MECHANISMS
OF DENTAL PLAQUE FORMATION
T
A.AA. SCHEIE
Department of Oral Biology
Dental Faculty, University of Oslo
Pb 1052, Blindern
N-0316 Oslo, Norway
Adv Dent Res 8(2):246-253, July, 1994
Abstract—Much effort has been placed on elucidating the
diverse mechanisms of microbial adhesion to tooth surfaces.
Both specific and non-specific types of adhesion have been
envisaged. Pioneer colonizers represent a selected part of the
oral microflora, and it has been assumed that specific adhesin-
receptor interactions between the microbial surface and the
pellicle account for this specificity. Whereas microbial adhesion
to tooth surfaces is a general prerequisite for initiation of
plaque formation, microbial multiplication is probably the
dominant feature in the build-up of dental plaque. Local
environmental factors which influence the establishment and
composition of the ultimate plaque community are therefore of
greater importance than initial adhesion per se. The highly
individual and site-related characteristics of the plaque flora
illustrate the selective power of the environment. Environmental
conditions are not uniform. Thus, each site represents its own
distinct ecosystem, and the microbial composition at the site
depends on the outcome of a variety of host-microbial and
microbial-microbial interactions. The relative in vivo
significance of these interactions is difficult to assess.
This manuscript was presented at a Symposium entitled
"Mechanisms and Agents in Preventive Dentistry", held
October 28-November 1, 1992, in Chester, England, under
the auspices of the Council of Europe Research Group on
Surface and Colloid Phenomena.
246
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he adhesive potential of micro-organisms is considered
to be an important ecologic and pathogenic determinant
in the development of infectious diseases. In the same
way, microbial adhesion to tooth surfaces has been
considered an important oral ecologic factor (van Houte et al.,
1970, 1971; Gibbons and van Houte, 1975). Much effort has
been placed on elucidating the diverse mechanisms of microbial
adhesion to tooth surfaces. The obvious reason for this particular
interest is the anticipated role of dental plaque in the development
of dental caries and periodontal disease, and the possibility for
prophylactic interventions through interference with microbial
adhesion.
The present paper is not intended to be a comprehensive
review of all aspects of microbial adhesion to tooth surfaces,
but rather a discussion of the clinical implications of our
present knowledge obtained from the numerous in vitro model
studies.
MICROBIAL COMPOSITION
OF THE EARLY PLAQUE FLORA
There is general agreement that the early plaque flora constitutes
a selected part of the oral microflora, namely, the oral
streptococci and, in particular, Streptococcus sanguis (Gibbons
andvanHoute, 1975;Socransky^a/., 1977; Syed and Loesche,
1978; Theilade et al., 1982; Nyvad and Kilian, 1987;
Macpherson etal., 1991). However, due to the recent changes
in taxonomy of the oral streptococci (Schmidhuber etal., 1987;
Kilian et al., 1989), variations in the nomenclature make
comparisons of new and old data on the numbers of the various
streptococcal species difficult. By applying the revised
taxonomy, Nyvad and Kilian (1987, 1990) and Macpherson
and co-workers (1991) provide new insight. They performed
quantitative and qualitative studies of the early plaque flora in
vivo. Several enamel and/or root surface slabs were mounted
in appliances, allowing for comparisons of undisturbed plaque
developed for various time periods under identical conditions.
Both studies confirm the dominance of streptococci in early
plaque. This is the case for both enamel and root surfaces
(Nyvad and Kilian, 1987), in caries-active as well as in caries-
inactive individuals (Nyvad and Kilian, 1990), and for enamel
surfaces regardless of the presence or absence of sucrose
(Macpherson etal., 1991).
The studies revealed that the early colonizing streptococci
belong mainly to three species: Streptococcus mitis (Nyvad
and Kilian, 1987, 1990); Streptococcus oralis, the previous S.
sanguis 11 and Streptococcus mitior; and S. sanguis (Nyvad
and Kilian, 1987, 1990; Macpherson etal., 1991).
The high levels of streptococci encountered in early plaque
may be taken to support specificity of microbial adhesion.
However, both the number of micro-organisms available for
adhesion (van Houte and Green, 1974) and their inherent
VOL. 8(2)
PLAQUE FORMATION BY ADHESION AND MICROBIAL GROWTH
Mode Rate
Adhesion 1% perh
Growth 2 h gen. time
0rstavikD, 1984.
ability to adhere determine the probability for adherence
(Gibbons and van Houte, 1975). Thus, caries-active individuals
who show higher salivary levels of micro-organisms may also
show a higher plaque formation rate (Ny vad and Kilian, 1990).
The high levels of streptococci in early plaque may therefore,
on the one hand, reflect the fact that they comprise a major
portion of the salivary flora. Streptococcus salivarius, on the
other hand, is the most prevalent salivary streptococcal species,
but constitutes only a minor portion of the plaque flora, thus
supporting the selectivity involved in early plaque formation.
Actinomyces spp. are the most prevalent among Gram-
positive rods, but constitute less than 10% of total viable
counts. Other Gram-positive rods encountered are
Propionibacteria, Corynebacteria, and Rothia (Nyvad and
Kilian, 1987). Interestingly, both the two previously mentioned
studies indicate variable and decreasing proportions of Gram-
positive rods during early phases of plaque formation. A lag
time for replication (Macpherson et a/., 1991) or a relatively
slow growth rate (Nyvad and Kilian, 1987; Nyvad and
Fejerskov, 1987b) of the Actinomyces spp., was suggested to
account for this phenomenon.
