Indigoidine and deoxyviolacein co-pigmentation in a single bacterial cell enables visual discrimination of Pb(ii) and Hg(ii) in environmental water matrices.

Heavy metal co-contamination of lead and mercury in aquatic environments poses escalating risks to public health and ecosystems. Conventional instrumental methods, while accurate, are ill-suited for routine on-site screening due to high costs and operational complexity. Here, we describe a dual-pigment whole-cell biosensor engineered in Escherichia coli TOP10 for simultaneous visual detection of Pb(ii) and Hg(ii). The sensor integrates a Pb(ii)-responsive PbrR regulatory module that drives indigoidine biosynthesis with an Hg(ii)-responsive MerR module that drives deoxyviolacein biosynthesis within a single construct. Upon exposure to metal, the engineered strain produces distinct blue and purple pigments that are quantifiable at 600 nm and 570 nm, respectively. The biosensor achieved limits of detection of 6.74 nM for Pb(ii) and 0.0631 nM for Hg(ii), with both channels exhibiting high selectivity against competing metal ions. Performance was further validated in spiked surface water and seawater matrices, where clear dose-response relationships were maintained. By enabling naked-eye discrimination of two toxic metals near regulatory thresholds in a standard microplate format, this platform offers a low-cost, high-throughput alternative for preliminary environmental monitoring and emergency pollution assessment.