Effect of acid stress on the physiology of biofilm cells of Streptococcus mutans

Streptococcus mutans is a component of the dental plaque biofilm and an important aetiological agent in dental caries. Although this organism growing in the suspended (planktonic) state has been well characterized, relatively little is known about its physiology in biofilms, particularly in the acidic environments associated with caries development. The authors determined the effect of biofilm age (1-5 days) and cell density on selected metabolic properties under conditions of glucose limitation in a biofilm-chemostat at pH 7(.)5 and compared these baseline values with those of 3 day biofilms subjected to acid stress. Biofilm cell biomass more than doubled over the 5 day experimental period under baseline conditions, with the glycolytic rate, glucose uptake, glucose-PTS (phosphotransferase system) activity and protein synthesis maximum at 1-2 days. DNA and RNA synthesis increased for the first 3 days before decreasing in the 5 day biofilms, while H+/ATPase activity was higher in 5 day biofilms than 1 day biofilms, with overall activity 5-13-fold higher per cell unit than in the associated planktonic cells. Glucose pulsing (50 mM final concentration) for three consecutive days without pH control for 5 h(pH 4(.)39 +/- 0(.)02) resulted in a progressive decrease in planktonic cell numbers; however, the rate of acid formation and glucose utilization in the chemostat by these cells increased per cell unit. Assays for carbohydrate metabolism in the latter cells showed increased activity, as did an assay for H+/ATPase (8-fold); however, DNA, RNA and protein synthesis were repressed (0(.)3-0(.)7-fold). Although the 3 day biofilm viable cell counts declined by 51% on glucose pulsing, all the physiological parameters measured by cell unit increased in activity, with notable increases in RNA and protein synthesis (4(.)6-7(.)6-fold). The results indicate that the maintenance of intracellular pH homeostasis is the basis of the enhanced physiological status and acid tolerance of biofilm cells.