CRISPR/Cas9-based engineered Escherichia coli biosensor for sensitive and specific detection of Cd(II) in drinking water

• Published in: Chemosphere (Oxford) 2024-08, Vol.362, p.142607, Article 142607
• Main Authors: Wei, Yijun, Shi, Danyang, Chen, Tianjiao, Zhou, Shuqing, Yang, Zhongwei, Li, Haibei, Yang, Dong, Li, Junwen, Jin, Min
• Format: Article
• Language: English
• Subjects: Bacterial whole-cell biosensor; Biosensing Techniques - methods; Cadmium - analysis; Cd(II); CRISPR-Cas Systems; CRISPR/Cas9 gene editing; cysI; Detection; Drinking Water - microbiology; Environmental Monitoring - methods; Escherichia coli - genetics; Gene Editing; Limit of Detection; Water Pollutants, Chemical - analysis
• ISSN: 0045-6535; 1879-1298; 1879-1298
• DOI: 10.1016/j.chemosphere.2024.142607

Cadmium (Cd) is a ubiquitous pollutant that poses a potential threat to human health. Monitoring Cd(II) in drinking water has significant implications for preventing potential threats of Cd(II) to human. However, the weak signal output and response to nontarget interference limit the detection of Cd(II) using bacterial biosensors. In this study, to enable sensitive and specific detection of Cd(II) in water, a stable whole-cell biosensor, K12-PMP-luxCDABE-△cysI, was constructed in a dual-promoter mode by fusing the mercury promoter Pmer, regulatory gene merR(m), and luciferase gene luxCDABE into the E.coli chromosome based on CRISPR/Cas9 gene editing technology. By knocking out the cadmium-resistance-gene cysI, the sensitivity of the biosensor to Cd(II) was further enhanced. The constructed E. coli biosensor K12-PMP-luxCDABE-△cysI exhibited good nonlinear responses to 0.005–2 mg/L Cd(II). Notably, among the three constructed E. coli biosensor, it exhibited the strongest fluorescence intensity, with the limit of detection meeting the allowable limit for Cd(II) in drinking water. Simultaneously, it could specifically detect Cd(II). Nontarget metal ions, such as Zn(II), Hg(II), and Pb(II), did not affect its performance. Furthermore, it exhibited superior performance in detecting Cd(II) in real drinking water samples by avoiding background interference, and showed excellent stability with the relative standard deviation under 5%. Thus, K12-PMP-luxCDABE-△cysI holds promise as a potential tool for the detection of Cd(II) in drinking water. [Display omitted] •E. coli Cd(II) biosensors were constructed based on CRISPR/Cas9 gene editing.•K12-PMP-luxCDABE-△cysI holds promise for detecting Cd(II) in drinking water.•It can approximately quantify Cd(II) in the range of 0.005–2 mg/L.•It specifically detects Cd(II) by avoiding nontarget interference from drinking water.•The knockout of cysI greatly helps improve the detection limit of Cd(II).