Double phage displayed peptides co-targeting-based biosensor with signal enhancement activity for colorimetric detection of Staphylococcus aureus

The development of simple, fast, sensitive, and specific strategies for the detection of foodborne pathogenic bacteria is crucial for ensuring food safety and promoting human health. Currently, detection methods for Staphylococcus aureus still suffer from issues such as low specificity and low sensitivity. To address this problem, we proposed a sensitivity enhancement strategy based on double phage-displayed peptides (PDPs) co-targeting. Firstly, we screened two PDPs and analyzed their binding mechanisms through fluorescent localization, pull-down assay, and molecular docking. The two PDPs target S. aureus by binding to specific proteins on its outer membrane. Based on this phenomenon, a convenient and sensitive double PDPs colorimetric biosensor was developed. Double thiol-modified phage-displayed peptides (PDP-SH) enhance the aggregation of gold nanoparticles (AuNPs), whereas the specific interaction between the double PDPs and bacteria inhibits the aggregation of AuNPs, resulting in an increased visible color change before and after the addition of bacteria. This one-step colorimetric approach displayed a high sensitivity of 2.35 CFU/mL and a wide detection range from 10–2 × 108 CFU/mL. The combination with smartphone-based image analysis improved the portability of this method. This strategy achieves the straightforward, highly sensitive and portable detection of pathogenic bacteria.