Filter-based template preparation for rapid and sensitive PCR detection of Staphylococcus aureus in meat / 生物工程学报

ABSTRACT

In the present study, an assay using Polymerase Chain Reaction (PCR) was developed for the detection of Staphylococcus aureus in meat. Based on flotation and solvent extraction technology, FTA filter was used to extract S. aureus DNA from artifically contaminated meat. Primers targeting the thermostable nuclease gene (nuc) were used to amplify a 279 bp DNA fragment which was confirmed by DNA sequencing. The detection limit of PCR was 10 cfu x g(-1) meat of S. aureus. This novel FTA-PCR assay allows for detection of Staphylococcus aureus in meat in <6 h, which is 12-24 h less than that of conventional PCR with enrichment method. Seventy-two samples were analyzed and the detection rate using the standard cultivation method was 70.8%, detection time was 5d. The detection rate of PCR amplification using filters was 73.6%, detection time was 6h. The detection rate of Baird-Parker R.P.F method was 69.4%, detection timewas 18 h. The detection rate of petrfilm RSA method was 61.1%, detection time was 18 h. Thus PCR amplification using filters provides a faster and more sensitive method of S. aureus detection than the standard cultivation method. At the same time,it provides an universal process for preparing DNA template.