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in Dental Research

Microbial Ecology of Dental Plaque and its Significance in Health and Disease
P.D. Marsh
ADR 1994 8: 263
DOI: 10.1177/08959374940080022001
The online version of this article can be found at:
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MICROBIAL ECOLOGY OF DENTAL PLAQUE

AND ITS SIGNIFICANCE IN HEALTH AND DISEASE
P.D. MARSH

Pathology Division
PHLS Centre for Applied Microbiology and Research
Porton Down
Salisbury SP4 OJG
England
Adv Dent Res 8(2):263-271, July, 1994

AbstractDental plaque forms naturally on teeth and is of

benefit to the host by helping to prevent colonization by
exogenous species. The bacterial composition of plaque remains
relatively stable despite regular exposure to minor
environmental perturbations. This stability (microbial
homeostasis) is due in part to a dynamic balance of both
synergistic and antagonistic microbial interactions. However,
homeostasis can break down, leading to shifts in the balance of
the microflora, thereby predisposing sites to disease. For
example, the frequent exposure of plaque to low pH leads to
inhibition of acid-sensitive species and the selection of
organisms with an aciduric physiology, such as mutans
streptococci and lactobacilli. Similarly, plaque accumulation
around the gingival margin leads to an inflammatory host
response and an increased flow of gingival crevicular fluid.
The subgingival microflora shifts from being mainly Grampositive to being comprised of increased levels of obligately
anaerobic, asaccharolytic Gram-negative organisms. It is
proposed that disease can be prevented or treated not only by
targeting the putative pathogens but also by interfering with the
processes that drive the breakdown in homeostasis. Thus, the
rate of acid production following sugar intake could be reduced
by fluoride, alternative sweeteners, and low concentrations of
antimicrobial agents, while oxygenating or redox agents could
raise the Eh of periodontal pockets and prevent the growth and
metabolism of obligately anaerobic species. These views have
been incorporated into a modified hypothesis (the "ecological
plaque hypothesis") to explain the relationship between the
plaque microflora and the host in health and disease, and to
identify new strategies for disease prevention.

This manuscript was presented at a Symposium entitled

"Mechanisms and Agents in Preventive Dentistry", held
October 28-November 7, 1992, in Chester, England, under
the auspices of the Council of Europe Research Group on
Surface and Colloid Phenomena.

he mouth is similar to other sections of the digestive

tract in having a resident microflora that develops
naturally, and which has a characteristic composition.
Owing to differences in local environmental conditions,
however, the microflora of mucosal surfaces differs in
composition from that of dental plaque. For similar reasons,
the plaque microflora varies in composition at distinct
anatomical sites on the toothfor example, in fissures, on
approximal surfaces, and in the gingival crevice. The resident
microflora of a site is of benefit to the host by acting as part of
the host defenses by preventing colonization by exogenous
(and often pathogenic) micro-organisms ("colonization
resistance"; van der Waaij et al., 1971).
The early colonizers of the tooth surface include members
of the genera Streptococcus, Actinomyces, Haemophilus,
Neisseria, and Veillonella (Liljemark et al., 1986; Nyvad and
Kilian, 1987). These bacteria adhere to the acquired enamel
pellicle by specific and non-specific molecular interactions
between adhesins on the cell and receptors on the surface
(Gibbons, 1989; Busscher etal, 1992). Once established, the
microflora at a site remains relatively stable over time despite
regular minor perturbations to the oral environment (Marsh,
1989). This stability (termed "microbial homeostasis") stems
not from any metabolic indifference among the components of
the microflora, but rather results from a dynamic balance of
microbial interactions, including both synergism and
antagonism (Sanders and Sanders, 1984).
MECHANISMS INVOLVED IN MAINTAINING
MICROBIAL HOMEOSTASIS

