Survey of Staphylococcal Enterotoxin Genes, Exfoliative Toxin Genes, and Toxic Shock Syndrome Toxin 1 Gene in Non- Staphylococcus aureus Species

Two multiplex PCR enzyme immunoassays (PCR-EIAs) were developed forStaphylococcus aureus exotoxin gene screening as an alternative to the conventional biological assays, which depend on detectable amounts of toxins produced. One set of oligonucleotide primers and probes was designed to search for enterotoxin A to E genes (entA, entB, entC,entD, and entE), and the other one was designed to detect the staphylococcal exfoliative toxin genes (etaand etb) and the toxic shock syndrome toxin 1 gene (tst). Oligonucleotide primers were used as published previously, modified or newly developed to meet the requirements of both good size-distinguishable amplification bands of multiplex PCR and the temperature limit of the uracil DNA glycosylase system for carryover protection. Amplification products were visualized by agarose gel electrophoresis, and specificity was controlled with the aid of a DNA EIA system using oligonucleotide probes derived from the sequences of the S. aureus toxin genes. PCR procedures were performed by using template nucleic acids extracted from a panel of S. aureus reference strains and from a collection of 50 clinical strains. The PCR results were compared with those of immunological toxin production assays. This multiplex PCR-EIA system offers an alternative method for the rapid, sensitive, specific, and simultaneous detection of the clinically important exotoxin potency of isolatedS. aureus strains for diagnostic purposes as well as research studies.

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Use of Multiplex PCR To Detect Classical and Newly Described Pyrogenic Toxin Genes in Staphylococcal Isolates

Journal of Clinical Microbiology ◽

10.1128/jcm.37.10.3411-3414.1999 ◽

1999 ◽

Vol 37 (10) ◽

pp. 3411-3414 ◽

Cited By ~ 217

Author(s):

Steven R. Monday ◽

Gregory A. Bohach

Keyword(s):

Staphylococcus Aureus ◽

Multiplex Pcr ◽

Toxic Shock Syndrome ◽

Food Poisoning ◽

Toxic Shock ◽

Cellular Targets ◽

Staphylococcal Enterotoxins ◽

Toxin Genes ◽

Toxic Shock Syndrome Toxin ◽

Single Reaction

Staphylococcus aureus may contain one or more genes that encode a variety of immunomodulatory pyrogenic toxins (PTs), including the staphylococcal enterotoxins and toxic shock syndrome toxin (TSST). The PTs interact with several cellular targets to produce disease, such as food poisoning and toxic shock syndrome. At present, nine serologically distinct enterotoxins and one immunoreactive form of TSST have been identified and characterized. As isolates of S. aureus are further assessed, it is anticipated that this number will increase. To facilitate screening, a multiplex PCR was designed to simultaneously determine which of these 10 currently known PT genes an individualS. aureus isolate possesses. We show here, using S. aureus isolates with characterized PT phenotypes, that this novel PCR technique reliably detects each of the known PTs in a single reaction.

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