Ultrastructural studies by both scanning and transmission
electron microscopy confirm the cultural findings (Nyvad and
Fejerskov, 1987a,b). However, whether the seemingly
qualitative selectivity relates to specificity of the initial
adsorption process or to varying ability to metabolize and grow
on the tooth surface at initial stages is not ascertained. It was
shown by use of a vital fluorescence technique that the relative
number of viable micro-organisms was lower in the early
(four-hour) than in later (24-hour) phases of plaque formation,
probably due to a high sensitivity to and accessibility for
salivary antimicrobial factors against early colonizers (Weiger
et al., 1993). This latter observation supports the view that,
besides microbial adhesive properties, local environmental
factors have decisive impact on early plaque formation.
MICROBIAL ATTACHMENT TO TOOTH SURFACES
General Considerations
Classic studies have outlined two stages in microbial adhesion:
the reversible sorption, which is an instantaneous attraction of
micro-organisms to a surface, followed by irreversible sorption
which involves biologically specific reactions providing firm
adhesion of micro-organisms to the surface (Marshall et aL,
1971). In a recent review, vanLoosdrechtefa/. (1990) illustrated
schematically microbial adhesion to surfaces in aquatic
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PLAQUE FORMATION 247
TABLE 1
Time to Reach 10n Cells per mm2
4
10 105 106 107 108
lh 10 h 4d 40 d lyr
lh 6h 12 h 24 h 36 h
environments as a four-stage sequence. This also may serve as
a schematic representation of the dynamic plaque formation
process: The first stage involves the initial transport of a
bacterium to the tooth surface. Random contact may occur, for
instance, through Brownian motions, through sedimentation
of micro-organisms, through liquid flow, or through active
movement of the micro-organisms.
The second stage is the initial, reversible adsorption which
results in the secondary minimum. These interactions are too
weak to provide firm attachment. They result from interplay
between attractive van der Waals' forces and repulsive
electrostatic interactions. These interactions are influenced by
pH and ionic strength of the suspending medium and are
determined by macroscopic, physico-chemical characteristics
of the interacting surfaces, i.e., the surface charge and surface
energy or hydrophobicity of the pellicle-covered tooth surface
and adhering micro-organisms.
Once these forces have been overcome, it is thought that
biologically specific microscopic characteristics of the pellicle-
covered tooth and microbial surfaces become determinant for
the following firm attachment, which constitutes the third
stage. The firm attachment is followed by surface colonization
and biofilm formation, which eventually reaches the "climax
community" of dental plaque.
Experimentally, the four stages may be separated, and a
number of model systems has been used to characterize the
mechanisms involved in the separate events, with the assumption
that they function similarly in vivo. In particular, the non-
specific, macroscopic surface characteristics of stage two and
the specific microscopic adhesin-receptor interactions involved
in stage three have been studied and provided important
knowledge. Nevertheless, data from such studies have not yet
led to a complete understanding of the processes as they occur
in vivo.
Unfortunately, most studies have been performed in vitro,
often under conditions which are irrelevant for a clinical
situation. It is extremely difficult to create in vitro systems
which simulate oral conditions. Minor alterations in the
composition, the pH, or the ionic strength of the suspending
medium, as occurs, for instance, in the presence of saliva, may
alter the adhesive properties in the system. Moreover, the role
of saliva has often been neglected. An important point is that
all surfaces in the oral cavity, including the micro-organisms,
adsorb salivary proteins, forming a protein film which may
alter surface properties and mask surface structures. Therefore,
the inherent effects of the surface characteristics of the naked
248
tooth or micro-organism are probably limited. Saliva also
provides available calcium ions which may bridge opposing
negatively charged tooth and microbial surfaces, thus providing
non-specific binding (R0lla, 1976). Furthermore, saliva contains
components with the ability both to promote and to inhibit
microbial adhesion (Mandel, 1989).
Another point is that by employing laboratory strains,
which is most often the case in model studies, one runs the risk
of studying micro-organisms which have lost or modified
surface components involved in adhesion, such as surface
appendages and hydrophobicity. Also, caution should be used
in generalizing such data, since many reactions are strain-
related rather than species-related. Thus, adhesive events in in
vitro model systems may well proceed through routes which
have no or little relevance to the in vivo situation.
Model systems are "clean systems" which allow for the
study of isolated factors, but they cannot mimic, in any way, the
individually varying physiological conditions at the various
sites within the dentition. Thus, the mechanisms delineated in
simplified systems represent only fractions of the numerous
interacting and competing factors operating among the entire
flora. Therefore, the relative in vivo significance of the various
factors cannot be assessed in such studies.
Most model studies fail to account for the considerable
contribution to the plaque mass of microbial proliferation
(Brecx et al, 1983) as well as the various environmental
factors which influence early stages of plaque formation.
In view of the complex environment within the oral cavity,
the true situation can be studied only in vivo. Clinical studies
show that micro-organisms colonize as individual organisms.