It has been proposed that the ability to maintain homeostasis

within a microbial community increases with its species
diversity (Alexander, 1971). In dental plaque, the diversity of
the microflora is enhanced by the development of food chains
between bacterial species, and their use of complementary
metabolic strategies for the catabolism of endogenous nutrients,
such as glycoproteins and proteins. Individual species possess
different but overlapping patterns of enzyme activity so that
certain mixed cultures of oral bacteria can synergistically
degrade complex host molecules (van der Hoeven and Camp,
1991). Several food chains have been recognized among
plaque bacteria (Mikx and van Campen, 1982; Grenier and
Mayrand, 1986), such as the utilization of lactic acid by
Veillonella spp. and succinate by spirochetes.
Antagonism is also a major mechanism in maintaining
microbial homeostasis in plaque. Bacteriocins and bacteriocinlike substances are produced by many genera of oral bacteria
(James and Tagg, 1988; Marsh, 1989). The precise benefit of
bacteriocins is not known for certainty, but animal studies have
shown that their production can confer an ecological advantage
on an organism during colonization (van der Hoeven and

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263

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ADV DENT RES JULY 1994

MARSH

TABLE 1
EFFECTS OF GLUCOSE PULSES, LOW pH, AND FLUORIDE ON THE STABILITY
OF A MIXED CULTURE OF NINE ORAL B ACTERIAa
Percentage Viable Count
After 10 glucose pulses:
Pre-pulsing

With pH control
(pH7)

Without pH control

Without pH control,
with NaF

28.3

25.0

0.2

0.002

S. oralis

15.2

16.9

1.3

4.6

S. mutans

0.3

1.0

18.9

0.2

A. viscosus

0.1
0.1

13.1
0.2

2.3

0.4

0.1

0.01

36.1
NDb

2 x lO 5

9.8
31.0
15.2

28.7

41.4

Bacterium
S. gordonii

L. casei
N. subflava
V. dispar
P. intermedia
F. nucleatum

5.6
9.5

36.5
57.8
4

6 x lO"
2 x 105

0.5
0.2

Final pH
7.0
7.0
3.83
4.49
The mixed culture was pulsed on 10 consecutive days to give 28 mmol/L glucose, with or without 1 mmol/L NaF. The pH was
either maintained automatically throughout at pH 7.0 0.1 or was allowed to fall for six h following each pulse before being
returned to pH 7.0 for 18 h prior to the next pulse.
b
ND = not detected.
(Data from Bradshaw et ai, 1989a, 1990)

Rogers, 1979). Other inhibitory factors produced by plaque

bacteria include organic acids (Donoghue and Tyler, 1975),
H2O2 (Holmberg and Hallander, 1973), and enzymes (Baba,
1986).
The production of such inhibitory substances might also be
a major factor in determining the composition of the plaque
microflora. It was found that subgingival plaque samples from
healthy subjects contained organisms that could inhibit the
growth of several periodontopathogens (Hillman and
Socransky, 1989). In contrast, plaque from sites with localized
juvenile periodontitis (LJP) or refractory periodontitis invariably
lacked organisms producing inhibitors. Subsequent studies
identified some of the antagonistic bacteria as Streptococcus
sanguis, and the inhibitor as H2O2 (Hillman and Socransky,
1989). Such interactions can also contribute to colonization
resistance. S. salivarius can produce "enocin" with activity
against Lancefield Group A streptococci (Sanders and Sanders,
1982) and may prevent colonization of this pathogen in the
mouth in a manner similar to that proposed for the pharynx.

PLAQUE MICROFLORA AND DISEASE

Plaque accumulates preferentially at stagnant or retentive
sites, unless removed by diligent oral hygiene. As plaque mass
increases, saliva is less able to penetrate plaque and protect
enamel. Microbial homeostasis can break down, and major
shifts in the composition of the microflora can occur.
For example, the frequent consumption of fermentable
dietary carbohydrates is associated with an increased risk of

