Designing a Whole-Cell Biosensor for Detection of Toxic Metals Using Intein Splicing Inhibition of Mycobacterium...

Heavy metal contamination, driven by anthropogenic disruption of natural geochemical cycles, has led to widespread bioaccumulation, posing a serious global health threat. Addressing this crisis demands the development of simple and sensitive strategies capable of monitoring metal pollution in the environment. Leveraging nongenetically modified native organisms, this research offers a promising low-cost, eco-friendly screening approach for detecting heavy metals in contaminated samples. Current study uses post-translational splicing of the Mycobacterium tuberculosis (Mtb) SufB precursor protein to detect metals by linking metal-induced splicing inhibition to viability loss of native mycobacterial cells. Toxic metal ions like Cd²⁺ and Hg²⁺ blocked splicing activity of Mtb SufB precursor protein over a concentration range of 25 μM-2 mM, while Pb²⁺ and Cr³⁺ failed to do so. An innovative biosensor platform was designed to detect metals by a simple Alamar Blue assay using attenuated Mtb H37Ra as indicator cells. Qualitative metal detection was assessed via colorimetric variation relating to mycobacterial viability, while concurrent spectral absorbance measurement enabled metal ion quantification. Loss of H37Ra cell viability by metal ions over the 25 μM-2 mM concentration range highlighted the sensitivity of the designed biosensor, while the addition of metal-specific chelators reversed the effect. Multiplexing ability was evaluated by including known splicing inhibitors like Cu²⁺, Zn²⁺, and Pt⁴⁺ over various concentration ranges alongside Cd²⁺ and Hg²⁺ in a simple 96-well plate format. The designed intein-based biosensor offers a user-friendly platform, readily standardizable for high-throughput detection using native organisms harboring metal-sensing precursor proteins. As a proof-of-concept, this study demonstrates the applicability of intein-based biosensing for initial heavy metal screening in environmental and industrial effluents, serving as a rapid and accessible tool prior to targeted advanced analysis.