商）和Cefas环境、渔业和水产科学中心（英国海洋科学和技术的世界领导者），设计，验证和部署在印度的实地测试。目前在印度和南亚其他地方使用的诊断测试是基于核酸检测，在资源充足的实验室中使用先进的设备需要几个小时才能完成。这有两个主要后果：（i）贫穷农民无法获得测试（大多数），要么根本不进行，要么以能够可靠地监测生产和及时发现潜在疫情的频率进行;（ii）在进行（例如在阿南达）时，要求将样本运送到有专门知识的地方，这导致结果的重大延误（往往是许多天）。这些延误可能导致不合时宜的不作为（并可能导致作物损失），紧急收割（导致经济产出下降），或者在某些情况下，滥用抗生素或化学治疗，这有可能增加耐药性并污染环境。在这个项目中，我们将逐步改变农民的能力，以减少生产损失的影响和与无效治疗相关的负担。该平台依赖于使用纸基过滤和磁珠的组合从虾组织中富集病原体到纸基生物传感器上。通过折叠纸，在类似于折纸的过程中，病原体的遗传物质被纯化并分布到特定区域，在那里核酸被扩增。这种扩增（使用小型手持加热器进行，在60 ℃左右的低功率，但也可以在保温瓶中进行）然后使用直接观察颜色变化（如在妊娠测试中）或使用移动的电话在45分钟内量化反应来检测。通过开发和部署低成本的使用点检测设备，我们将使小规模农民能够及早识别病原体，从而使他们能够采取补救措施，并与社区联系，以确保疫情得到控制。

The aquaculture industry in South Asia, and in particular shrimp farming in India, has been growing rapidly and now has significant potential to alleviate poverty within low-income regions and communities. In these settings, farms are often small (90% of the shrimp farmers in India use <2ha of land) and are run by poor families. However, they can generate benefits across a large portion of the local population, through improved employment, increased income (from jobs associated with the industry and its supply-chain, as well as through export for example). These benefits can flow to wider community, locally, alleviating poverty. However, these benefits are generally not realised, as the production from these farms is not sustainable, mainly due to the high frequency and impact of outbreaks of infectious diseases, which destroy crops and yield slow growth, resulting in significant impact on the farms themselves but also on the wider communities and economy. At a national level, this translates into ca. 50,000 Metric Tonne (MT) losses (ca. £1B) and loss of employment of 2.15 million man days per annum.In this project, we will translate a novel, low-cost, battery-powered handheld platform for the early, multiplexed detection of pathogens at the farm or hatchery, to empower local farmers and communities to control and reduce the occurrence and impact of disease outbreaks. We will work with our partners Ananda (a well-established, seafood company, based in the hub of Indian shrimp farming in Andhra Pradesh), ICAR-Central Institute of Fisheries Technology, (in Cochin), Mologic (a provider of rapid diagnostic tests), Epigem (a manufacturer of low-cost devices) and the Centre for Environment, Fisheries and Aquaculture Sciences Cefas (aworld leader in marine science and technology in the UK), to design, validate and deploy the tests in the field in India. The current diagnostic tests, which are used in India and elsewhere in South Asia, are based on nucleic acid-based detection, which takes a few hours to perform, in a well-resourced laboratory, using sophisticated equipment. This has two major consequences: (i) the tests are not available to poor farmers (the majority), either at all, or at a frequency that would allow to monitor the production reliably and detect potential outbreaks timely; and (ii) when they are carried out (e.g. at Ananda), they require the samples to be transported to where expertise is available, which leads to significant delays in results (often many days). These delays can result in mis-timed inaction (and the potential to lose the crop), emergency harvesting (leading to decreased economic output), or, in some cases, the mis-use of antibiotic or chemical treatment, which has the potential to increase drug resistance and contaminate the environment. In this project, we will deliver a step-change in the farmer's abilty to reduce the impact of production-based losses and the burden associated with ineffective treatments. The platform relies upon the enrichment of pathogens from shrimp tissue using a combination of paper-based filtration and magnetic beads, onto a paper-based biosensor. By folding the paper, in a process akin to origami, the genetic material of the pathogens is purified and distributed into specific areas, where nucleic acids are amplified. This amplification (performed using a small hand-held heater, with low power at around 60oC, but which could also be performed in a thermos flask) is then detected using either direct visualisation of a colour change, as in pregnancy tests, or using a mobile phone to quantify the response, within 45min. By developing and deploying a low-cost, point-of-use detection device, we will empower small-scale farmers with the ability to identify pathogens early, thus allowing them to take remedial actions as well as liaise with the community to ensure outbreaks are contained.