dental caries (Loesche, 1986). Such diets lead to a rise in the

proportions of mutans streptococci and lactobacilli, with a
concomitant fall in levels of other streptococci, especially
members of the Streptococcus oralis group, which include S.
sanguis, S. oralis, and S. mitis (de Stoppelaar et ai, 1970;
Dennis^ a/., 1975;Staat6tfa/., 1975;Minahefa/., 1985). The
metabolism of plaque also changes from a heterofermentative
pattern to one in which sugars are converted primarily to lactic
acid.
Gingivitis is associated with the accumulation of plaque
around the gingival margin. The host mounts an inflammatory
response to this microbial challenge, and the flow of gingival
crevicular fluid (GCF) is increased. The composition of
subgingival plaque shifts away from a streptococci-dominated
microflora (Slots, 1977) to one with higher levels ofActinomyces
spp. and an increase in capnophilic and obligately anaerobic
bacteria such as Capnocytophaga, Fusobacterium, and
Prevotella species (Savitt and Socransky, 1984; Moore et aL,
1987). Gingivitis may lead to more advanced forms of
periodontal disease, in which the microflora can become even
more diverse. Depending on the type of disease, bacteria
belonging to the genera Actinobacillus, Campy lobacter,
Selenomonas, Treponema, and Wolinella may be isolated.
Tissue damage can result directly from the activity of the
subgingival microflora and indirectly from the release of
lysosomal enzymes during phagocytosis or to the production
of cytokines that stimulate resident connective tissue cells to
release metalloproteinases (Reynolds, 1994).

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VOL.

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265

MICROBIAL ECOLOGY OF DENTAL PLAQUE

TABLE 2

TABLE 3

COMPONENTS OF GINGIVAL CREVICULAR FLUID

PREVENTION STRATEGIES

THAT MIGHT AFFECT THE COMPOSITION

AND THE ECOLOGICAL PLAQUE HYPOTHESIS

OF THE SUBGINGIVAL MICROFLORA

Host Defenses
Novel Nutrients
Hemin, iron
IgG
IgA
IgM
Complement

1. Reduced sugar/low pH challenge

(a) Fluoride
(b) Sugar substitutes

Albumin

(c) Stimulation of saliva flow

a-2-globulin
Transferrin

(d) Antimicrobial agents (sub-MIC)

2. Altered subgingival environment

Hemopexin

(a) Oxygenating or redox agents

Hormones

(b) Anti-inflammatory agents

B and T lymphocytes

Haptoglobin

(c) Antimicrobial agents (sub-MIC)

Neutrophils

Hemoglobin

Macrophages

Proteins, glycoproteins

3. Replacement therapy
(a) Pre-emptive colonization
(b) Competitive displacement

POTENTIAL FACTORS DISRUPTING MICROBIAL

HOMEOSTASIS IN DENTAL PLAQUE
Observations from clinical and laboratory studies have enabled
potential factors to be recognized that may disrupt microbial
homeostasis in plaque. Several in vitro model systems have
been devised to study interactions among the oral microflora,
the environment, and the host. Some have tried to simulate
important physical aspects of the oral cavity by incorporating
surfaces for biofilm formation (the "artificial mouth"; see
Tatevossian, 1991), while an alternative approach has been to
exploit the unique advantages of the chemostat to grow mixed
cultures of oral bacteria under a range of defined but controllable
conditions (Marsh, 1993). Individual parameters can be varied
independently in the chemostat, and their effects on the
composition and metabolism of the culture can be determined,
so that cause-and-effect relationships can be established. Several
factors that may be responsible for the transition of the oral
microflora from having a commensal to a pathogenic
relationship with the host have been identified by means of
continuous-culture techniques.
Mixed-culture chemostat studies have been performed to
distinguish whether the increases in mutans streptococci and
lactobacilli following repeated sugar intake are due to
differences in the ability of oral bacteria to (a) transport and
catabolize sugars, or to (b) tolerate and grow in the low-pH
environment so generated. A system for growing nine oral
bacteria, stably and reproducibly, in a chemostat at constant
temperature (37C) and pH (7.0 0.1) in a habitat-simulating
medium has been developed (see Marsh, 1993). The effect on
the balance of the mixed culture of pulsing on 10 consecutive
days with glucose, either with or without pH control, was
determined (Table 1). It was found that low pH rather than the
availability of carbohydrate per se was the factor driving the
selection of potentially cariogenic species. This selection was
at the expense of acid-sensitive species, some of which are
associated more with oral health (Bradshaw et aL, 1989a). The

