Rapid, sensitive and wide-spectrum colourimetric detection of viable Staphylococcus aureus in food using phage cocktail

Although bacteriophages (phages) are ideal biorecognition elements employed for Staphylococcus aureus detection, existing single-phage-based approaches are limited by the specific affinity of phages and false-negative results. To develop an improved phage-based colourimetric detection system and investigate its detection mechanism, we prepared a phage cocktail composed of three S. aureus phages (SapYZU10, SapYZU11, and SapYZUM13), immobilised it on Co-Mn nanozymes, and applied it to S. aureus for evaluation. The phage cocktail@Co-Mn demonstrated a sensitive, specific, and fast colourimetric evaluation of live S. aureus counts in approximately 17 min, with a detection limit of 92 CFU/mL. Moreover, the phage cocktail@Co-Mn was successfully used to detect S. aureus in food samples, with recovery rates ranging from 92.21 % to 105.56 %. The phage cocktail@Co-Mn also exhibited resilience to changes in pH (3–9), five food additives, and NaCl concentration (2 %–10 %). The detection mechanism of the phage cocktail@Co-Mn involves blockage of the active sites of the nanozyme producing O₂^•–. Notably, owing to the synergistic recognition spectra of the three phages in the cocktail, the recognition rate for the strains was a remarkable 100 %, compared with the much lower recognition rates of individual phages (between 74.7 % and 76.9 %). The recognition mechanism involves adsorption between phage tail fibres and S. aureus hosts. This pioneering study revealed that the phage cocktail is an effective biorecognition element suitable for the rapid, sensitive, specific, reliable, and colourimetric detection of viable S. aureus strains in the food industry, providing innovative directions for the phage-based detection methods.