Establishment and evaluation of high-throughput fluorescent PCR assay for identifying 16 kinds of Staphylococcus aureusenterotoxins

Objective To establish a high-throughput fluorescent PCR assay (fluorescent probe melting curve method) for the identification of 16 kinds of Staphylococcus aureus enterotoxins.

Methods Primers were designed according to S. aureus enterotoxin SEA-SEQ gene sequences with the different annealing temperature, and the PCR assay was established, then the limit detection, sensitivity, specificity and repeatability of the assay were evaluated by detecting 158 strains of food borne S. aureus in comparison with conventional PCR.

Results The limit detection of the established PCR assay ranged from 0.80 ng/μl to 2.15 ng/μl. The specificity was 100.0%, no cross fluorescence signal was produced, and the coefficient of variation was lower than 1%. Compared with conventional PCR, the sensitivity and specificity of new PCR assay were 95.4% and 100.0%, the Kappa was 0.88.

Conclusion The established PCR assay is rapid, accurate and specific and can be used to identify all the 16 kinds of enterotoxins of S. aureus.