experiment was repeated to determine if there was a "critical

pH" for this breakdown in homeostasis to occur. The culture
was again pulsed with glucose in three replicate experiments
but in which the pH was allowed to fall only to fixed values of
pH 5.5,5.0, or 4.5, respectively. The microbial community was
disrupted irreversibly only when the pH fell regularly below
pH 5.0 (Bradshaw et aL, 1989b), and the predominant species
always became Streptococcus mutans, Lactobacillus casei,
and Veillonella dispar. These three species have been associated
with nursing caries (Milnes and Bowden, 1985) and progressing
caries (Boyar and Bowden, 1985) in humans. Pure culture
studies have also shown that the growth of these three species
is less sensitive to low pH than other oral bacteria (Harper and
Loesche, 1984; Bradshaw et aL, 1989a). Furthermore,
mouthrinsing with acidic buffers (pH 3.9) was found to increase
the proportions of mutans streptococci in human fissure plaque
(Svanberg, 1980). Collectively, these findings show that the
selection of cariogenic species following regular sugar
consumption is likely to be a consequence of their aciduric
physiology, which enables them to compete successfully at
low pH.
In periodontal diseases, the redox potential of pockets is
lower than that at healthy sites (Kenney and Ash, 1969). The
inflammatory host response also leads to increased secretion of
GCF and a small rise in local pH from just below neutrality in
health to around pH 7.5 during disease (Eggert et aL, 1991).
GCF not only delivers components of the host defenses but also
provides a continuous supply of proteins, glycoproteins, and
co-factors that can act as novel nutrients for bacteria, especially
asaccharolytic and obligately anaerobic species (Table 2). In
an early longitudinal clinical study, black-pigmented anaerobes
increased from 0.01 to 0.2% of the subgingival flora when
gingivitis progressed to a bleeding stage (Loesche and Syed,
1978). This is noteworthy, because these organisms require
hemin for growth, and this co-factor can be derived from the

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266

ADV DENT RES JULY 1994

MARSH

ECOLOGICAL PLAQUE HYPOTHESIS

AND PREVENTION OF CARIES

FERMENTABLE
SUGAR

REMINERALIZATION

DEMINERALIZATION

ECOLOGICAL PLAQUE HYPOTHESIS AND

PREVENTION OF PERIODONTAL DISEASE

Plaque
Reduction

Plaque
Accumuli

Fig. 1The ecological plaque hypothesis and the etiology

of caries. The diagram depicts a dynamic relationship
whereby an environmental change in plaque (low pH)
produces a shift in the balance of the resident microflora,
thereby favoring demoralization. Caries could be
prevented not only by inhibiting the putative pathogens but
also by interfering with the environmental change driving
the ecological shift. MS = mutans streptococci.

Fig. 2The ecological plaque hypothesis and the

1
etiology ofperiodontal diseases. The diagram depicts the
dynamic relationship whereby the inflammatory response
results in an environmental change, subgingivally, which
produces a shift in the balance of the resident microflora. Such
a shift predisposes a site to disease. Disease could be prevented
not only by inhibiting theperiodontopathogens directly but
also by interfering with thefactors driving the transition.

degradation of host molecules in GCF. Similarly, it was

reported recently that some species that predominate in
periodontitis, but which are not detectable in the healthy
gingiva, can be found as a small proportion of the microflora
in gingivitis (Moore et al., 1987). This also suggests that
environmental conditions which develop during gingivitis
{e.g., bleeding, increased GCF flow) may favor the growth of
species implicated in periodontitis.
The possible effect of GCF on the stability of the subgingival
microflora has been studied in the laboratory by repeated
passaging of plaque through human serum (used as a substitute
for GCF) (ter Steeg et al.,1981) or by the prolonged continuous
culture of plaque on serum (ter Steeg et al., 1988). Both
experimental approaches resulted in the enrichment of species
implicated in periodontal disease (e.g., anaerobic streptococci,

