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Use of Tetrazolium Dyes in an Agar Medium for Differentiation of Streptococcus Lactis and Streptococcus Cremoris¹

Department of Microbiology, Oregon State University, Corvallis

ABSTRACT

A medium consisting of 0.5% Tryptone, 0.5% yeast extract, 0.2% L-arginine, 1.5% agar, 0.2% K₂HPO₄, 0.05% glucose, and 0.005% 2,3,5-triphenyltetrazolium chloride (TTC) has been developed which may be used to distinguish Streptococcus lactis from Streptococcus cremoris. S. lactis organisms, by virtue of their ability to produce ammonia from arginine, formed red colonies; colonies of S. cremoris were white. The incorporation of tetrazolium blue or tetrazolium red into the basal arginine medium in place of TTC produced analogous results.

To insure that the production of colored colonies by S. lactis was dependent on an unimpaired degradation of arginine, catabolism of the amino acid by this organism was studied. Mutants of S. lactis Strain 27, which lacked the ability to produce ammonia from arginine, were obtained by ultraviolet light irradiation and selection on arginine-TTC agar. Whole cells and cell-free extracts of mutant and wild type organisms were compared for their abilities to produce CO₂ from arginine and citrulline. Cell-free extracts of the wild type organism produced more than 9 µmoles of CO₂ from 10 µmoles of either arginine or citrulline; cell-free extracts of the mutant produced about 6 µmoles of CO₂ from either substrate. Whole cells of the wild type organism produced nearly 10 µmoles of CO₂ from 10 µmoles of arginine, but only 3 µmoles of CO₂ from 10 µmoles of citrulline; whole cells of the mutant produced less than 1 µmole of CO₂ from either substrate.

No enzymes for arginine degradation were found in cell-free extracts of S. cremoris Strain 144.

¹ Technical paper no. 1609 Oregon Agricultural Experiment Station. Supported in part by research grants from the National Science Foundation (G14703) and the American Dairy Association.

² Boyce Thompson Plant Research Institute, Yonkers, New York.

³ Department of Food Science and Technology, Oregon State University, Corvallis.

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