As stated earlier, individuals who frequently consume sugar in their diet generally have elevated levels of cariogenic bacteria such as mutans streptococci and lactobacilli in their plaque, and are at greater risk of dental caries. In animal studies or epidemiological surveys of humans, it can never be determined whether the rise in cariogenic bacteria is due to the sudden availability of sugar
per se (
e.g. because of more efficient sugar transport systems in these bacteria), or is a response to the inevitable conditions of low pH following sugar consumption. Exploitation of the unique benefits of parameter control in the chemostat, coupled with the reproducibility of a defined mixed culture inoculum, enabled these linked effects to be separated for the first time [
28]. Two mixed culture chemostats were inoculated with 9 or 10 species (representative of those in health and disease) in a growth medium at pH 7.0 in which mucin was the main source of carbohydrate; under these conditions,
S. mutans and
Lactobacillus rhamnosus were noncompetitive and made up <1% of the total microflora (Table
1). Once the consortia were stably established, both chemostats were pulsed daily for ten consecutive days with a fermentable sugar (glucose). In one chemostat, the pH was maintained automatically throughout the study at neutral pH (as is found in the healthy mouth) in order to determine the effect of the addition of a fermentable sugar on culture stability, while in the other the pH was allowed to fall by bacterial metabolism for six hours after each pulse (as occurs
in vivo); the pH was then returned to neutrality for 18 hours prior to the next pulse [
28]. Daily pulses of glucose for 10 consecutive days at a constant pH 7.0 had little or no impact on the balance of the microbial community, and the combined proportions of
S. mutans and
L. rhamnosus stayed at ca. 1% of the total microflora (Table
1). In contrast, however, when the pH was allowed to change after each pulse, there was a progressive selection of the cariogenic (and acid-tolerating) species at the expense of bacteria associated with dental health. After the final glucose pulse, the community was dominated by species implicated in caries (
S. mutans and
L. rhamnosus comprised ca. 55% of the microflora) [
28]. When this study was repeated, but the pH fall was restricted after each glucose pulse to either pH 5.5, 5.0, or 4.5 in independent experiments [
29], a similar enrichment of cariogenic species at the expense of healthy species was observed, but their rise was directly proportional to the extent of the pH fall (Table
1). Collectively, these studies showed conclusively that it was the low pH generated from sugar metabolism rather than sugar availability that led to the breakdown of microbial homeostasis in dental plaque. This finding has important implications for caries control and prevention; the data suggest that the selection of cariogenic bacteria could be prevented if the pH changes following sugar metabolism could be reduced (see later).
Table 1
The effect of glucose and low pH on the stability of a microbial community. A mixed culture of 9 oral bacteria (representative of those found in health and disease) were grown in a mucin-based medium at a constant pH 7.0. In independent experiments, the culture was pulsed daily on 10 consecutive days with 28 mM glucose. In one culture the pH was maintained throughout the pulsing at pH 7.0, while in other experiments the pH was allowed to fall by bacterial metabolism for 6 hours to pre-set levels of pH 5.5, 5.0 or 4.5. In a final study, the pH was allowed to fall without any pH control imposed [28,29]. Viable counts were determined on selective and non-selective media; proportions are shown after the final glucose pulse for two species implicated in caries and for two species associated with sound enamel
Streptococcus mutans
| 0.3 | 1.0 | 4.2 | 7.9 | 9.6 | 18.9 |
Lactobacillus rhamnosus
| 0.1 | 0.2 | 2.4 | 6.2 | 13.9 | 36.1 |
Streptococcus gordonii
| 28.3 | 25.0 | 2.6 | 6.9 | 2.7 | 0.2 |
Fusobacterium nucleatum
| 15.2 | 9.5 | 7.4 | 2.3 | 2.7 | <0.001 |