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

南亚的水产养殖业，特别是印度的对虾养殖业一直在快速增长，目前在减轻低收入区域和社区的贫困方面具有很大潜力。在这些环境中，养殖场通常很小(印度90%的养虾户使用2公顷土地)，由贫困家庭经营。然而，它们可以通过改善就业、增加收入(例如，通过与该行业及其供应链相关的工作以及通过出口)，为大部分当地人口带来好处。这些好处可以在当地惠及更广泛的社区，从而减轻贫困。然而，这些好处通常没有实现，因为这些农场的生产是不可持续的，主要是由于传染病爆发的频率和影响很高，这些疾病摧毁了农作物，产量增长缓慢，对农场本身以及更广泛的社区和经济造成了重大影响。在国家层面上，这意味着每年约50,000公吨(MT)损失(约合GB 1B)和215万个工日的就业损失。在这个项目中，我们将转换一个新颖、低成本、电池供电的手持平台，用于在农场或孵化场早期、多路检测病原体，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Ananda(一家历史悠久的海鲜公司，总部设在安得拉邦的印度对虾养殖中心)、ICAR-中央水产技术研究所(在科钦)、莫洛奇(一家快速诊断测试提供商)、Eigem(一家低成本设备制造商)和环境、渔业和水产养殖科学中心(英国海洋科学和技术的世界领先者)合作，设计、验证和部署印度现场测试。目前在印度和南亚其他地方使用的诊断测试是基于核酸的检测，这种检测需要几个小时才能在资源充足的实验室使用复杂的设备进行。这有两个主要后果：(I)贫穷的农民(大多数)根本得不到检测，或者检测的频率不足以可靠地监测生产并及时发现潜在的疫情；(Ii)在进行检测时(例如在阿南达)，检测要求将样本运送到有专门知识的地方，这导致结果严重延迟(往往是好几天)。这些拖延可能导致不当的不采取行动(并有可能失去作物)、紧急收获(导致经济产量下降)，或者在某些情况下，滥用抗生素或化学治疗，这可能会增加抗药性并污染环境。在这个项目中，我们将使农民的能力发生阶段性变化，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相结合的方式从虾组织中浓缩病原体，并将其添加到纸基生物传感器上。通过折叠纸张，在类似于折纸的过程中，病原体的遗传物质被提纯并分布到特定的区域，在那里核酸被放大。这种放大(使用60摄氏度左右的低功率手持加热器进行，也可以在保温瓶中进行)然后在45分钟内通过直接观察颜色变化或使用手机量化反应来检测。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行动，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.