Preparation of Carbon Dots with Peroxidase-like Activity and Their Application in Staphylococcus aureus Detection and Antimicrobial Susceptibility Test

Abstract

Carbon dot (CD) nanozymes with excellent biocompatibility, optical properties, and catalytic activity show great promise for microbial detection and drug sensitivity testing. This study reports the synthesis of metal-doped green-emitting CDs with good peroxidase-like activity, which were synthesized via a one-step hydrothermal route using thiourea, N-[3-(trimethoxysilyl)propyl]ethylenediamine, and catechol as the starting materials and FeCl₃ as the doping agent. In the presence of H₂O₂, CDs catalyze the oxidation of 3,3′,5,5′-tetramethylbenzidine (TMB), producing a blue product; however, in the presence of bacteria and H₂O₂, the bacterial catalase enzyme decomposes H₂O₂ and inhibits the catalytic activity of CDs, preventing the color change. The bacterial catalase enzyme neutralizes H₂O₂, which prevents the CDs from producing the color-changing reaction with TMB. Based on the CDs-TMB-H₂O₂ cascade system of bioenzymes and nanozymes, we developed a rapid, sensitive, and direct colorimetric detection method for Staphylococcus aureus (S. aureus) with a detection limit of 2 × 10³ CFU/mL and a linear range of 2 × 10³–2 × 10⁶ CFU/mL. This visual detection method was successfully applied to the detection of S. aureus in food samples. Antibiotics have different effects on the proliferation of sensitive and resistant bacterial strains, leading to different levels of hydrolysis of H₂O₂ in the bacterial solution and resulting in varying intensities of the solution color; therefore, we developed a simple and visual antibiotic susceptibility test. The applications of CD nanozymes provide a powerful tool for detecting pathogenic bacteria in food, clinical, and environmental samples and infections.