Abstract: Specific primers were designed based on the nuc gene, and the reaction system of the isothermal multiple self-matching-initiated amplification (IMSA) method was optimized through gradient experiments by setting different temperatures, incubation times, enzyme amounts, and other conditions; its specificity and sensitivity were evaluated and compared with the PCR method. The results showed that the reaction system of the IMSA method achieved optimal amplification efficiency and specificity under the conditions of 63 ℃ reaction temperature, 45 min incubation time, 16 U/reaction Bst DNA polymerase, 4.0 mmol/L Mg²⁺, 2.0 mmol/L dNTPs, and 1.2 mol/L betaine. In the detection of three types of samples (raw milk, meat products, and fruit and vegetable products), both the positive agreement rate and negative agreement rate between the IMSA method and the PCR method reached 100%, with Kappa values both being 1.00 (P<0.01), indicating that the detection results of Staphylococcus aureus in the three types of samples by the two methods were completely consistent. The rapid detection method for Staphylococcus aureus based on the IMSA method exhibited high specificity, high sensitivity, and good practicality in food testing, providing an efficient and reliable means for food safety detection.

Key words: food, Staphylococcus aureus, isothermal multiple self-matching-initiated amplification (IMSA)method, optimization, application