Key Laboratory of Advanced Mass Spectrometry and Molecular Analysis of Zhejiang Province, Institute of Mass Spectrometry, School of Material Science and Chemical Engineering, Ningbo University, Ningbo, 315211, China.
Department of Hematology, Nanfang Hospital, Southern Medical University, Guangzhou, 510515, China.
Biosens Bioelectron. 2023 Jan 15;220:114852. doi: 10.1016/j.bios.2022.114852. Epub 2022 Oct 31.
Live foodborne pathogens proliferate rapidly and do great harm to human health, which requires appropriate methods to supervise. In this work, a portable adenosine triphosphate (ATP) bioluminescence sensor with high specificity for live E. coli O157:H7 strain synergistically enhanced by orientated phage-modified stir bar extraction and bio-proliferation was developed. In brief, the selected phages were directionally immobilized on the poly(diallyldimethylammonium chloride)-modified gold stir bar as the bioreceptor. Following the simple stir bar absorptive extraction and bio-proliferation in the Luria-Bertani medium, the number of captured E. coli O157:H7 exploded. Finally, it was quantified by a portable ATP bioluminescence sensor. Benefitting from the high specificity of phage and simple signal dual-amplification strategy, the proposed biosensor achieved the recognition of live bacteria at strain level with superior sensitivity. Also, the portable signal readout made it suitable for on-site detection. Under optimal conditions, this bioassay provided a detectable range of 10-10 CFU mL with a low detection limit of 30 CFU mL within 30 min. The detection results for real samples demonstrated that there were no differences between the assay and the plate counting method, while the detection time was largely shortened. Furthermore, the assay gives a novel path for the point-of-care test (POCT) of live E. coli strain, which is promising to be extended to other virulent strains measurement with corresponding phages.
食源性致病菌繁殖迅速，对人体健康危害极大，需要用适当的方法进行监督。在这项工作中，开发了一种便携式三磷酸腺苷（ATP）生物发光传感器，该传感器对活的 E. coli O157:H7 菌株具有高度特异性，通过定向噬菌体修饰搅拌棒提取和生物增殖协同增强。简而言之，选择的噬菌体被定向固定在聚（二烯丙基二甲基氯化铵）修饰的金搅拌棒上作为生物受体。在简单的搅拌棒吸收提取和在 LB 培养基中的生物增殖后，捕获的 E. coli O157:H7 数量剧增。最后，用便携式 ATP 生物发光传感器进行定量。受益于噬菌体的高特异性和简单的信号双重放大策略，该生物传感器实现了对活细菌的菌株水平识别，具有卓越的灵敏度。此外，便携式信号读出使其适用于现场检测。在最佳条件下，该生物测定法提供了 10-10 CFU mL 的检测范围，检测限为 30 CFU mL，检测时间为 30 分钟。实际样品的检测结果表明，该测定法与平板计数法之间没有差异，而检测时间大大缩短。此外，该测定法为即时检测（POCT）提供了一条新途径活的大肠杆菌菌株，有望通过相应的噬菌体扩展到其他毒力菌株的测量。
