Phage Cocktail-Mediated Ultrasensitive Real-Time Electrochemical Whole-Cell Detection of Klebsiella pneumoniae on a SPCE

Rapid and selective detection of pathogenic bacteria is essential for timely clinical diagnosis and environmental surveillance. In this work, we report a bacteriophage-assisted electrochemical sensing platform for the selective detection of Klebsiella pneumoniae using an Er2O3/Co3O4/Au/polyethylenimine (PEI)-modified screen-printed carbon electrode (SPCE). The hierarchical nanocomposite was engineered to provide high surface area, enhanced charge transfer kinetics, and abundant functional sites for efficient phage immobilization. PEI served as a cationic linker to facilitate electrostatic attachment of lytic bacteriophages while maintaining their biological activity and host specificity. Upon target recognition, phage-bacteria binding at the electrode interface produced pronounced impedance changes due to interfacial blocking and altered electron transfer dynamics. The developed biosensor exhibited high selectivity for Klebsiella pneumoniae over nontarget bacteria and could distinguish live from dead bacteria. The sensor showed a wide linear detection range of 10-107 CFU/mL with a low detection limit of 7.36 CFU/mL and good repeatability with an RSD of ∼1.17%. The real-world applicability of the sensor was tested using aquarium water samples and demonstrated good performance, with recovery percentages ranging from ∼91.71 to 96.26%. This work establishes a robust nano-bio hybrid interface strategy that integrates rare-earth/transition metal oxide electrocatalysts with phage biorecognition on disposable SPCE platforms. The developed system shows strong potential for point-of-care pathogen monitoring in clinical, environmental, and food safety applications.