The health and well-being of cattle is an important issue in maintaining and increasing global agricultural output. In dairy production within low and middle income countries (LMICs), there is a significant biosensing challenge in detecting sexually transmitted infection (STI) pathogens during animal husbandry, due in part to difficulties associated with the limited infrastructure for veterinary medicine. Here we demonstrate low-cost, multiplexed, and sample-to-answer paper-origami tests for the detection of three bovine infectious reproductive diseases in semen samples, collected at a test site in rural India. Pathogen DNA from one viral pathogen, bovine herpes virus-1 (BoHV-1), and two bacteria (Brucella and Leptospira) was extracted, amplified (using loop-mediated isothermal amplification, LAMP), and detected fluorescently, enabling <1 pg (∼ from 115 to 274 copies per reaction) of target genomic DNA to be measured. Data was collected as a fluorescence signal either visually, using a low-cost hand-held torch, or digitally with a mobile-phone camera. Limits of detection and sensitivities of the paper-origami device for the three pathogens were also evaluated using pathogen-inoculated semen samples and were as few as 50 Leptospira organisms, 50 CFU Brucella, and 1 TCID₅₀ BoHV-1. Semen samples from elite bulls at a germplasm center were also tested in double-blind tests, as a demonstrator for a low-cost, user-friendly point-of-care sensing platform, for in-the-field resource-limited regions. The sensors showed excellent levels of sensitivity and specificity, and for the first time a demonstrated ability of the application of paper microfluidics devices for the diagnosis multiple infectious diseases from semen samples.

中文翻译：

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使用纸折DNA DNA微流控技术快速诊断精液中的牛生殖传染病

牛的健康和福祉是维持和增加全球农业产量的重要问题。在低收入和中等收入国家（LMIC）的乳制品生产中，畜牧业在检测性传播感染（STI）病原体方面存在重大的生物传感挑战，部分原因是与兽医基础设施有限相关的困难。在这里，我们演示了低成本，多路复用和从样本到答案的纸折纸测试，用于检测在印度农村的一个测试地点收集的精液样本中的三种牛传染性生殖疾病。来自一种病毒病原体，牛疱疹病毒1（BoHV-1）和两种细菌（布鲁氏菌和钩端螺旋体）的病原体DNA提取，扩增（使用环介导的等温扩增，LAMP），并进行荧光检测，可测量目标基因组DNA <1 pg（每个反应约115至274拷贝）。使用低成本的手持式手电筒以可视方式或通过移动电话摄像头以数字方式收集数据作为荧光信号。还使用病原体接种的精液样本评估了纸折纸装置对三种病原体的检测极限和敏感性，其检出限为50种钩端螺旋体生物，50种CFU布鲁氏菌和1种TCID ₅₀BoHV-1。来自种质中心的精英公牛的精液样品也进行了双盲测试，作为低成本，用户友好型即时医疗点传感平台的演示者，用于现场资源有限的地区。传感器显示出极高的灵敏度和特异性水平，并首次证明了纸微流控设备在诊断精液样本中多种传染性疾病中的应用能力。