Rapid and sensitive detection of Staphylococcus aureus via an all-in-one staggered strand exchange amplification platform

Abstract

Staphylococcus aureus is a common foodborne pathogen that causes food poisoning and infectious diseases in humans and animals. Rapid detection of S. aureus with high sensitivity is of great significance to prevent the spread of this pathogen. In this study, we developed a staggered strand exchange amplification (SSEA) method by refining denaturation bubble-mediated strand exchange amplification (SEA) to detect S. aureus at a constant temperature with high specificity and efficiency. This method employs a DNA polymerase and two sets of forward and reverse primers arranged in tandem that invade denaturation bubbles of double-stranded DNA. In comparison, the sensitivity of SSEA was 20 times that of SEA. Subsequently, magnetic bead (MB)-based DNA extraction was introduced into SSEA to establish an all-in-one SSEA platform that incorporated sample processing, amplification and detection in a single tube. The use of MBs further enhanced the sensitivity of SSEA by two orders of magnitude. Specificity tests showed that the all-in-one SSEA could specifically identify S. aureus and had no cross-reaction with other common foodborne pathogens. For artificially spiked meat samples, the method could detect 1.0 × 10² CFU g⁻¹S. aureus in pork and 1.0 × 10³ CFU g⁻¹ in either duck or scallop samples without a bacterial enrichment step. The entire assay can be completed sample-to-answer within 1 h. Thus, we believe that this easy-to-operate diagnostic platform enables sensitive and accurate detection of S. aureus and holds great promise for the food safety industry.