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Sensitive, Rapid, Quantitative and in Vitro Method for the Detection of Biologically Active Staphylococcal Enterotoxin Type E

Author:
Rasooly, Reuven, Do, Paula, Hernlem, Bradley
Source:
Toxins 2016 v.8 no.5 pp. 1-11
ISSN:
2072-6651
Subject:
B-lymphocytes, Staphylococcus aureus, T-lymphocytes, bioassays, bioluminescence, detection limit, dose response, enterotoxins, gastroenteritis, gene expression regulation, genetic engineering, kittens, laboratory animals, luciferase, major histocompatibility complex, mixed culture, monkeys, poisoning, reporter genes, staphylococcal intoxication, superantigens, France, United Kingdom, United States
Abstract:
Staphylococcus aureus is a major bacterial cause of clinical infections and foodborne illnesses through its production of a group of enterotoxins (SEs) which cause gastroenteritis and also function as superantigens to massively activate T cells. In the present study, we tested Staphylococcal enterotoxin type E (SEE), which was detected in 17 of the 38 suspected staphylococcal food poisoning incidents in a British study and was the causative agent in outbreaks in France, UK and USA. The current method for detection of enterotoxin activity is an in vivo monkey or kitten bioassay; however, this expensive procedure has low sensitivity and poor reproducibility, requires many animals, is impractical to test on a large number of samples, and raises ethical concerns with regard to the use of experimental animals. The purpose of this study is to develop rapid sensitive and quantitative bioassays for detection of active SEE. We apply a genetically engineered T cell-line expressing the luciferase reporter gene under the regulation of nuclear factor of activated T-cells response element (NFAT-RE), combined with a Raji B-cell line that presents the SEE-MHC (major histocompatibility complex) class II to the engineered T cell line. Exposure of the above mixed culture to SEE induces differential expression of the luciferase gene and bioluminescence is read out in a dose dependent manner over a 6-log range. The limit of detection of biologically active SEE is 1 fg/mL which is 109 times more sensitive than the monkey and kitten bioassay.
Agid:
63154
Handle:
10113/63154