The increase in number from 12-24 h is accounted for mainly
by cell division (Ny vad and Fejerskov, 1987b). This is supported
by the calculations made by 0rstavik (1984, Table 1). According
to his assumptions, that 1% of the micro-organisms adhere per
hour and that the generation time is 2 h, the time to reach 106
micro-organisms would be 4 days by adhesion, but only 12 h
by microbial growth. The importance of growth may be
overestimated and the role of adhesion may have been
underestimated in these calculations, since the generation time
in plaque is generally longer than 2 h and only a few of the
micro-organisms in dental plaque are actually dividing
(Critchley and Bowen, 1970). Moreover, the number of micro-
organisms adhering may be higher than 1 %. Notably, the inter-
individual differences in plaque formation become evident
during the proliferative phase from 12-24 h (Nyvad and
Fejerskov, 1987a), thus supporting the importance of microbial
growth for dental plaque formation and for its diversity in
composition.
Whereas microbial adhesion is a prerequisite for initiation
of plaque formation in vivo, microbial proliferation is probably
the dominant feature in the build-up of plaque. Studies of
plaque formation mechanisms should therefore include the
study of factors involved in the stage from firm attachment to
the established biofilm, i.e., factors which affect microbial
proliferation and intermicrobial binding.
In the following, comments will be made on recent in vivo
and in vitro findings, with particular reference to their clinical
significance.
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SCHEIE ADV DENT RES JULY 1994
Role of Macroscopic Surface Characteristics
Macroscopic cell surface characteristics relevant for microbial
adhesion include surface energy, zeta potential, and
hydrophobicity, which are interrelated phenomena.
Theoretically, modification of the chemical characteristics of
the tooth surface would seem to be an attractive non-
antimicrobial way of controlling plaque formation.
Surface Energy
Critical surface tension (Glantz, 1969; Christersson et al,
1989) or interfacial surface free-energy (Busscher and
Weerkamp, 1987) has been postulated as a driving force for
initial adhesion of micro-organisms to solid surfaces.
Surface energy determinations are based on in vitro contact
angle measurements of the substratum (Glantz, 1969;
Christersson et al, 1989) or of the substratum and the micro-
organisms (Busscher and Weerkamp, 1987). It is shown in
vitro that adsorption of micro-organisms depends on critical
surface tension of the substratum and the flow rate of saliva.
Very hydrophobic and very hydrophilic surfaces retain few
micro-organisms (Christersson et al, 1989), whereas surfaces
of medium critical surface tension (35-38 mN/m) retain the
highest number of micro-organisms from human whole saliva.
Adsorption of Streptococcus and Actinomyces spp. is similar
in this system, yet streptococci dominate in early plaque in vivo
(Nyvad and Kilian, 1987, 1990; Macpherson et al, 1991).
By using hydroxyapatite made hydrophilic by various agents,
Olsson and co-workers (Olsson et al, 1990) showed, in vitro,
that adhesion of a hydrophobic S. mutans strain was reduced.
However, when the same agents were tested clinically, only
minor, statistically non-significant effects on plaque formation
were obtained (Olsson et al, 1990).
According to the model of interfacial surface free-energy,
the change in free energy as materials form interfaces is
important. Adhesion is favored if it is accompanied by a
decrease in free energy of the system, i. e., interfacial free energy
of micro-organisms, substratum, and suspending medium.
Accordingly, low surface free-energy micro-organisms adhere
to low surface free-energy substrata, and high surface free-
energy micro-organisms to high surface free-energy substrata
(Busscher and Weerkamp, 1987). In vitro measurements show
large differences in the surface free-energy of individual
microbial isolates (37.8-122.6 mJnr2, Weerkamp et al, 1989).
Most oral micro-organisms have high surface free-energy,
which theoretically implies that they adhere preferentially to
surfaces with high surface free-energy, such as dental enamel
(88 mJm2; Busscher and Weerkamp, 1987; Weerkamp et al,
1989). Theoretically, lowering the surface free-energy of teeth
would thus reduce colonization. Support for this assumption
has been found in in vivo comparative studies of plaque
formation on enamel (88 mJnr2), cellulose-acetate (58 mJnr2),
and Teflon® (20 mJnr2) (Quirynen et al, 1989; Weerkamp et
al, 1989). Significantly fewer micro-organisms colonized the
Teflon and cellulose-acetate surfaces than enamel, which has a
higher surface free-energy than the two synthetic test surfaces.
Also, low surface free-energy micro-organisms were found to
colonize the low energy surfaces preferentially (Quirynen etal,
VOL. 8(2)
1989; Weerkamp et al., 1989). Mean surface free-energy of
micro-organisms adhering to Teflon, cellulose acetate, and
enamel were 84 mJm2, 86 mJm2, and 93 mJm2, respectively
(Weerkamp et al., 1989). Moreover, the adhesion to the low
surface free-energy surfaces seemed weak (Quirynen et al.,
1989).
Interestingly, in a recent clinical study, the influence of
surface roughness was found to be more important for plaque
formation in vivo than surface free-energy per se. The rough
surfaces (13-15 |im) were more heavily colonized, and the
plaque was at a more mature stage after six days than plaque on
a comparable smooth surface (0.75 jam, Quirynen etal., 1990).
This corroborates the clinical findings by Nyvad and Kilian
(1987) and Nyvad and Fejerskov (1987b) that root surfaces are
more heavily colonized than enamel surfaces, and underlines
the modulating potential of the multitude of factors operating
in vivo.
