In addition to two known staphylococcal enterotoxin-like genes (selj and selr), two novel genes coding for two superantigens, staphylococcal enterotoxins S and T (SES and SET), were identified in plasmid pF5, which is harbored by food poisoning-related Staphylococcus aureus strain Fukuoka 5. This strain was implicated in a food poisoning incident in Fukuoka City, Japan, in 1997. Recombinant SES (rSES) specifically stimulated human T cells in a T-cell receptor V␤9-and V␤16-specific manner in the presence of major histocompatibility complex (MHC) class II ؉ antigen-presenting cells (APC). rSET also stimulated T cells in the presence of MHC class II ؉ APC, although its V␤ skewing was not found in reactive T cells. Subsequently, we examined the emetic activity of SES and SET. We also studied SElR to determine emetic activity in primates. This toxin was identified in previous studies but was not examined in terms of possession of emetic activity for primates. rSES induced emetic reactions in two of four monkeys at a dose of 100 g/kg within 5 h of intragastric administration. In one monkey, rSET induced a delayed reaction (24 h postadministration) at a dose of 100 g/kg, and in the other one, the reaction occurred 5 days postadministration. rSElR induced a reaction in two of six animals within 5 h at 100 g/kg. On this basis, we speculate that the causative toxins of vomiting in the Fukuoka case are SES and SER. Additionally, SES, SER, and SET also induced emesis in house musk shrews as in the monkeys.Staphylococcus aureus produces a variety of superantigenic toxins (SAGTs), which selectively activate a vast number of T cells, depending on V␤ elements in the ␤ chain of a T-cell receptor (TCR), in direct association with major histocompatibility complex (MHC) class II molecules on antigen-presenting cells (APC) (14, 31). Staphylococcal SAGTs can be divided into three large groups and one minor group on the basis of similarity of amino acid sequences (31). Most toxins of the three groups, including staphylococcal enterotoxins A and B (SEA and SEB), exhibit strong emetic activity in primates (4, 16, 25); toxic shock syndrome toxin-1, grouped as the minor group, does not possess emetic activity in primates (14, 31). It is noteworthy that toxins designated SE-like toxins, such as SElP and SElR, which either have not been examined for emetic activity or have been reported not to have emetic activity, have been discovered in S. aureus strains (12,13,20,27). S. aureus strain Fukuoka 5 was isolated from food as the causative microbe in a food poisoning outbreak in Fukuoka City, Japan, in 1997, although this strain did not carry any well-recognized SAGT genes with emetic activity (19). Subsequently, Omoe et al. (19) discovered, using a plaque hybridization with a seg probe, that 2.8 kbp of the EcoRI fragment of plasmid pF5, carried by Fukuoka 5, carries two genes, a novel SAGT gene designated selr and a previously reported gene, selj (33).We undertook research to explore, using a PCR walking technique, whether there were addition...

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g Staphylococcal enterotoxins (SEs) are a common causative agent of food poisoning. Recently, many new SE-like (SEl) toxins have been reported, although the role of SEls in food poisoning remains unclear. In this study, the emetic potentials of SElK, SElL, SElM, SElN, SElO, SElP, and SElQ were assessed using a monkey-feeding assay. All the SEls that were tested induced emetic reactions in monkeys at a dose of 100 g/kg, although the numbers of affected monkeys were significantly smaller than the numbers that were affected after consuming SEA or SEB. This result suggests that these new SEs may play some role in staphylococcal food poisoning. Staphylococcal enterotoxins (SEs) are exotoxins that cause staphylococcal food poisoning in humans worldwide (1-4). Several classical types (SEA, SEB, SEC, SED, and SEE) have been characterized (1, 2, 5). Recently, SEG, SEH, SEI, SER, SES, and SET were identified as potential agents of food poisoning (6-10). In addition, new proteins (SE-like toxin J [SElJ], SElK, SElL, SElM, SElN, SElO, SElP, SElQ, SElU, SElV, and SElX) with amino acid sequences that are similar to those of the above-mentioned SEs have been identified (11)(12)(13)(14)(15)(16)(17)(18)(19). SEs are also known to be members of the superantigen family (3,20). During the last few decades, numerous studies have been conducted on the nature of SEs and their superantigenic activities (3,20). However, the emetic activities of these toxins have not been studied. To better understand the etiologic nature of staphylococcal food poisoning, the emetic potentials of SEls should be evaluated in a primate model.We demonstrated the emetic activities of SElK, SElL, SElM, SElN, SElO, SElP, and SElQ using a primate model (cynomolgus monkeys); the number of vomiting events, the time until the first vomiting event (latency period), and behavioral changes were recorded for each animal. We compared the emetic activities of classical and new SEs as well as the activities of three large groups of SEs that were grouped according to the similarity of their amino acid sequences. MATERIALS AND METHODS Preparation of SEls.To construct the SElK, SElL, SElM, SElN, SElO, and SElQ expression plasmids, PCR primers were designed to amplify the gene fragment corresponding to the mature forms of these SEls (Table 1). The selk and selq genes were amplified from genomic DNA of the Staphylococcus aureus S6 strain, which harbors the sea, seb, selk, and selq genes (21). The sell gene was amplified from genomic DNA of the S. aureus bov1117 strain (harboring sea, sec, sell, sed, selj, selr, and tst-1), isolated from cow's milk, and the selm, seln, and selo genes were amplified from genomic DNA of the S. aureus Fukuoka 2 strain (21, 22). The PCR products were digested with BamHI and EcoRI or SalI and then subcloned into a pGEX6P-1 glutathione S-transferase (GST) fusion expression vector. These clones were designated pKKX (including the selk gene), pKLX (including the sell gene), pKMX (including the selm gene), pKNX (including the seln gene), pKOX (includin...

We analyzed the in vivo dynamics of peritoneal exudate cells (PECs) in mice injected with group A streptococcus (GAS). A live low-virulence strain, as well as heat-killed low- and high-virulence strains, significantly increased the number of PECs (primarily neutrophils), whereas a live high-virulence strain did not. When coinjected with thioglycollate, the live high-virulence strain, as well as most other GAS strains, suppressed the ability of thioglycollate to induce neutrophil exudation. This suppression was due to a cytocidal effect of GAS on exuded neutrophils rather than an inhibition of neutrophil migration. In addition, GAS enhanced the apoptosis of neutrophils. These cytocidal effects were significantly reduced by the deletion of functional streptolysin S from GAS. Our findings suggest that, in addition to the production of antiphagocytic factors and survival inside phagocytes, GAS uses a more aggressive method--the elimination of neutrophils--to evade the host's innate immune system.

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