organisms that constitute the resident plaque microflora, only

a very limited number are actively involved in causing disease.
Problems can arise with this hypothesis, however, when
attempting to explain those occasions when either disease is
diagnosed in the apparent absence of the putative pathogens, or
when pathogens are present at sites with no evidence of
disease. When sensitive detection methods are used, mutans
streptococci can be found quite commonly in plaque, albeit in
low numbers (Bratthall, 1991), as can periodontopathogens
such as F. nucleatum, P. intermedia, and, on occasion, A.
actinomycetemcomitans (Ashley et al., 1988; Frisken et al.,
1987; Zimmer et al., 1991). In contrast, the "non-specific
plaque hypothesis" purports that many of the heterogeneous
mixture of organisms in plaque could play a role in disease, and
that disease is a result of the overall interaction of the plaque
microflora with the host (Theilade, 1986). Some of the
arguments surrounding these hypotheses may be, in part, about
semantics (e.g., the definition of "specific" or "non-specific"),
since plaque-mediated diseases, while not necessarily having
a totally specific etiology, do show evidence of specificity.
Consequently, a modified hypothesis (the "ecological plaque
hypothesis") was proposed recently (Marsh, 1991) in an attempt
to unify some of these clinical and laboratory observations. In
this hypothesis, it is proposed that a change in a key
environmental factor (or factors) will trigger a shift in the
balance of the resident plaque microflora, and this might
predispose a site to disease (Figs. 1 and 2). The occurrence of
potentially pathogenic species as minor members of the resident
plaque microflora would be consistent with this proposal.
Under the conditions that prevail in health, these organisms
would be only weakly competitive and may also be suppressed
by inter-microbial antagonism, so that they would comprise
only a small percentage of the plaque microflora and would not
be significant clinically. Microbial specificity in disease would
be due to the fact that only certain species are competitive

Prevotella intermedia, Fusobacterium nucleatum, and

Treponema denticola) that were present in the inoculum at
levels too low to be detected. Likewise, the effect of the
clinically-observed rise in pH during inflammation on the
proportions of three black-pigmented anaerobes has been
studied in mixed-continuous culture. At or below pH 7.0, the
culture was dominated by Prevotella melaninogenica. As the
pH was increased to pH 7.25, P. intermedia became
predominant, whereas at pH 7.5 and above, the culture was
comprised of >99% Porphyromonas gingivalis (McDermid et
al., 1990). These studies have demonstrated the significant
influence an altered supply of nutrients and even a small
change in local pH can have in determining the balance of the
microflora.
IMPLICATIONS FOR THE ETIOLOGY
OF CARIES AND PERIODONTAL DISEASES

There are two main schools of thought on the role of plaque

bacteria in disease. The "specific plaque hypothesis" (Loesche,
1976) proposes that, out of the diverse collection of micro-

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VOL.

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MICROBIAL ECOLOGY OF DENTAL PLAQUE

under the new (changed) environmental conditions. It is a basic

tenet of microbial ecology that a major change to an ecosystem
produces a corresponding disturbance to the stability of the
resident microbial community (Brock, 1966; Alexander, 1971;
Fletcher et aL, 1987). Examples of this have been reported in
ecosystems as diverse as soil (Heal and Ineson, 1984), skin
(Noble and Pitcher, 1978), and water (Pomeroy, 1984).
In subjects with a conventional low-sugar diet, the
composition of the plaque microflora would be stable, and only
small amounts of acid would be produced at main-meal times.
In these circumstances, the processes of de- andremineralization
would be in equilibrium. If the frequency of fermentable
dietary carbohydrate intake were to increase, however, plaque
would spend more time at low pH (Loesche, 1986). Such
conditions would favor the proliferation of mutans streptococci
and lactobacilli at the expense of more acid-tolerant species,
and this would tip the equilibrium toward demineralization
(Fig. 1). Factors reducing the flow of saliva (e.g., xerostomia)
would lead to similar shifts in the microflora. Greater numbers
of mutans streptococci and lactobacilli would lead to even
faster rates of acid production from sugars, enhancing
demineralization still further, while the elevated levels of lactic
acid in plaque would also select for Veillonella spp., as has
been reported in the aforementioned clinical studies. Acidsensitive species, such as members of the S. oralis group (e.g.,
S. sanguiSy S. oralis, and S. mitis), would decline in proportion,
thereby accounting for the widely reported inverse relationship
between S. sanguis and mutans streptococci seen in plaque.
Other bacteria could also produce significant amounts of acid
under similar conditions, at slower rates (van Houte, 1992), but
nevertheless providing an explanation for demineralization in
the absence of mutans streptococci.
Likewise, in periodontal diseases, the changes in nutrient
profile of the gingival crevice due to GCF secretion will lead
to increased plaque biomass, the metabolism of which will
lower the redox potential of the site and raise the pH. These
changes will tend to enrich the previously low levels of
obligately anaerobic and often asaccharolytic Gram-negative
bacteria, and so fuel the inflammatory response (Fig. 2). Other
predisposing factors might include the state of the host defenses,
so that leukotoxin-producing
strains
of
A.
actinomycetemcomitans might exploit the pocket environment
in individuals with neutrophil deficiencies and gain a
competitive advantage (Genco and Slots, 1984). These
sequences of events can go some way toward explaining the
lack of total specificity in the microbial etiology of dental
caries and periodontal diseases, and account for the pattern of
bacterial succession often seen during disease progression in
clinical studies.

