{"status":"ok","message-type":"work","message-version":"1.0.0","message":{"indexed":{"date-parts":[[2026,9,23]],"date-time":"2026-09-23T15:45:32Z","timestamp":1790178332935,"version":"4.1.0"},"reference-count":19,"publisher":"MDPI AG","issue":"6","license":[{"start":{"date-parts":[[2026,6,22]],"date-time":"2026-06-22T00:00:00Z","timestamp":1782086400000},"content-version":"vor","delay-in-days":0,"URL":"https:\/\/creativecommons.org\/licenses\/by\/4.0\/"}],"content-domain":{"domain":[],"crossmark-restriction":false},"short-container-title":["Microorganisms"],"abstract":"<jats:p>Staphylococcus aureus is a major bacterial pathogen that can cause clinical infections and foodborne illnesses through the production of 25 exotoxin types. The most frequently implicated toxins in food poisoning outbreaks are Staphylococcal enterotoxins type A\u2013E (SEA-SEE), which are the first enterotoxins discovered. While in vitro detection methods are available to identify the presence of enterotoxins, they cannot distinguish between biologically active and inactive forms of the toxins. Detection of biologically active enterotoxins currently relies on in vivo testing, using the emetic response in kittens or monkeys. Here, we show the development of an in vitro assay to detect the active forms of SEB, a potential biological warfare agent and leading cause of food poisoning, and SEC1, a frequent cause of staphylococcal food poisoning. The novel assay involves the implementation of a genetically engineered Jurkat T-cell line expressing TCR V\u03b23, resulting in a dose response of IL-2 production when exposed to active toxin. We also show that at a concentration of 100 ng\/mL, the biological activity of SEB is significantly decreased at temperatures over 70 \u00b0C, while pasteurization at 63 \u00b0C only slightly reduces the biological activity of the toxin. Our studies provide an alternative method to animal testing to determine the presence of active toxins and provide possible inactivation methods of the toxins.<\/jats:p>","DOI":"10.3390\/microorganisms14061383","type":"journal-article","created":{"date-parts":[[2026,6,22]],"date-time":"2026-06-22T09:49:55Z","timestamp":1782121795000},"page":"1383","update-policy":"https:\/\/doi.org\/10.3390\/mdpi_crossmark_policy","source":"Crossref","is-referenced-by-count":1,"title":["In Vitro Detection of Biologically Active Staphylococcal Enterotoxins Type B and C1 as an Alternative to In Vivo Testing"],"prefix":"10.3390","volume":"14","author":[{"given":"Reuven","family":"Rasooly","sequence":"first","affiliation":[{"name":"Emeritus, Foodborne Toxin Detection & Prevention Research Unit, Agricultural Research Service, USDA, Albany, CA 94710, USA"}],"role":[{"vocabulary":"crossref","role":"author"}]},{"given":"Naomi","family":"Balaban","sequence":"additional","affiliation":[{"name":"Naomi Balaban, StaphOff Biotech Inc., Dudley, MA 01571, USA"}],"role":[{"vocabulary":"crossref","role":"author"}]}],"member":"1968","published-online":{"date-parts":[[2026,6,22]]},"reference":[{"key":"ref_1","unstructured":"Miyagishima, K. (2015). WHO Estimates of the Global Burden of Foodborne Diseases: Food-Borne Disease Burden Epidemiology Reference Group 2007\u20132015, WHO Press."},{"key":"ref_2","doi-asserted-by":"crossref","first-page":"7","DOI":"10.3201\/eid1701.P11101","article-title":"Foodborne illness acquired in the United States\u2014Major pathogens","volume":"17","author":"Scallan","year":"2011","journal-title":"Emerg. Infect. Dis."},{"key":"ref_3","doi-asserted-by":"crossref","unstructured":"Fisher, E.L., Otto, M., and Cheung, G.Y. (2018). Basis of Virulence in Enterotoxin-Mediated Staphylococcal Food Poisoning. Front. Microbiol., 9.","DOI":"10.3389\/fmicb.2018.00436"},{"key":"ref_4","doi-asserted-by":"crossref","first-page":"8941","DOI":"10.1073\/pnas.86.22.8941","article-title":"Interaction of Staphylococcus aureus toxin \u201csuperantigens\u201d with human T cells","volume":"86","author":"Choi","year":"1989","journal-title":"Proc. Natl. Acad. Sci. USA"},{"key":"ref_5","doi-asserted-by":"crossref","first-page":"1775","DOI":"10.1016\/j.str.2005.08.015","article-title":"Structural basis of affinity maturation and intramolecular cooperativity in a protein-protein interaction","volume":"13","author":"Cho","year":"2005","journal-title":"Structure"},{"key":"ref_6","doi-asserted-by":"crossref","first-page":"422","DOI":"10.1128\/CMR.00104-12","article-title":"Staphylococcal and streptococcal superantigen exotoxins","volume":"26","author":"Spaulding","year":"2013","journal-title":"Clin. Microbiol. Rev."