Inosine-Assisted Signal-On Electrochemical Biosensor for Selective Bacterial DNA Detection

Abstract

Accurate and rapid identification of Shigella sonnei and Stenotrophomonas maltophilia remains challenging due to their close genomic similarity to other Gram-negative bacteria and their limited coverage in routine molecular diagnostic panels. In this study, we report a label-free electrochemical DNA biosensing strategy for the sequence-selective detection detection of these two clinically relevant but under-recognized pathogens. The biosensor is based on a pencil graphite electrode modified with carboxyl-functionalized single-walled carbon nanotubes (SWCNT-COOH), providing a conductive and chemically active interface for probe immobilization. To minimize intrinsic guanine-related background signals, inosine-substituted DNA probes were employed, enabling a hybridization-induced “signal-on” response that arises exclusively from the target DNA. Stable and reproducible probe attachment was achieved through optimized –COOH/–NH₂ interactions between the SWCNT interface and amine-terminated probes. Under optimized conditions, the biosensor exhibited linear responses in the 20–80 µg/mL range, with limits of detection of 7.3 µg/mL for S. sonnei and 10.2 µg/mL for S. maltophilia. The platform demonstrated effective discrimination against mismatched and non-complementary sequences and retained analytical performance in artificial saliva matrices, supporting its applicability in non-invasive sample environments. This work presents a simple and low-cost electrochemical approach for DNA-based detection of S. sonnei and S. maltophilia. The proposed sensing strategy highlights the potential of inosine-assisted, signal-on electrochemical readout as a modular platform for nucleic-acid detection, with relevance for decentralized and resource-limited diagnostic settings.