Zeta Potential
The zeta potential of an organism has also been described as
being an important surface characteristic in adhesion (Olsson
et al., 1976). The zeta potential is determined by the nature and
number of ionogenic groups on the cell surface and depends on
the pH and ionic strength of the suspending medium. A low
negative charge expectedly favors adhesion. According to this
concept, the higher cariogenicity of Streptococcus sobrinus
over other mutans streptococci (de Soet et al., 1991) was
ascribed to a comparatively smaller negative surface charge
(van der Mei et al., 1991). However, adsorption to saliva-
coated glass surfaces was found to be the same for all tested
species, including S. sobrinus, regardless of both surface free-
energy and surface charge (Busscher et al., 1992).
Hydrophobicity
Although microbial hydrophobicity is ill-defined (van der
Mei, 1989), hydrophobicity of micro-organisms has been
implicated as being important for adherence. Various methods
have been used to measure microbial hydrophobicity, which
varies widely among various species and strains. Hydrophobic
micro-organisms are attracted to solid surfaces by rejection
from the aqueous phase. It is suggested that ionic, ion-dipole,
or hydrogen bond interactions may be stabilized by hydrophobic
bonds (Nesbitt etal., 1982). Fimbriae which may bridge gaps
between micro-organisms and tooth surfaces have been
associated with hydrophobicity. It has also been suggested that
lipoteichoic acid may confer hydrophobicity to the microbial
surface.
Hydrophobic streptococci adhere better to hydroxyapatite
in vitro than do less hydrophobic strains. Along this line,
hydrophobic strains of S. mutans persist to a higher degree than
less hydrophobic strains upon in vivo implantation in humans
(Svanberg etal., 1984). In contrast, however, hydrophobic and
hydrophilic strains of 5. mutans were found to implant equally
well in a more recent study in hamsters (Olsson and Emilson,
1988). Notably, induction of streptomycin resistance into the
micro-organism being implanted, as was done in the human
study (Svanberg et al., 1984), may impair colonization
(Bammann et aL, 1978). It also cannot be excluded that
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PLAQUE FORMATION 249
essential surface structures of the micro-organism may be lost
upon repeated subculture, along with the loss of hydrophobicity.
Thus, no definite conclusions can be drawn from these two
latter studies about the clinical significance of surface
hydrophobicity.
ROLE OF MICROSCOPIC SURFACE CHARACTERISTICS
Salivary Components
Several salivary components have been shown to aggregate
micro-organisms. This aggregating ability has been taken to
support a role of certain salivary components in microbial
adhesion to the pellicle-covered tooth surface and to confer
specificity to the adhesive process of the early colonizers
(Ericson and Magnusson, 1976). For instance, salivary
oligosaccharide-containing glycoproteins may serve as
receptors for oral streptococci in the salivary pellicle (Gibbons
and Qureshi, 1978), and the salivary proline-rich protein 1 and
statherin have been implicated as receptors for type 1 fimbriae
of A. viscosus (Gibbons etal., 1988).
It seems paradoxical, however, that the same salivary
components which serve as receptors for microbial adhesion
when parts of the pellicle would inhibit microbial adhesion
by binding to microbial adhesins when they are present in
solution in saliva. Gibbons and co-workers (Gibbons 1989;
Gibbons et al., 1990) recently pointed to the conformational
changes which may occur in proteins upon adsorption to
surfaces. They showed in vitro, for instance, ihatActinomyces
spp., which bind to proline-rich proteins in the adsorbed
state, fail to interact with the same proteins when present in
solution (Gibbons etal., 1990). The explanation may be that,
upon adsorption, previously hidden molecular segments which
may serve as receptors are being exposed. Such "cryptitopes"
may be revealed by the adsorption of the molecule to the
tooth surface, or by the enzymatic cleavage of terminal
residues. For instance, neuraminidase removes terminal sialic
acid residues, resulting in exposure of galactosyl. This may,
on the one hand, reduce the adhesion of micro-organisms
which bind to neuraminidase-sensitive receptors such as S.
sanguisandS. mitis strains (Liljemarker al., 1989),but may,
on the other hand, increase the adhesion of other successor
micro-organisms (Gibbons, 1989). Proteases, for instance,
may cleave peptide bonds, thus exposing arginine residues,
making them available for binding. The presence of proteases
in crevicular fluid of periodontitis patients and elevated
levels of neuraminidase in oral fluids in patients with poor
oral hygiene are taken to support a possible role in vivo (for
review, see Gibbons, 1989). Such mechanisms may also
explain the rapid establishment of a complex microbial flora
as observed in test subjects with neglected oral hygiene
(Brecx et al., 1980). However, definite evidence for an in
vivo function of cryptitopes is yet to be obtained.
Microbial Adhesins
The prevalence of microbial species on a surface has been
found to correlate with the organism's inherent ability to
adhere to the surface (Gibbons and van Houte, 1975). General
microbiologic research has revealed that most micro-organisms
250
possess a vast number of adhesins which may bind
stereochemically to complementary receptors on the interacting
surface. Adhesins often are associated with filamentous
appendages or fimbria, or with high-molecular-weight proteins
extending from the microbial surface. Electron microscopic
studies of dental plaque reveal that early colonizers of the tooth
surfaces are connected by microbial surface fimbriae (Nyvad
and Fejerskov, 1987b). Fimbriae which can act as adhesins
often possess hydrophobic domains consisting of non-polar
amino acids which confer adhesion.