IMPLICATIONS FOR DISEASE PREVENTION

Implicit in the ecological plaque hypothesis is the possibility
that disease might be prevented, not only by inhibiting the
putative pathogen(s), but also by interfering with the factors
responsible for the transition of the plaque microflora from
having a commensal to a pathogenic relationship with the host
(Marsh, 1991). A consideration of the principles behind the
ecological plaque hypothesis can lead to the identification of

267

new strategies to prevent disease, as well as enabling existing

approaches to be seen from a fresh perspective (Table 3). These
strategies will now be discussed in terms of their ecological
implications.
Strategies to prevent dental caries

One of the primary mechanisms by which microbial homeostasis

in the mouth can be disrupted is by the repeated exposure of
plaque to low pH following the frequent intake of fermentable
dietary carbohydrates (Loesche, 1986). Consistent with the
prevention of disease under the ecological plaque hypothesis
would be the reduced frequency as well as the depth of such
acid challenges. This could be achieved by (a) inhibitors of
acid production, (b) simple avoidance, between main meals, of
food or drinks containing fermentable sugars, (c) the
consumption of items that contain alternative sweeteners that
are only weakly metabolized, if at all, by oral bacteria, or (d)
stimulation of saliva flow after main meals. Some of these
strategies will now be considered in more detail.
Fluoride

The principal mode of action of fluoride is to increase the

resistance of enamel to demineralization and to promote
remineralization. Fluoride can also inhibit bacterial growth,
but usually at concentrations much higher than those found in
dental plaque (Hamilton and Bowden, 1988). The anti-caries
properties of fluoride, therefore, have not generally been
considered to involve its antimicrobial activity. At sub-MIC
levels, however, fluoride can reduce glycolysis, while its
antibacterial (Bradshaw et al., 1990) and anti-metabolic
(Hamilton and Bowden, 1988) properties are enhanced
markedly at low pH. Concentrations of fluoride that might be
without effect at neutral pH could be inhibitory during acid
production and thereby reduce the deleterious shifts in the
plaque microflora by slowing the rate of change in pH.
For this proposal to be tested, sub-MIC levels (1 mmol/L)
of sodium fluoride were pulsed along with glucose into the
nine-member mixed-culture system described earlier. This
low level of fluoride prevented the selection of S. mutans under
otherwise favorable growth conditions (Table 1). The cultures
were still dominated by L casei and V. dispar after 10 pulses,
but the rate of acid production was reduced by fluoride, and
acid-sensitive species persisted at higher levels than in the
absence of fluoride (Bradshaw etai, 1990). Thus, fluoride was
able to stabilize the composition of the microflora, at least
partially, by reducing the pressure exerted by a rapid fall in
environmental pH, confirming an earlier suggestion by
Hamilton and Bowden (1982).
Antimicrobial agents

Antimicrobial agents, such as chlorhexidine, can be used as an

adjunct to mechanical cleaning for plaque control (Addy,
1986). Such agents are generally selected on the basis of their
spectrum of inhibitory activity, and on their bactericidal or
bacteriostatic mode of action (Lang and Brecx, 1986). However,
agents delivered from dental products have a relatively short
half-life in the mouth and may be present for considerable
periods at sub-MIC levels. At such concentrations, they might

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268

ADV DENT RES JULY 1994

MARSH

still serve a valuable function by interfering with bacterial

metabolism, e.g., by inhibiting acid production (Scheie, 1989;
Cummins, 1991;Marsh, 1992). This mode of action would be
consistent with the ecological plaque hypothesis by again
decreasing the impact of rapid changes in pH on both the
stability of the microflora and on demineralization.
Sugar substitutes and stimulation of saliva flow

