| Budget Amount *help |
¥3,900,000 (Direct Cost: ¥3,900,000)
Fiscal Year 2003: ¥1,300,000 (Direct Cost: ¥1,300,000)
Fiscal Year 2002: ¥2,600,000 (Direct Cost: ¥2,600,000)
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| Research Abstract |
In recent years, many new staphylococcal enterotoxins (SEs) have been reported. To investigate pathobiology and epidemiology of these newly identified SEs, we had undertaken studies shown in below.
1. To establish immunological detection methods of newly identified SEs, we have developed E.coli expression system of SEs to obtain large amount of each purified SE. These purified SEs were immunized to rabbits, and we obtained monospecific antibodies to each SEs.
2. We identified and characterized a novel staphylococcal entertoxin (SE)-like putative toxin, named SER. Nucleotides sequencing analysis of ser gene revealed that ser was most closely related to seg gene. The ser gene product, SER, was successfully expressed as recombinant protein in an Escherichia coli expression system, and rSER showed significant T cell stimulation activity. MHC class II molecules were required for T cell stimulation by SER. SER stimulated T cells bearing receptors Vβ 3, 11, 12, 13.2, and 14. These results sugge
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sted that SER acts as a superantigen. The SER production in ser harboring S.aureus strains was confirmed by Western blot analysis using anti-rSER antibody. Moreover, ser was seen to be encoded by at least two types of plasmids. In particular, one kind of plasmid encoding the ser gene has been known as sed-and sej-encoding pIB485-related plasmid (Infection and Immunity 71:6088, 2003 ; Infection and Immunity 72:3664, 2004).
3. To investigate the distribution of staphylococcal enterotoxin (SE) A to R (SEA to SER) genes (sea to ser) in Staphylococcus aureus, we have developed multiplex PCR for detection of 17 kinds of SE genes (sea to see, seg to selr) and TSST-1 gene. Using this multiplex PCR system, we investigated SE-genotype of Staphylococcus aureus 155 isolates obtained from bovine raw milk in Japan. In almost strains, these SE-genotypes were consistent with known SEs gene profile of Genomic islands or plasmids. These results suggest that SE-genotype of most S.aureus would be determined by combination of harboring mobile genetic elements. (manuscript in preparation).
4. We have investigated growth and enterotoxin (SEA) production of S.aureus in different concentrated skim milk (15, 25, 35%) at different storage (15, 25, 35℃). The growth rate of S.aureus was more rapid at 35℃ than 15 and 25℃ and SEA was produced most highly at 35℃ storage. Less
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