Abstract:

【Objective】 Staphylococcus aureus is an important pathogen of zoonose, mainly initiates its transmission through food contamination and causes food poisoning, suppurative inflammation of tissues and organs. Thus, the development of a technique that can identify the microbe quickly and accurately is of great importance. 【Method】 Loop-mediated isothermal amplification (LAMP) for DNA is a novel nucleic acid amplification method with rapidity and high sensitivity. In this study, a LAMP method for detecting Staphylococcus aureus was developed. Four primers including two outer primers F3, B3 and two inner primers FIP, BIP were designed according to highly conserved Sa442 gene of Staphylococcus aureus. Genomic DNA extracted from 45 bacteria strains including 20 Staphylococcus aureus strains and 25 other bacterial strains were amplified using the primers with Bst DNA polymerase at 65oC, only Staphylococcus aureus strains were LAMP positive result, which indicated that the LAMP primers designed were highly specific for target bacteria. 【Result】 The detection sensitivity of the LAMP method for Staphylococcus aureus purely cultured was 25 CFU/mL and for Staphylococcus aureus in contaminated food was 42 CFU/g, which could complete the test in 40-60 min. In practice, 115 pieces of samples detected from 958 pieces of meat samples, egg, dairy products and artificial contamination samples were positive using the LAMP method, which accorded with the detection result by GB. 【Conclusion】 The LAMP assay established in this study is a sensitive, rapid and simple tool for detecting Staphylococcus aureus.

Key words: Staphylococcus aureus')">Staphylococcus aureus, Sa442 gene, loop-mediated isothermal amplification (LAMP), rapid detection