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Antimicrobial Agents and Chemotherapy, August 1998, p. 1906-1910, Vol. 42, No. 8
0066-4804/98/$04.00+0
Copyright © 1998, American Society for Microbiology. All rights reserved.

Generation of Bioluminescent Streptococcus mutans and Its Usage in Rapid Analysis of the Efficacy of Antimicrobial Compounds

Vuokko Loimaranta,1,* Jorma Tenovuo,1 Leeni Koivisto,2,dagger and Matti Karp2

Institute of Dentistry and Turku Immunology Centre1 and Department of Biotechnology,2 University of Turku, FIN-20520 Turku, Finland

Received 30 September 1997/Returned for modification 31 January 1998/Accepted 27 April 1998

The oral bacterium Streptococcus mutans was transformed by electroporation with a shuttle vector (pCSS945) containing insect luciferase gene from a click beetle (Pyrophorus plagiophthalamus) resulting in a bioluminescent phenotype. This S. mutans strain was used in experiments in which light emission was used as a rapid and, compared to conventional CFU counting, more convenient means of estimating the effects of various antimicrobial treatments. The basic parameters affecting in vivo light production by the strain were studied. It was found that pH 6.0 was optimal for incorporation of the substrate D-luciferin for the luciferase reaction. The optimum concentration of D-luciferin was approximately 150 µM at room temperature. Under optimum conditions the light emission in vivo increased rapidly to a constant level and thereafter had a decay of 0.6%/min when logarithmic-growth-phase cells were used. The light emission closely paralleled the numbers of CFU, giving a detectable signal from 30,000 cells and having a dynamic measurement range over 4 log CFU/relative light unit. The cells were treated with various antimicrobial agents, and the emitted bioluminescence was measured. With the bioluminescent measurements, the results were obtained within hours compared to the days required for agar plates, and also, the kinetics of the antibacterial actions could be followed. Thus, the light emission was found to be a reliable, sensitive, and real-time indicator of the bacteriostatic actions of the antimicrobial agents tested.

* Corresponding author. Mailing address: Institute of Dentistry and Turku Immunology Centre, University of Turku, Lemminkäisenkatu 2, FIN-20520 Turku, Finland. Phone: 358-2-3337918. Fax: 358-2-3338356. E-mail: vuokko.loimaranta{at}utu.fi.

dagger Present address: Department of Oral Medical and Surgical Sciences, Faculty of Dentistry, University of British Columbia, Vancouver, British Columbia, Canada V6T 1Z3.

Antimicrobial Agents and Chemotherapy, August 1998, p. 1906-1910, Vol. 42, No. 8
0066-4804/98/$04.00+0
Copyright © 1998, American Society for Microbiology. All rights reserved.


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