Saliva is significant in a number of respects in terms of

caries prevention. Its flow is important in the clearance of
fermentable sugars from the mouth, while also providing
buffering capacity to restore the pH of plaque to resting
values. Furthermore, saliva contains antimicrobial factors,
as well as urea and peptides from which base can be generated
to raise the local pH; saliva can also remineralize enamel.
Two approaches have been adopted to exploit these beneficial
properties of saliva. First, the acid challenge to enamel,
especially during between-meal periods, can be reduced by
the consumption of snack foods, drinks, and confectionery
that contain non-metabolizable sweeteners (sugar substitutes).
The sweetness of these agents stimulates saliva flow in the
absence of significant acid production, and this can promote
remineralization of enamel. Some sugar substitutessuch
as aspartame, saccharin, and xylitolare also able to inhibit
bacterial growth (Grenby and Saldanha, 1986), with saccharin
and xylitol being particularly effective against mutans
streptococci (Best and Brown, 1987;Makinen, 1989; Scheie,
1989). Second, saliva flow can be stimulated for extended
periods after a mealfor example, by the chewing of gum
sweetened with sorbitol (Jensen, 1986; Jensen and Wefel,
1989). It has been proposed that the regular use of sorbitol
chewing gum is not only non-cariogenic but also therapeutic,
in that the stimulation of saliva flow can promote
remineralization (Leach et al., 1989). Xylitol-containing
chewing gum can be used in a similar manner; indeed, in
general, the regular consumption of xylitol-containing gum
gives a greater reduction in caries than similar sorbitolcontaining products (Makinen, 1989). Thus, both approaches
reduce the anti-homeostatic effect of low pH on the balance
of the plaque microflora.

Strategies to prevent periodontal diseases

Most conventional methods of treating disease involve
mechanical removal of subgingival plaque and, sometimes,
the use of antimicrobial agents, especially in advanced or
refractory periodontal disease. Despite less being known about
specific factors that result in changes in the subgingival
microflora, an alternative (ecological) approach would be to
alter the environment of the pocket to prevent the growth of the
putative pathogens.
Anti-inflammatory and antimicrobial agents

Anti-inflammatory agents might break the cycle of tissue

destruction caused by both bacterial and host-derived proteases
(Johnson and Curtis, 1994). This would also reduce the supply
of GCF, and thereby restrict the availability of nutrients
essential for the growth of some periodontopathogens.
Some of the antimicrobial agents being used in dental

health products contain broad-spectrum antimicrobial agents

which can reduce plaque, especially at sites that are difficult
to clean. Care has to be taken with the regular, unsupervised
use of such agents, so that the natural ecology of dental
plaque will not be disrupted (Page, 1989). However, recent
studies have suggested that, at their concentrations in the
mouth, the activity of some of these agents may be more
selective than hitherto suspected. For example, in the same
way that chlorhexidine can be used for selective suppression
of mutans streptococci in plaque (Kohler et al., 1984), so
Triclosan and zinc citrate, in combination, have been found
to have greatest activity against periodontopathogens in vitro
(Bradshaw etal, 1993) and in vivo (Jones etal, 1990), while
leaving streptococci associated with sound enamel and a
healthy periodontium relatively unaffected (Bradshaw et al.,
1993). Again, at sub-MIC levels, several antimicrobial agents
may have additional potentially valuable properties by inhibiting
bacterial proteases implicated in tissue destruction (Scheie,
1989; Cummins, 1991; Marsh, 1992).
Oxygenating and redox agents

Another approach has been to try to raise the redox potential of

the pocket (which is lowered during disease) to create an
environment incompatible with the growth of obligate
anaerobes. This has been tried with molecular oxygen or an
oxygenating agent (Chasens, 1978), with various rates of
success. Recently, the use of redox dyes has been proposed
which, while not releasing oxygen, can raise the redox potential
of an ecosystem (Wilson et al., 1992). Methylene blue was
applied subgingivally on a daily basis for 7 days at 25 test sites;
control sites in the same patients received water. Treatment led
to a significant reduction in flow of GCF, and reduced the
proportions of obligate anaerobes and motile organisms in the
subgingival microflora; this was accompanied by a concomitant
increase in facultatively anaerobic and coccal bacteria (Wilson
et al., 1992). In vitro studies also demonstrated that methylene
blue could raise the Eh of pre-reduced culture medium, and
significantly reduce the viable counts of a suspension of P.
gingivalis (Fletcher and Wilson, 1993). These early studies
confirm the theoretical basis of the ecological plaque hypothesis
by showing that a preventive strategy that interferes with a
critical event in the breakdown of microbial homeostasis in
plaque can shift the ecological balance of plaque back toward
that which is compatible with dental health.