In vitro studies suggest that the fibrillar, hydrophobic
surface-protein PI is involved in both salivary aggregation
and in adherence of 5. mutans to saliva-coated hydroxyapatite
(Lee et al, 1989; Bowen et al, 1991). It would thus be
expected that a mutant lacking this protein would adhere less
well to teeth. However, the PI-deficient mutant was found to
colonize rat teeth as well as the intact parent strain when the
rats were fed a high-sucrose diet (Bowen et al, 1991),
indicating a diversity of adhesive mechanisms for one microbial
strain. The results again demonstrate the shortcomings of
model systems and the often-seen inconsistencies of in vitro
and in vivo findings, due to the variability and range of
prevailing in vivo environments.
Intermicrobial Co-aggregation
Species- or strain-specific co-aggregation based on cell-to-cell
recognition has been shown to occur among a diversity of
micro-organisms in vitro. The so-called "corn cob" structures
observed microscopically in dental plaque have been taken as
evidence for the importance of such co-aggregations in in vivo
plaque formation. However, in a recent study by Skopek and
co-workers (Skopek et al, 1993), co-aggregation was found to
have only limited effect on in vivo plaque formation. The
results, on the other hand, pointed to the importance of other
environmental factors, such as microbial growth (Skopek et
al, 1993).
INFLUENCE OF ENVIRONMENTAL CONDITIONS
As outlined in the previous sections, in vitro model studies
reveal a vast variety of factors involved in microbial adhesion
to surfaces, but to what degree they are operative within the
complex environment of the oral cavity is uncertain. The
influences of environmental conditions and of microbial-
derived components for microbial adhesion—and in particular
for the subsequent growth of plaque micro-organisms—have
often been overlooked in such model studies. Their importance
is clearly demonstrated in the marked inter-individual
variation in plaque formation rate and the highly individual
characteristics of the structural composition of dental plaque at
the various locations within the dentition (Nyvad and Fejerskov,
1987a, 1989). This underlines the wide variability of host and
microbial factors and their possible multiple interactions,
which may affect both microbial adhesive properties and
microbial growth. First, environmental conditions and microbial
interactions affect the microbial growth rate, which in turn may
affect the microbial surface structures involved in microbial
adhesion (Knox and Wicken, 1985; van der Hoeven et al,
1985, Table 2; van Loosdrecht et al., 1990). Second, such
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SCHEIE ADV DENT RES JULY 1^4
factors may affect subsequent plaque accumulation, as shown
by the well-established microbial shift taking place within
dental plaque with time (Ritz, 1967).
The microbial load on tooth surfaces in caries-active
individuals is found to be higher than in caries-inactive
individuals (Nyvad and Kilian, 1990). Moreover, distinct
differences appear as to the relative proportions of the various
streptococcal species between caries-active and caries-inactive
individuals. Caries-inactive individuals are colonized by higher
numbers of S. sanguis and lower numbers of 5. mitis and S.
mutans than are caries-active individuals. The numbers of S.
oralis, on the other hand, are similar in caries-active and caries-
inactive individuals (Nyvad and Kilian, 1990).
Colonization is also dependent on the surface structure. For
instance, root surfaces are more heavily colonized than enamel
surfaces, and a more advanced stage of development is reached
earlier than on enamel surfaces (Nyvad and Fejerskov, 1987b).
The sequence of colonization and the relative proportions of
the various species are, however, similar on enamel and root
surfaces (Nyvad and Kilian, 1987; Nyvad and Fejerskov
1987a).
The potential selective power of a micro-environment and
the adaptability of the microflora may well be demonstrated by
the emergence of aciduric bacteria such as mutans streptococci
and lactobacilli in a niche created by a loosely fitted orthodontic
band (Arneberg et al, 1984). Within four weeks, the proportion
of mutans streptococci emerged from less than 1 % to more than
10%. Similarly, lactobacilli emerged from undetectable levels
to 1-10%. Another example is the powerful selection of the
microflora in an infected root canal, shown experimentally in
monkeys by Fabricius and co-workers (Fabricius et al, 1982).
When the microbial strains were isolated from the canal,
grown and re-inoculated in equal proportions into 12 canals,
the almost exact microbial distribution as found initially was
re-established (Fabricius et al, 1982). Though plaque on
unprotected surfaces constitutes more open systems than these
two examples, each site represents distinct ecosystems with
characteristic environmental conditions which support the
growth of a characteristic microbial community adapted to the
environment. The gradient of mutans streptococci within the
dentition illustrates this very clearly (Kristoffersson et al,
1984).
Marsh (1991) showed the impact of the pH, the carbohydrate
supply, and the presence of fluoride on maintenance of the
homeostasis of a mixed culture in a chemostat. A carbohydrate
challenge had little effect at neutral pH, but when pH was
allowed to fall following carbohydrate metabolism, S. mutans,
Lactobacillus, and Veillonella dispar increased and became
the dominant species. This environmental selective pressure
could be counteracted by addition of low levels of NaF (1
mmol/L) to the system (Marsh, 1991), showing the ability of
the microbial mass to adapt to changing environmental
conditions.
