Development of a multiplex PCR assay for the identification of Staphylococcus aureus enterotoxigenic strains isolated from milk and dairy products

Detection by multiplex PCR of Staphylococcus aureus , S. intermedius and S. hyicus in artificially contaminated milk

“…In another study with mPCR, CREMONESI et al (2005) detected simultaneously sequences of genes 23S of rRNA, of coagulase, of nuc and of enterotoxin A, B, C, D, G, H, I, J and L from a concentration of 1pg of DNA of S. aureus isolates, obtained from samples of milk and dairy products. In the same research, the global efficiency of the process was not calculated, but the quantity of DNA template obtained from microbial concentrations equal to or lower than 10 2 UFC mL -1 was shown to vary between 0.33mg mL -1 (extracted from the concentration of 50UFC mL -1 in bovine whole milk artificially contaminated with S. aureus) and 1.63mg mL -1 (extracted from concentration of 10 2 UFC mL -1 in bovine whole milk artificially contaminated with S. aureus).…”

Standardization of a multiplex PCR for the identification of coagulase-positive Staphylococcus

“…The choice of the nuc gene was based on the fact that the three species under study were able to produce the thermonuclease enzyme and, therefore, carry the nuc gene, and on the high correlation between this gene and the production of enterotoxins (CREMONESI et al, 2005). The oligonucleotides (primers) were synthetized by Invitrogen.…”

“…Several authors have suggested the use of molecular methods, for example, amplification of specific sequences of the microbial genome by PCR, as a viable alternative to the traditionally used morphological and biochemical methods (SILVA et al, 2003;BARON et al, 2004;BECKER et al, 2005;CREMONESI et al, 2005;YANG et al, 2007).…”

Surface‐activated chemical ionization and cation exchange chromatography for the analysis of enterotoxin A

“…[34] For detection of the staphylococcal enterotoxin gene (sea), a multiplex PCR assay was used as described. [35] This PCR assay includes species-specific primers for 23S rRNA, the coagulase gene (coa), the thermonuclease gene (nuc) and staphylococcal enterotoxin genes such as sea, sec, sed, seg, seh, sei, sej and sel. The reference strain ATCC 700699 (harbouring sea, sec, seg, sei and sel) was included as a positive control for the PCR.…”

“…The presence of a gene does not necessarily indicate detectable levels of its expression, and differences between the presence of a gene and its expression can be resolved by further molecular tests. [35] Conclusions A reliable and sensitive approach for screening milk samples for S. aureus contamination based on LC-CEC-SACI-MS/MS was developed.…”