A dual-color whole-cell biosensor for differential visual detection of As(III) and Hg(II) at drinking water limits

A single-plasmid dual-color biosensor for As(III) and Hg(II) visual detection. • Detection limits near or below drinking water regulatory thresholds. • Independent blue and purple pigments enable direct color discrimination. • Validated for environmental water matrices including seawater samples. A novel dual-color whole-cell biosensor was engineered for simultaneous visual detection of As(III) and Hg(II) within a single microbial host. The biosensor integrates an As(III)-responsive ArsR module driving indigo biosynthesis and an Hg(II)-responsive MerR module driving deoxyviolacein (DV) biosynthesis into a single plasmid designated pAs-indigo-Hg-DV. Upon exposure, the engineered Escherichia coli TOP10 generates distinct blue and purple pigments with absorption maxima at 620 nm and 570 nm, respectively. The system achieved limits of detection of 0.39 μM for As(III) (linear range 0.39–6.25 μM) and 0.49 nM for Hg(II) (linear range 7.8–125 nM), both approaching or falling below the respective drinking water regulatory limits. By using two independent color outputs from a single host, a dual-color biosensor for microscopic differential detection of two toxic metals was achieved, addressing the key unmet need of a flexible multi-channel multiplexed whole-cell biosensing system. The sensor supports naked-eye preliminary screening in a standard 96-well microplate format without dependence on instrumentation. It exhibited excellent Hg(II) selectivity and maintained robust performance in surface water and seawater matrices. This pigment-based dual-target strategy offers a low-cost, high-throughput platform for rapid co-exposure assessment of toxic heavy metals, providing a practical complement to conventional instrumental analysis and advancing the development of innovative biosensors for environmental monitoring. Download: Download high-res image (164KB) Download: Download full-size image