Role of Sucrose
With the presence of glucosyltransferases in saliva and pellicle
(Rolla et al, 1983; Scheie and Rolla, 1984; Scheie, 1985;
Scheie etal, 1987), sucrose may be an environmental factor of
VOL. 8(2)
MINIMAL INFECTIVE DOSE AND ADHERENCE OF S. mutans T2 IN CONVENTIONAL RATS
Growth Conditions of Minimal Infective Dose
the Inoculum Sucrose
Chemostat lxlO5
Batch 5xlO 5
van der Hoeven et aL, 1985
* Number of adherent cells (CFU) per 106 inoculated cells.
great importance (Rolla et aL, 1985). Due to the universal use
of sucrose, the primary surface for microbial colonization in
vivo may in fact be a glucan-covered tooth surface. In the
presence of sucrose, glucan formation by glucosyltransferase
adsorbed to teeth or to micro-organisms may hide potential
specific adhesins and receptor sites. But, on the other hand,
glucan also may provide receptors for a number of micro-
organisms possessing glucan-binding proteins. Moreover, non-
adhering or loosely adhering micro-organisms may possibly
be entrapped in the glucan meshwork forming in the presence
of sucrose. This may explain why the minimal infective dose
of 5. mutans T2 in conventional rats was found to be significantly
lower in the presence of sucrose than in the presence of glucose
(Table 2), and why chemostat- and batch-grown cells adhere
equally well in the presence of sucrose, whereas adhesion of
batch-grown cells is higher than that of chemostat-grown cells
in the presence of glucose (Table 2). Such phenomena may also
explain why the protein-Pi -deficient S. mutans, which were
unable to adhere to saliva-coated surfaces, were able to adhere
to a glucan-covered surface (Bowen et aL, 1991).
In a recent in vivo study by Steinberg and co-workers
(Steinberg et aL, 1993), some actinomyces strains were shown
to possess glucan-binding proteins, and thus adhesion to glucan-
covered hydroxyapatite was enhanced. For other strains of
actinomyces, adhesion to pre-formed glucan was decreased
compared with adhesion to saliva-coated hydroxyapatite. These
data clearly demonstrate the complex events of microbial
adhesion.
In vivo human studies show that sucrose increases the initial
number of micro-organisms per surface area. This is the case
for Gram-positive cocci and rods, as well as for Gram-negative
cocci and rods, except for S. sanguis and Bacteroides spp. As
to the relative proportions of the various species, exposure to
sucrose has seemingly no effect, except for the S. sanguis and
Bacteroides spp., whose relative proportions thus may be
lowered (Macpherson et aL, 1991).
It was found in another clinical study that sucrose increases
the level of glucosyltransferase in saliva and that sucrose
supply increased plaque formation in these subjects (Scheie,
1985), supporting a possible role in plaque formation of
glucosyltransferase in saliva and pellicle.
In the absence of sucrose, the number of colonizing micro-
organisms increases steadily, whereas colonization seems to
reach a plateau after 30 h during sucrose exposure (Macpherson
etaL, 1991). This may indicate that the capacity of the enamel
sites is reached more quickly (Macpherson et aL, 1991),
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PLAQUE FORMATION 251
TABLE 2
Adherence*
Glucose Sucrose Glucose
5xlO 7 107 7
5xlO 7 129 82
whereby a "climax community" is reached within a shorter
period in the presence of sucrose.
CONCLUSION
Numerous in vitro studies have provided circumstantial
evidence for a diversity of potential adhesive mechanisms for
microbial adhesion to teeth. Though adhesion is essential in the
colonization of the tooth surface, environmental factors are
quantitatively more important for the ultimate composition of
dental plaque. The composition of dental plaque at a given site
depends on the outcome of a variety of competing microbial-
microbial and host-microbial interactions. The relative in vivo
significance of these interactions is difficult to assess due to
lack of appropriate in vivo assays.
REFERENCES
Arneberg P, Ogaard B, Scheie AAa, R0lla G (1984). Selection
of Streptococcus mutans and Lactobacilli in an intra-oral
human caries model. Scand J Dent Res 63:1197-1200.
Bammann LL, Clark WB, Gibbons RJ (1978). Impaired
colonization of gnotobiotic and conventional rats by
streptomycin-resistant strains of Streptococcus mutans.
Infect Immun 22:721-726.
Bowen WH, Schilling K, Giertsen E, Pearson S, Lee SF,
Bleiweis A, et aL (1991). Role of a cell surface-associated
protein in adherence and dental caries. Infect Immun 59:4606-
4609.
Brecx M, Theilade J, Attstrom R (1980). Influence of optimal
and excluded oral hygiene on early formation of dental
plaque on plastic films. J Clin Periodontol 7:361-373.
Brecx M, Theilade J, Attstrom R (1983). An ultrastructural
quantitative study of the significance of microbial
multiplication during early dental plaque growth. JPeriodont
Res 18:177-186.
Busscher HJ, Weerkamp AH (1987). Specific and non-specific
interactions in bacterial adhesion to solid substrata. FEMS
Microbiol Rev 46:165-173.
Busscher HJ, Doornbusch GI, van der Mei HC (1992). Adhesion
of mutans streptococci to glass with and without a salivary
coating as studied in a parallel-plate flow chamber. J Dent
Res 71:491-500.
Christersson CE, Dunford RG, Glantz PO, Baier RE (1989).
Effect of critical surface tension on retention of oral
microorganisms. Scand J Dent Res 97:247-256.
Critchley P, Bowen WH (1970). Correlation of the biochemical
composition of plaque with the diet. In: McHugh WD,
252
Editor. Dental plaque. Dundee: Thompson & Co Ltd., 157-
169.
de Soet JJ, van Loveren C, Lammens AJ, Pavicic MJAMP,
Homburg CHE, ten Cate JM, et al. (1991). Difference in
cariogenicity between fresh isolates of Streptococcus
sobrinus and Streptococcus mutans. Caries Res 25:116-
122.