Strategies to enhance colonization resistance

The phenomenon by which one member of an ecosystem can
inhibit the growth of another member is termed "bacterial
interference". The possibility that antagonistic organisms could
be used to control pathogens and prevent disease has been
proposed for over 100 years and is termed "replacement
therapy". This approach has the potential advantage that it
provides life-long protection with minimal cost or compliance
on behalf of the recipient, once colonization by the "effector"
strain has been achieved (Hillman and Socransky, 1989).
There are two main approaches by which replacement therapy
is being considered as a means of enhancing colonization
resistance in plaque to prevent caries and periodontal diseases.

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MICROBIAL ECOLOGY OF DENTAL PLAQUE

Pre-emptive colonization

In this approach, ecological niches (functions) within plaque

are filled by a harmless or potentially beneficial organism
before the undesirable strain has had an opportunity to colonize
or become established. The initial colonizer becomes integrated
into the ecosystem and subsequently excludes the pathogen
(Donoghue, 1990). Low-virulence mutants of mutans
streptococci have been produced that are deficient in
glucosyltransferase, intracellular polysaccharide, or lactate
dehydrogenase (LDH) activity, and which are designed to
prevent subsequent colonization by "natural" mutans
streptococci. However, wild-type revertants can occur, the
degree of colonization by the mutant can vary with the animal
host, and mutants do not always compete successfully in vivo.
Competitive displacement

An alternative approach has been to derive a more competitive

strain that would displace a pre-existing organism from plaque
(Donoghue, 1990). Competitive displacement is of potentially
greater clinical value, since it is not dependent on treatment
with the "effector" strain at or before colonization by the
undesired organism (Hillman and Socransky, 1989). An unusual
strain of S. salivarius (TOVE-R) was shown to displace S.
mutans from the teeth of rats and to inhibit tooth decay (Tanzer,
1989), but this organism was less effective when attempts were
made to implant it into human plaque (Fisher and Tanzer,
1984). A strain of S. mutans, selected on the basis of enhanced
bacteriocin production, was able to colonize the teeth of human
volunteers persistently, and it reduced the indigenous strain of
S. mutans in some subjects (for a review, see Hillman and
Socransky, 1989). The properties of a number of potential
"effector" strains for replacement therapy have been reviewed
(Tanzer, 1989).
Competitive displacement has also been considered in the
treatment of LJP. As stated earlier, plaque from periodontallyhealthy sites contained organisms, such as H2O2-producing
strains of S. sanguis, that inhibited the growth of A.
actinomycetemcomitans, whereas the converse was true with
plaque taken from sites with LJP (Hillman and Socransky,
1982). Levels of A. actinomycetemcomitans were reduced
markedly in gnotobiotic rats when the animals were
superinfected by wild-type S. sanguis, but variable findings
have been reported when attempts have been made to implant
S. sanguis in humans (Hillman and Socransky, 1989). Thus, at
present, conventional approaches using debridement and
antibiotics remain the optimal form of treatment.
Before replacement therapy can be considered as a
practical alternative to existing treatment, the problems of
implanting effective "effector" strains will have to be
overcome, and assurances of the safety of these strains will
be required. Molecular biology techniques are being exploited
to develop suitable "effector" strains with the desired
properties. Nevertheless, the use of bacterial interference
to produce plaque either with a lower disease potential or
with an increased level of colonization resistance would be
consistent with the principles of the ecological plaque
hypothesis.

ACKNOWLEDGMENTS
The assistance of Dr. DJ. Bradshaw with the Figs, and of Dr.
A.H. Rogers with the section on replacement therapy is
gratefully acknowledged.
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