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Rapid detection of S. aureus and coliforms using firefly luciferase Satoshi Fukuda*1; I Masuda-Nishimura1; H Igarashi2; S Igimi3; H Tatsumi1 (1) R & D Division, Kikkoman Corp., 399 Noda, Noda City, Chiba 278-0037, Japan; (2) Kokusaigakuin Saitama Junior College, 2-5 Kishiki-Cho, Ohmiya City, Saitama 330-8548, Japan, ; (3) National Institute of Infectious Disease, 1-23-1 Toyama, Shinjuku-ku, Tokyo 162-8640, Japan      *(sfukuda@mail.kikkoman.co.jp) The number of reports documenting outbreak of food borne illness caused by pathogenic microorganisms is now increasing in all countries. The need for a rapid and easy-to-use detection method for food pathogens with high sensitivity led to the development of bioluminescent assays. We have developed a bioluminescent enzyme immunoassay (BLEIA) using biotinylated firefly luciferase for the detection of protein A in Staphylococcus aureus and a bioluminescent assay for that of beta-galactosidase in coliforms. The BLEIA enabled us to detect a protein A at 1 pg/ml and 103 CFU/ml levels of S. aureus. After 7 hours of cultivation followed, all of 57 S. aureus strains tested were detected in this BLEIA. The bioluminescent assay of beta-galactosidase has been developed using a luciferin derivative, D-luciferin-O-beta-galactopyranoside (LuGal). The detection limit for beta-galactosidase was 3 x 10-20 mol/assay, which was approximately 50-fold more sensitive than the test using a fluorescent substrate. Observations made after 7-h of cultivation of 26 coliform strains followed by a 10-min enzyme assay using LuGal were comparable to those made after a 22 to 24 hours cultivation of them on desoxycholate agar and Chromocult agar. [Poster: fukuda.satosh.86902]

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