Ericson T, Magnusson I (1976). Affinity for hydroxyapatite of
salivary substances inducing aggregation of oral
streptococci. Caries Res 10:8-18.
Fabricius L, Dahlen G, Holm SE, Moller AjR (1982). Influence
of combinations of oral bacteria on periapical tissues of
monkeys. Scand J Dent Res 90:200-206.
Gibbons RJ (1989). Bacterial adhesion to oral tissues: a model
for infectious diseases. J Dent Res 68:750-760.
Gibbons RJ, Qureshi JV (1978). Selective binding of blood
group-reactive salivary mucins by Streptococcus mutans
and other oral organisms. Infect Immun 22:665-671.
Gibbons RJ, van Houte J (1975). Bacterial adherence in oral
microbial ecology. Ann Rev Microbiol 29:19-44.
Gibbons RJ, Hay DI, Cisar JO, Clark WB (1988). Adsorbed
salivary proline-rich protein 1 and statherin: receptors for
type 1 fimbriaeof Actinomyces viscosus T14V-11 on apatite
surfaces. Infect Immun 56:2990-2993.
Gibbons RJ, Hay DI, Childs WC III, Davis G (1990). Role of
cryptic receptors (cryptitopes) in bacterial adhesion to oral
surfaces. Arch Oral Biol 35:107S-l 14S.
Glantz PO (1969). On the wettability and adhesiveness. Odont
/tevy20(Suppl 17): 1-132.
Kilian M, Mikkelsen L, Henrichsen J (1989). Taxonomic study
of viridans streptococci: description of Streptococcus
gordonii sp. nov. and emended descriptions of Streptococcus
sanguis (White and Niven 1946), Streptococcus oralis
(BidgeandSneath 1982), and Streptococcus mitis( Andrewes
and Horder 1906). Int J System Bacteriol 39:471-484.
Knox KW, Wicken AJ (1985). Environmentally induced
changes in the surfaces of oral streptococci and lactobacilli.
In: Mergenhagen SE, Rosan B, Editors. Molecular basis of
oral microbial adhesion. Washington (DC): Am Soc
Microbiol, 212-219.
Kristoffersson K, Axelsson P, Bratthall D (1984). Effect of a
professional tooth cleaning program on interdental localized
Streptococcus mutans. Caries Res 18:385-390.
Lee SF, Progulske-Fox A, Erdos GW, Piacentini DA, Ayakawa
GY, Crowley PJ, et al. (1989). Construction and
characterization of isogenic mutants of Streptococcus mutans
deficient in major surface protein antigen PI (I/II). Infect
Immun 57:3306-3313.
Liljemark WF, Bloomquist CG, Fenner LJ, Antonelli PJ,
Coulter MC (1989). Effect of neuraminidase on the
adherence to salivary pellicle of Streptococcus sanguis and
Streptococcus mitis. Caries Res 23:141-145.
Macpherson LMD, Macfarlane TW, Stephen KW (1991). An
in situ microbiological study of the early colonization of
human enamel surfaces. Microb Ecol Health Dis 4:39-46.
Mandel ID (1989). The role of saliva in maintaining oral
homeostasis. JAmDentAssoc 119:298-304.
Marsh PD (1991). Sugar, fluoride, pH and microbial
Downloaded from adr.sagepub.com at University of Victoria on August 27, 2015 For personal use only. No other uses without permission.
SCHEIE ADV DENT RES JULY 1994
homeostasis in dental plaque. Proc Finn Dent Soc 87:515-
525.
Marshall KC, Stout R, Mitchell R (1971). Mechanism of the
initial events in the sorption of marine bacteria to surfaces.
J Gen Microbiol 68:337-348.
Nesbitt WE, Doyle RJ, Taylor KG (1982). Hydrophobic
interactions and the adherence of Streptococcus sanguis to
hydroxyapatite. Infect Immun 38:637-644.
Nyvad B, Fejerskov O (1987a). Scanning electron microscopy
of early microbial colonization of human enamel and root
surfaces in vivo. Scand J Dent Res 95:287-296.
Nyvad B, Fejerskov O (1987b). Transmission electron
microscopy of early microbial colonization of human enamel
and root surfaces in vivo. Scand J Dent Res 95:297-307.
Nyvad B, Fejerskov O (1989). Structure of dental plaque and
the plaque-enamel interface in human experimental caries.
Caries Res 23:151-158.
Nyvad B, Kilian M (1987). Microbiology of the early
colonization of human enamel and root surfaces in vivo.
Scand J Dent Res 95:369-380.
Nyvad B, Kilian M (1990). Comparison of the initial
streptococcal microflora on dental enamel in caries-active
and in caries-inactive individuals. Caries Res 24:267-272.
Olsson J, Emilson CG (1988). Implantation and cariogenicity
in hamsters of Streptococcus mutans with different
hydrophobicity. Scand J Dent Res 96:85-90.
Olsson J, Glantz PO, Krasse B (1976). Surface potential and
adherence of oral streptococci to solid surfaces. Scand J
Dent Res 84:240-242.
Olsson J, Carlen A, Holmberg K (1990). Modulation of bacterial
binding to salivary pellicle by treatment with hydrophilizing
compounds. Arch Oral Biol 35:137S-140S.
