Paper-based colourimetric biosensor for simultaneous detection of UTI-associated nitrite and β-lactamase-mediated resistance using an iodometric method

Urinary tract infections (UTIs) are among the most prevalent bacterial infections worldwide and are increasingly complicated by the emergence of antibiotic resistance, particularly β- lactam resistance. Conventional diagnostic methods for UTI identification and antimicrobial susceptibility testing are often labour-intensive and time-consuming and require laboratory infrastructure, thereby restricting their use in resource-limited settings. To address this gap, this study developed a smartphone-assisted, paper-based colourimetric biosensor for the simultaneous detection of UTI-associated nitrite and β-lactamase-mediated antibiotic resistance, integrating two clinically relevant indicators into a low-cost, single platform with smartphone-assisted quantitative readout. The sensing device was fabricated from Whatman Grade-1 filter paper with hydrophobic barriers to form separate detection zones for nitrite and β-lactamase analysis. The nitrite detection zone employed the Griess reaction, while the antibiotic resistance detection zone utilized an agarose- based iodometric system containing penicillin G, iodine, starch, and chitosan- capped silver nanoparticles (AgChNPs). The synthesised AgChNPs were characterised by FT-IR, UV-visible spectroscopy, HRTEM, and zeta potential analysis, confirming successful synthesis. Artificial urine samples were analysed using smartphone-based image acquisition and ImageJ-based intensity quantification. The nitrite detection zone exhibited limits of detection (LoD) and limits of quantification (LoQ) of 0.670 and 2.031 ppm, respectively, while the β-lactamase detection zone showed corresponding values of 0.324 and 0.983 U/mL. The proposed platform offers a rapid, affordable, portable, and integrated approach for simultaneous UTI screening and β-lactamase-mediated antibiotic resistance detection, with potential for point-of-care applications in resource-limited settings.