Isolation and Detection of Enterotoxigenic Staphylococcus Aureus Type A by Multiplex PCR

This method is a rapid, sensitive, specific and inexpensive alternative for traditional biochemical and serological assays and can be used for detection of agent produced type A staphylococcal enterotoxin.  To investigate the distribution of staphylococcal enterotoxin (sea) gene in S. aureus, 98 samples obtained from environment were analyzed by Multiplex PCR and 89 confirmed as S. aureus. Six (6.74%) S. aureus isolates were found to be positive for enterotoxin type A gene. Nine (9.18%) bacteria isolates were found to be positive for S. aureus by biochemical methods but did not confirm by molecular analysis. In this study, S. aureus strains were isolated from nose of carriers and patients, and environmental specimens were confirmed by biochemical test. Primers were designed and the PCR was used to amplify the staphylococcal enterotoxin A gene (sea) and the staphylococcal nuclease gene (nuc). DNA amplification fragments of 279 bp for the staphylococcal nuclease gene (nuc) and 552 bp for staphylococcal enterotoxin A gene (sea) was confirmed by digestion enzymes. S. epidermidis used as negative control and did not yield a PCR product. Staphylococcus aureus produces different extra-cellular protein toxins and virulence factors. One of the most important extra-cellular proteins is enterotoxin which cause staphylococcal food poisoning (SFP) effect. These toxins pass from the digestive tract to the blood circulation and consecutively activate the nerve centre of the emetic reflex, causing nausea, vomiting, abdominal cramps and diarrhea, which can lead to dehydration of the organism. Numerous detection methods are based on the evidence of the enterotoxins directly (ELISA, reversal passive latex agglutination and others), but DNA amplification methods like PCR (polymerase chain reaction) can show the presence of enterotoxigenic strains of S. aureus before the expression of enterotoxins on the base of specific gene sequences.