It was believed that food poisoning in Osaka in 2000 was due to small amounts of staphylococcal enterotoxin A (SEA) in reconstituted milk. Results of this study clearly indicate that SEH was also present in the raw material of reconstituted milk, indicating that the food poisoning was caused by multiple staphylococcal enterotoxins.Exotoxins produced by Staphylococcus aureus are enterotoxins and elicit an emetic response. Fourteen staphylococcal enterotoxins, designated staphylococcal enterotoxin A (SEA), SEB, SEC, SED, SEE, SEG, SEH, SEI, SEJ, SEK, SEL, SEM, SEN, and SEO, have been identified (2,4,7,9,10,11,13). However, only the first five (SEA through SEE) can be detected using a commercial kit. Moreover, as commercial kits are generally employed to detect the presence of staphylococcal enterotoxins in food poisoning, SEG, SEH, or SEI was rarely detected and implicated as the source of food poisoning.A mass outbreak of food poisoning caused by the consumption of reconstituted milk occurred in Osaka, Japan, in June 2000, and more than 10,000 cases were reported (6). A small amount of SEA and sea gene were detected in the reconstituted milk and the skim milk powder, which was the raw material for the reconstituted milk (6). In an outbreak of food poisoning in United States, caused by SEA present in chocolate milk, 200 ng or less SEA was presumed to be the cause (3). Although it was considered that the outbreak of food poisoning in Osaka could have been caused by SEA, the quantity of SEA detected, i.e., approximately 80 ng, was insufficient to cause food poisoning on such a large scale. Hence, we investigated the possibility of staphylococcal enterotoxins other than SEA as the cause of the outbreak.We got 11 batches of skim milk powder, which were the raw material of the food causing the outbreak. Ten of these batches were manufactured on 1 April 2000 and designated SM1, SM100, SM200, SM300, SM400, SM500, SM600, SM700, SM800, and SM830 in the order of manufacture, and the 11th batch, manufactured on 10 April 2000, was designated SM500 (10/April).Since the manufacturing process of skim milk required heat treatment for 3 s at 130°C, viable S. aureus strains were not isolated from the skim milk powder samples. However, Gram staining of these samples revealed the presence of gram-positive cocci in large numbers. The skim milk solution (10%, wt/vol) reconstituted in sterile water was centrifuged at 12,000 rpm for 3 min, and the DNA was extracted from the precipitate using a DNeasy tissue kit (QIAGEN GmbH, Hilden, Germany). PCR was performed to detect the sea, seb, sec, sed, see, seg, seh, and sei genes (5, 8). The seb, sec, sed, and see genes were not detected in any of the samples, the sea and seh genes were detected in 10 samples, and the seg and sei genes were detected in 7 samples ( Fig. 1 and Table 1).Since the skim milk samples that contained the sea gene also showed the presence of the seh gene and the seh gene showed stronger expression of enterotoxin compared to that shown by the seg or sei gene (8), there wa...

ABSTRACT. A large-scale outbreak of food poisoning caused by consumption of skimmed milk powder contaminated with staphylococcal enterotoxin A (SEA) occurred in Japan. No viable Staphylococcus aureus was detected in the skimmed milk powder, however, sea and nuc genes of S. aureus were detected in it by PCR. The number of S. aureus in skimmed milk powder was estimated by quantitative real-time PCR.

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