Colorimetric detection of Staphylococcus aureus with enhanced sensitivity based on phage covalently immobilized Co₃O₄...

Staphylococcus aureus (S. aureus) is a common foodborne pathogen, posing a serious threat to public health. Consequently, it is crucial to establish a platform for sensitive and specific determination of S. aureus in food. Herein, phage SapYZUH₅, isolated by our lab, was covalently immobilized on Co₃O₄ to synthesize SapYZUH₅@Co₃O₄. Notably, SapYZUH₅@Co₃O₄ exhibited remarkable oxidase-like activity, enabling the catalysis of dissolved oxygen to generate superoxide anion free radicals and accelerate the TMB chromogenic reaction. Upon introduction of S. aureus, specific capture by SapYZUH₅@Co₃O₄ resulted in inhibiting its oxidase-like activity and decelerating the 3,3',5,5'-tetramethylbenzidine (TMB) chromogenic reaction. Moreover, S. aureus can be lysed to release the reductive bacterial contents, which can further inhibit the TMB chromogenic reaction. Based on this principle, SapYZUH₅@Co₃O₄ + TMB reaction system was employed for detection with enhanced sensitivity of S. aureus, yielding an equation: A =  - 0.092 Log (C_SA) + 0.79 (R² = 0.987), with an ultralow limit of detection (LOD) of 28 CFU mL^-1. This system exhibited remarkable specificity and anti-interfere towards S. aureus, owing to the excellent affinity of SapYZUH₅ towards S. aureus. In addition, S. aureus in the actual food samples was detected using this system, yielding recoveries ranging from 96.34 to 109.43%, demonstrating its exceptional accuracy. Hence, our proposed covalent immobilization of phage on the nanozyme can realize sensitive and specific colorimetric determination of S. aureus in food samples.