School of Public Health, Wannan Medical College, 241002, Wuhu, Anhui, China.
Key Laboratory of Environmental Toxicology and Pollution Control Technology of Anhui Province, Hefei Institutes of Physical Science, Chinese Academy of Sciences, 230031, Hefei, Anhui, China.
Anal Chim Acta. 2024 Nov 22;1330:343299. doi: 10.1016/j.aca.2024.343299. Epub 2024 Oct 3.
As a threat to human health and public health, cadmium (Cd) pollution has received widespread social concern. Our previously constructed CadR-based bacterial whole cell biosensor (WCB) epCadR5 showed high sensitivity and specificity in cadmium detection. However, the application of the sensor is still hindered by the need for laboratory equipment to read the fluorescence signal output. In this study, we aimed to optimizing the sensor to make it available for visual detection of environmental cadmium and simplify the detection process to advance practical application of the sensor.
By replacing the constitutive promoter with J110, the fluorescence signal output of the sensor was significantly increased and the fluorescence leakage was decreased. In addition, the fluorescence signal output of green fluorescence protein (GFP) was enhanced by the addition of a 5' untranslated region (5'-UTR) mlcR10. The fluorescence signal output of the WCB is sufficiently robust to be visible and distinguishable to the naked eye, which is of paramount importance for visual detection. The sensor readout can be conveniently recorded by mobile phone camera and quantified. For ease of on-site application, the WCB's visual detection procedures and conditions were further optimized and simplified. The WCB demonstrated good linearity and detection limit (1.81 μg/L) for visual detection of Cd without the assistance of bulky laboratory equipment. For the detection of real environmental samples, the WCB visual detection results were close to those of WCB-flow cytometry (FACS) and graphite furnace atomic absorption spectroscopy (GFAAS).
In this work, we developed an easy-to-use, on-site and visual detection biosensor for monitoring environmental Cd. It will advance the utilization of cadmium WCBs in practical settings. The optimization and simplification strategy in the study also provide new insights into the visualization of other bacterial biosensors, and will advance the practical application of WCBs.
作为对人类健康和公共卫生的威胁，镉（Cd）污染受到了广泛的社会关注。我们之前构建的基于 CadR 的细菌全细胞生物传感器（WCB）epCadR5 在镉检测中表现出了高灵敏度和特异性。然而，由于需要实验室设备读取荧光信号输出，传感器的应用仍然受到限制。在这项研究中，我们旨在对传感器进行优化，使其能够用于环境镉的可视化检测，并简化检测过程，以推进传感器的实际应用。
通过用 J110 替换组成型启动子，传感器的荧光信号输出显著增加，荧光泄漏减少。此外，通过添加 5'非翻译区（5'-UTR）mlcR10，增强了绿色荧光蛋白（GFP）的荧光信号输出。WCB 的荧光信号输出足以肉眼可见且可区分，这对于可视化检测至关重要。传感器读数可以通过手机摄像头方便地记录和定量。为了便于现场应用，进一步优化和简化了 WCB 的可视化检测程序和条件。该 WCB 无需使用大型实验室设备即可对 Cd 进行可视化检测，具有良好的线性和检测限（1.81μg/L）。对于真实环境样品的检测，WCB 可视化检测结果与 WCB-流式细胞术（FACS）和石墨炉原子吸收光谱法（GFAAS）接近。
在这项工作中，我们开发了一种易于使用、现场和可视化的环境 Cd 监测生物传感器。它将推进 Cd WCB 在实际环境中的应用。研究中的优化和简化策略也为其他细菌生物传感器的可视化提供了新的思路，并将推进 WCB 的实际应用。
