Division of Biomedical Engineering, School of Engineer, University of Glasgow , Oakfield Avenue, Glasgow G12 8LT, United Kingdom.
FAO/WHO Collaborating Centre for Brucellosis, OIE Brucellosis Reference Laboratory, Department of Bacteriology, Animal & Plant Health Agency , Woodham Lane, Addlestone, Surrey KT15 3NB, United Kingdom.
ACS Sens. 2018 Feb 23;3(2):403-409. doi: 10.1021/acssensors.7b00825. Epub 2018 Feb 5.
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 TCID 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.
牛只的健康和福祉对于维持和增加全球农业产量至关重要。在中低收入国家（LMICs）的乳品生产中，由于兽医医学基础设施有限，因此在畜牧业中检测性传播感染（STI）病原体存在重大生物传感挑战。在这里，我们展示了低成本、多重和即取即检的纸折叠测试，用于检测精液样本中的三种牛传染性生殖疾病，这些样本是在印度农村的一个测试点收集的。从一种病毒病原体（牛疱疹病毒-1（BoHV-1））和两种细菌（布鲁氏菌和钩端螺旋体）中提取、扩增（使用环介导等温扩增，LAMP）并荧光检测病原体 DNA，使 <1 pg（约 115 至 274 个拷贝/反应）的目标基因组 DNA 可以被测量。数据可以通过低成本的手持火炬进行视觉收集，也可以通过手机摄像头进行数字收集。还使用接种病原体的精液样本评估了纸折叠设备对三种病原体的检测限和灵敏度，最低检测限低至 50 个钩端螺旋体、50CFU 布鲁氏菌和 1TCID BoHV-1。在种质中心，还对精英公牛的精液样本进行了双盲测试，作为低成本、用户友好的即时护理传感平台的演示，用于在资源有限的野外地区。该传感器显示出极好的灵敏度和特异性水平，这是首次证明了应用纸微流控器件从精液样本诊断多种传染病的能力。
