Horst Audrey L, Rosenbohm Justin M, Kolluri Nikunja, Hardick Justin, Gaydos Charlotte A, Cabodi Mario, Klapperich Catherine M, Linnes Jacqueline C
Department of Biomedical Engineering, Boston University, Boston, MA, USA.
School of Medicine, Johns Hopkins University, Baltimore, MD, USA.
Biomed Microdevices. 2018 Apr 11;20(2):35. doi: 10.1007/s10544-018-0280-x.
Globally, the microbe Neisseria gonorrhoeae (NG) causes 106 million newly documented sexually transmitted infections each year. Once appropriately diagnosed, NG infections can be readily treated with antibiotics, but high-risk patients often do not return to the clinic for treatment if results are not provided at the point of care. A rapid, sensitive molecular diagnostic would help increase NG treatment and reduce the prevalence of this sexually transmitted disease. Here, we report on the design and development of a rapid, highly sensitive, paperfluidic device for point-of-care diagnosis of NG. The device integrates patient swab sample lysis, nucleic acid extraction, thermophilic helicase-dependent amplification (tHDA), an internal amplification control (NGIC), and visual lateral flow detection within an 80 min run time. Limits of NG detection for the NG/NGIC multiplex tHDA assay were determined within the device, and clinical performance was validated retroactively against qPCR-quantified patient samples in a proof-of-concept study. This paperfluidic diagnostic has a clinically relevant limit of detection of 500 NG cells per device with analytical sensitivity down to 10 NG cells per device. In triplicate testing of 40 total urethral and vaginal swab samples, the device had 95% overall sensitivity and 100% specificity, approaching current laboratory-based molecular NG diagnostics. This diagnostic platform could increase access to accurate NG diagnoses to those most in need.
在全球范围内，淋病奈瑟菌（NG）每年导致1.06亿例新记录的性传播感染。一旦得到恰当诊断，NG感染可用抗生素轻松治疗，但高危患者如果在即时护理时未得到检测结果，往往不会返回诊所接受治疗。一种快速、灵敏的分子诊断方法将有助于增加NG感染的治疗率并降低这种性传播疾病的流行率。在此，我们报告一种用于NG即时护理诊断的快速、高灵敏度纸质微流控装置的设计与开发。该装置在80分钟的运行时间内集成了患者拭子样本裂解、核酸提取、嗜热解旋酶依赖性扩增（tHDA）、内部扩增对照（NGIC）以及可视化侧向流动检测。在该装置内确定了NG/NGIC多重tHDA检测法的NG检测限，并在一项概念验证研究中针对经qPCR定量的患者样本追溯验证了临床性能。这种纸质微流控诊断方法的临床相关检测限为每个装置500个NG细胞，分析灵敏度低至每个装置10个NG细胞。在对40份尿道和阴道拭子样本进行的三次重复检测中，该装置的总体灵敏度为95%，特异性为100%，接近当前基于实验室的分子NG诊断方法。这种诊断平台可为最需要的人群增加获得准确NG诊断结果的机会。
