Response of Freshly Isolated Strains of Streptococcus mutans and Streptococcus mitior to Change in pH in the Presence and Absence of Fluoride During Growth in Continuous Culture

Abstract

A study was undertaken to compare the effects of pH and fluoride on the growth and metabolic properties of Streptococcus mutans 2452 and Streptococcus mitior 572, strains recently isolated from 8-year-old school children and grown in continuous culture with a glucose limitation. Each experiment had four consecutive stages of growth: (i) pH 7.0, (ii) no pH control, (iii) pH 7.0, and (iv) no pH control plus 50 μg of fluoride per ml in the medium. At a dilution rate (D) of 0.13 h ⁻¹, cells of S. mitior possessed high glycolytic activity at pH 7.0 in the initial stage, but were washing out of the chemostat within 24 h after the pH control was shut off and the pH fell to 5.1. Once the culture was reestablished at pH 7.0, fluoride (50 μg/ml) was added to the medium and the pH control was again turned off. Whereas cell numbers fell from 24.0 × 10⁸ to 0.9 × 10⁸/ml within 24 h, the culture remained relatively constant during the following 6 days despite the fall in pH to 5.4. The cells from this culture also maintained an intermediate glycolytic rate of 0.44 μmol mg⁻¹ min⁻¹. The cells in this latter stage developed phenotypic resistant to fluoride at concentrations up to 16 mM. Growth of S. mitior at D = 0.034 h⁻¹ resulted in a slower response to environmental change such that cells were able to grow to pH values as low as 5.2 in the absence of fluoride. In contrast to S. mitior, S. mutans 2452 under the same conditions at D = 0.13 h⁻¹ grew to higher cell numbers and higher yields and was able to maintain significant cell numbers to pH 4.8 once the pH control was shut off in the presence and absence of fluoride. S. mutans had 40% less glycolytic activity but was fourfold more resistant to fluoride at the start of the experiment, and cells were shown to adapt to growth at low pH and to fluoride at levels as high as 20 mM. This fluoride resistance by freshly isolated S. mutans 2452 was significantly higher than that of S. mutans DR0001 grown under identical conditions in the chemostat. S. mutans DR0001 is a strain which has been subcultured in vitro for several years. This study demonstrated that S. mutans 2452 was more aciduric than S. mitior 572 and, unlike the latter organism, could grow at pH values below 5.1. The addition of fluoride to the medium stabilized the S. mitior culture in the absence of pH control, indicating that whereas fluoride does suppress growth and glycolytic activity it also results in higher environmental pH values, which permit the survival of the less aciduric bacteria.

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