A self-contained and fully integrated fluidic cassette system for multiplex nucleic acid detection of bacteriuria
A self-contained and fully integrated fluidic cassette system for multiplex nucleic acid detection of bacteriuria
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用于菌尿多重核酸检测的独立且完全集成的流体盒系统
DOI：
10.1039/c9lc00994a
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发表时间：
2020-01-21
期刊：
影响因子：
6.1
通讯作者：
Xu, Youchun
中科院分区：
文献类型：
--
作者：
Li, Nan;Lu, Ying;Xu, Youchun
诊断传染病的黄金标准是基于培养的细菌病原体鉴定，这是耗时且费力的。当前分子诊断和微流体技术的进步使得快速检测临床标本中的细菌或病毒成为可能。然而，以“进样答出”方式快速、灵敏、多重检测病原体的需求尚未完全满足。在这项研究中，构建了一个独立且完全集成的流体盒及其配套分析仪，用于细菌的多重检测，以加速尿路感染（UTI）的诊断。完全集成的盒包含细菌分析所需的所有组件和试剂。所有细菌分析过程，包括细菌裂解、基于磁性硅胶珠的 DNA 提取、DNA 洗脱和多重环介导扩增 (LAMP)，均在盒中自动进行。该试剂盒成功应用于尿路感染中四种主要致病菌的检测，即大肠杆菌、奇异变形杆菌、鼠伤寒沙门氏菌和金黄色葡萄球菌。前三个在尿液样本中以 1 菌落形成单位 (CFU) mu L-1 的检测限 (LoD) 成功检测到，最后一个在尿液样本中以 10 CFU mu L-1 的 LoD 成功检测到，这表明该盒与手动方案相比具有相似的灵敏度，低于 UTI 所需的灵敏度。这种基于盒的样本到答案系统的周转时间约为 100 分钟，检测灵敏、完全自动化且准确，展示了成为 UTI 有用诊断工具的潜力。
The gold standard for diagnosing infectious diseases is culture-based identification of bacterial pathogens, which is time-consuming and labour-intensive. Current advances in molecular diagnostics and microfluidic technologies have made the rapid detection of bacteria or viruses in clinical specimens possible. However, the need for rapid, sensitive and multiplex detection of pathogens in a "sample-in and answer-out" manner has not been fully satisfied. In this study, a self-contained and fully integrated fluidic cassette and its supporting analyser were constructed for multiplex detection of bacteria to accelerate the diagnosis of urinary tract infections (UTIs). The fully integrated cassette contains all the necessary components and reagents for bacterial analysis. All of the bacterial analysis processes, including bacterial lysis, magnetic silica bead-based DNA extraction, DNA elution and multiplex loop-mediated amplification (LAMP), are automatically conducted in the cassette. This cassette was successfully applied for the detection of four major pathogenic bacteria in UTIs, i.e., Escherichia coli, Proteus mirabilis, Salmonella typhimurium and Staphylococcus aureus. The first three were successfully detected with a limit of detection (LoD) of 1 colony-forming unit (CFU) mu L-1 and the last was with a LoD of 10 CFU mu L-1 in urine samples, demonstrating that the cassette has similar sensitivity compared to that of the manual protocol, which is lower than that required by UTIs. The turnaround time for this cassette-based sample-to-answer system was approximately 100 minutes, and the detection is sensitive, fully automated, and accurate, demonstrating the potential to be a useful diagnostic tool for UTIs.