0rstavik D (1984). Initial bacterial adhesion to surfaces:
ecological implications in dental plaque formation. In: ten
Cate JM, Leach SA, Arends J, Editors. Bacterial adhesion
and preventive dentistry. Washington (DC): IRL Press,
153-166.
Quirynen M, Marechal M, Busscher HJ, Weerkamp AH,
Arends J, Darius PL, et al. (1989). The influence of surface
free-energy on planimetric plaque growth in man. J Dent
Res 68:796-799.
Quirynen M, Marechal M, Busscher HJ, Weerkamp AH,
Darius PL, van Steenberghe D (1990). The influence of
surface free energy and surface roughness on early plaque
formation. / Clin Periodontol 17:138-144.
Ritz HL (1967). Microbial population shifts in developing
human dental plaque. Arch Oral Biol 12:1561-1568.
R0lla G (1976). Inhibition of adsorption—general
considerations. In: Stiles HM, Loesche WJ, O'Brien TC,
Editors. Microbial aspects of dental caries. Washington
(DC): IRL Press, 309-324.
R0lla G, Ciardi JE, Bowen WH (1983). Identification of IgA,
IgG, lysozyme, albumin, a-amylase andglucosyltransferase
in the protein layer adsorbed to hydroxyapatite from whole
saliva. Scand J Dent Res 91:186-190.
R0lla G, Scheie AAa, Ciardi JE (1985). Role of sucrose in
plaque formation. Scand J Dent Res 93:105- 111.
Scheie AAa (1985). Bacterium derived cell-free poly saccharide
VOL. 8(2)
producing enzymes in human saliva—the effect of
mouthrinses with sucrose or glucose. In: Glantz P-O,
Leach SA, Ericson T, Editors. Oral interfacial reactions
of bone, soft tissue and saliva. Washington (DC): IRL
Press, 53-61.
Scheie AAa, R0lla G (1984). Polysaccharide production by
cell free transferases in saliva in relation to salivary
microflora. Scand J Dent Res 92:43-49.
Scheie AAa, Eggen KH, R0lla G (1987). Glucosyltransferase
activity in human in vivo formed enamel pellicle and in
whole saliva. Scand J Dent Res 95:212-215.
Schmidhuber S, Kilpper-Balz R, Schliefer KH (1987). A
taxonomic study of Streptococcus mitis, S. oralis and S.
sanguis. System Appl Microbiol 10:74-77.
Skopek RJ, Liljemark WF, Bloomquist CG, Rudny JD (1993).
Dental plaque development on defined streptococcal
surfaces. Oral Microbiol Immunol 8:16-23.
Socransky SS, Manganiello AD, Propas D, Oram V, van Houte
J (1977). Bacteriological studies of developing supragingival
dental plaque. J Periodont Res 12:90-106.
Steinberg D, Kopec LK, Bowen WH (1993). Adhesion of
actinomyces isolates to experimental pellicles. J Dent Res
72:1015-1020.
Svanberg M, Westergren G, Olsson J (1984). Oral implantation
in humans of Streptococcus mutans strains with different
degrees of hydrophobicity. Infect Immun 43:817-821.
Syed SA, Loesche WJ (1978). Bacteriology of human
experimental gingivitis: effect of plaque age. Infect Immun
21:821-829.
Theilade E, Theilade J, Mikkelsen L (1982). Microbiological
studies on early dento-gingival plaque on teeth and mylar
strips in humans. J Periodont Res 17:12-25.
Downloaded from adr.sagepub.com at University of Victoria on August 27, 2015 For personal use only. No other uses without permission.
PLAQUE FORMATION 253
van der Hoeven JS, de Jong MH, Kolenbrander PE (1985). In
vivo studies of microbial adherence in dental plaque. In:
Mergenhagen SE, Rosan B, Editors. Molecular basis of oral
microbial adhesion. Washington (DC): Am Soc Microbiol,
220-227.
van der Mei HC (1989). Physico-chemical surface properties
of oral streptococci (thesis). Groningen (The Netherlands).
van der Mei HC, de Soet JJ, de Graaff J, Rouxhet PG, Busscher
HJ (1991). Comparison of the physicochemical surface
properties ofStreptococcus rattus with those of other mutans
streptococcal species. Caries Res 25:415-423.
van Houte J, Green DB (1974). Relationship between the
concentration of bacteria in saliva and the colonization of
teeth in humans. Infect Immun 9:624-630.
van Houte J, Gibbons RJ, Banghart SB (1970). Adherence as
a determinant of the presence of Streptococcus salivarius
and Streptococcus sanguis on the human tooth surface.
Arch OralBiol 15:1025-1034.
van Houte J, Gibbons RJ, Pulkkinen AJ (1971). Adherence as
an ecological determinant for streptococci in the human
mouth. Arch Oral Biol 16:1131-1141.
van Loosdrecht MCM, Lyklema J, Norde W, Zehnder AJB
(1990). Influence of interfaces on microbial activity.
Microbiol Rev 54:75-87.
Weerkamp AH, Quirynen M, Marechal M, van der Mei HC,
van Steenberghe D, Busscher HJ (1989). The role of surface
free energy on the early in vivo formation of dental plaque
on human enamel and polymeric substrata. Microbial Ecol
2:11-18.
Weiger R, Netuschil L, van Ohle C, Brecx M (1993). Microbial
vitality during early dental plaque formation (abstract).
NOF/CED meeting, Kolding, Denmark, 152.