Egbo Timothy E, Blancett Candace D, Payne Jackie M, Stefan Christopher P, Minogue Timothy D, Sellers John H, Koehler Jeffrey W
Diagnostic Systems Division, United States Army Medical Research Institute of Infectious Diseases, Fort Detrick, Maryland, 21702, United States of America.
BMC Infect Dis. 2025 Jan 14;25(1):63. doi: 10.1186/s12879-024-09573-w.
Point of need diagnostics provide efficient testing capability for remote or austere locations, decreasing the time to answer by minimizing travel or sample transport requirements. Loop-mediated isothermal amplification (LAMP) is an appealing technology for point-of-need diagnostics due to its rapid analysis time and minimal instrumentation requirements.
Here, we designed and optimized nine LAMP assays that are sensitive and specific to targeted bacterial select agents including Bacillus anthracis, Francisella tularensis, Yersinia pestis, and Brucella spp. Evaluation of each assay determined preliminary limit of detection (LOD) with LOD confirmed across 60 replicates (≥ 95% positivity rate). Testing across a robust set of strains of the target agent, common DNA agents, and near-neighbors documented sensitivity and specificity for independent assays.
Specifically, all assays were 100% specific and sensitive except for Y. pestis Caf1 (90% inclusive across Y. pestis strains).
Here, we optimized assay turn-around-time, decreasing a standard 60 min traditional polymerase chain reaction (PCR) to 30 min using LAMP with positive results in as little as 5-10 min. Incorporating point of need sample processing and evaluating the potential inhibitory impact of sample matrices such as whole blood and soil would be needed to enable this test system for use on field-forward clinical and environmental sample testing.
即时诊断可为偏远或条件艰苦的地区提供高效检测能力，通过减少运输或样本运送需求来缩短获得结果的时间。环介导等温扩增技术（LAMP）因其分析速度快且对仪器设备要求低，是一种适用于即时诊断的有吸引力的技术。
在此，我们设计并优化了9种LAMP检测方法，这些方法对包括炭疽芽孢杆菌、土拉弗朗西斯菌、鼠疫耶尔森菌和布鲁氏菌属在内的目标细菌选择剂敏感且特异。对每种检测方法的评估确定了初步检测限（LOD），并在60次重复实验中得到确认（阳性率≥95%）。对一系列丰富的目标菌株、常见DNA试剂和近缘菌进行检测，记录了各独立检测方法的敏感性和特异性。
具体而言，除鼠疫耶尔森菌Caf1检测方法（对鼠疫耶尔森菌菌株的总体阳性率为90%）外，所有检测方法的特异性和敏感性均为100%。
在此，我们优化了检测周转时间，使用LAMP技术将标准60分钟的传统聚合酶链反应（PCR）缩短至30分钟，阳性结果在短短5 - 10分钟内即可获得。若要使该检测系统用于现场临床和环境样本检测，还需要整合即时样本处理方法，并评估全血和土壤等样本基质的潜在抑制作用。