},{"key":"ref_7","doi-asserted-by":"crossref","unstructured":"Tuffs, S.W., Haeryfar, S.M.M., and McCormick, J.K. (2018). Manipulation of innate and adaptive immunity by staphylococcal superantigens. Pathogens, 7.","DOI":"10.3390\/pathogens7020053"},{"key":"ref_8","doi-asserted-by":"crossref","first-page":"975","DOI":"10.1007\/s10295-008-0372-3","article-title":"Staphylococcus aureus enterotoxin C2 mutants: Biological activity assay in vitro","volume":"35","author":"Hui","year":"2008","journal-title":"J. Ind. Microbiol. Biotechnol."},{"key":"ref_9","doi-asserted-by":"crossref","unstructured":"Rasooly, R., Do, P., He, X., and Hernlem, B. (2019). T cell Receptor V\u03b29 in Method for Rapidly Quantifying Active Staphylococcal Enterotoxin Type-A without Live Animals. Toxins, 11.","DOI":"10.3390\/toxins11070399"},{"key":"ref_10","doi-asserted-by":"crossref","first-page":"1857","DOI":"10.4315\/0362-028X.JFP-17-196","article-title":"Interleukin 2 Secretion by T Cells for Detection of Biologically Active Staphylococcal Enterotoxin Type E","volume":"80","author":"Rasooly","year":"2017","journal-title":"J. Food Prot."},{"key":"ref_11","doi-asserted-by":"crossref","first-page":"1467","DOI":"10.1021\/bi00857a032","article-title":"Purification and Partial Characterization of Enterotoxin C Produced by Staphylococcus aureus Strain 137","volume":"6","author":"Borja","year":"1967","journal-title":"Biochemistry"},{"key":"ref_12","doi-asserted-by":"crossref","unstructured":"Fang, R., Cui, J., Cui, T., Guo, H., Ono, H.K., Park, C.H., Okamura, M., Nakane, A., and Hu, D.L. (2019). Staphylococcal Enterotoxin C Is an Important Virulence Factor for Mastitis. Toxins, 11.","DOI":"10.3390\/toxins11030141"},{"key":"ref_13","doi-asserted-by":"crossref","unstructured":"Etter, D., Schelin, J., Schuppler, M., and Johler, S. (2020). Staphylococcal Enterotoxin C-An Update on SEC Variants, Their Structure and Properties, and Their Role in Foodborne Intoxications. Toxins, 12.","DOI":"10.3390\/toxins12090584"},{"key":"ref_14","doi-asserted-by":"crossref","unstructured":"Romano, A., Carrella, S., Rezza, S., Nia, Y., Hennekinne, J.A., Bianchi, D.M., Martucci, F., Zuccon, F., Gulino, M., and Di Mari, C. (2023). First Report of Food Poisoning Due to Staphylococcal Enterotoxin Type B in D\u00f6ner Kebab (Italy). Pathogens, 12.","DOI":"10.3390\/pathogens12091139"},{"key":"ref_15","doi-asserted-by":"crossref","first-page":"125","DOI":"10.1007\/978-1-4939-3344-0_11","article-title":"Monkey Feeding Assay for Testing Emetic Activity of Staphylococcal Enterotoxin","volume":"1396","author":"Seo","year":"2016","journal-title":"Methods Mol. Biol."},{"key":"ref_16","doi-asserted-by":"crossref","first-page":"324","DOI":"10.1016\/S0076-6879(88)65048-8","article-title":"Monkey feeding test for staphylococcal enterotoxin","volume":"165","author":"Bergdoll","year":"1988","journal-title":"Methods Enzymol."},{"key":"ref_17","doi-asserted-by":"crossref","first-page":"705","DOI":"10.1126\/science.2185544","article-title":"The staphylococcal enterotoxins and their relatives","volume":"248","author":"Marrack","year":"1990","journal-title":"Science"},{"key":"ref_18","unstructured":"Doyle, M.P. (1989). Staphylococcus aureus. Foodborne Bacterial Pathogens, Marcel Dekker Inc."},{"key":"ref_19","unstructured":"Lampel, K.A., Al-Khaldi, S., and Cahill, S.M. (2012). Pathogenic Bacteria, Bad Bug Book: Handbook of Foodborne Pathogenic Microorganisms and Natural Toxins."}],"container-title":["Microorganisms"],"original-title":[],"language":"en","link":[{"URL":"https:\/\/www.mdpi.com\/2076-2607\/14\/6\/1383\/pdf","content-type":"unspecified","content-version":"vor","intended-application":"similarity-checking"}],"deposited":{"date-parts":[[2026,6,22]],"date-time":"2026-06-22T10:48:32Z","timestamp":1782125312000},"score":1,"resource":{"primary":{"URL":"https:\/\/www.mdpi.com\/2076-2607\/14\/6\/1383"}},"subtitle":[],"short-title":[],"issued":{"date-parts":[[2026,6,22]]},"references-count":19,"journal-issue":{"issue":"6","published-online":{"date-parts":[[2026,6]]}},"alternative-id":["microorganisms14061383"],"URL":"https:\/\/doi.org\/10.3390\/microorganisms14061383","relation":{},"ISSN":["2076-2607"],"issn-type":[{"value":"2076-2607","type":"electronic"}],"subject":[],"published":{"date-parts":[[2026,6,22]]}}}