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Cariology IV

Microbiology of dental caries


Zdenek Broukal, Erika Lencova
Institute of Dental Research

Ontogenesis of oral microbial ecosystem


Cariogenic microorganisms
- Streptococcus mutans
- Heterofermentative lactobacilli
- Other oral microorganisms

Dental caries as an infectious disease


- Concept of the window of infectivity
- Mechanisms of oral infection
- Contributing factors

Ontogenesis of oral microbial


ecosystem I.
Several hrs after delivery
Transient flora of the birth canal and St. epidermidis from skin surface

Up to 5-6 months
S. viridans, S. oralis, S. sanguinis, lactobacilli, diphteroids

Teeth eruption
1. Infectious window period for S. mutans, G- cocci (Veilonella,
Neisseria), anaerobic streptococci, gramm negative rods

Beginning of physiological dentition exchange


Infectious window period for S. mutans, G+ anaerobic rods
(actinomycetes), G+ aerobic rods (Nocardia), G+ Rothia dentocariosa

Adolescence
G- anaerobes (Prevotella), Porphyromonas spp. , Capnocytophaga spp.,
anaerobic Vibrio spp., spirillae

Ontogenesis of oral microbial


ecosystem II.
In edentulous oral cavity of a newborn/toddler
microorganisms adhere partially to mucous
membranes, partially form planktonic microflora in
the saliva and in salivary biofilm covering mucous
membranes
After the teeth eruption 90-95 % of microbial
population forms plaque, the rest covers mucous
membranes and form planktonic microflora

Ontogenesis of oral microbial


ecosystem III.
G+ G+ cocci and
rods
G- cocci and rods

100%
80%

filaments and
fusobacteria
spirili and vibrio
spp.

60%
40%
20%
0%
0

Toothbrushing

11

No oral hygiene

13

15

days

Toothbrushing

Investigating association of
microbial flora with dental caries

1878 J. Tomes microorganisms in dentin tubules

1892 W.D. Miller chemo-parasite theory of dental caries aetiology

1922 J. Clark streptococci in dentin tubules S. mutans

1938 G. Soggnaes experimental dental caries in rodent model


Physiological breed of laboratory rodents
Cariogenic diet
Choosing suitable animal models for experimental dental caries
hamster, rat, (mice, gerbil, )
Selecting cell lines (rat Sprague-Dawley)

1963 R.W. Schaedler gnotobiotic and germ-free animal models in


research

Investigating association of microbial


flora with dental caries - evidence

1960 P.H. Keyes, R.J. Fitzgerald


Caries transmitted experimentally in hamster and rat models
with cariogenic diet that were kept together via bedding,
excrements etc.
germ-free animals with cariogenic diet had no dental caries
Mono-contamination of germ-free animals with individual oral
microbial strains of rodent and human origin
1964 cariogenic streptococci, actinomycetes

1965 J.van Houte, J. Carlsson cariogenic streptococci =


S. mutans

1967 B. Mejre and others levels of S. mutans in saliva correlate


with caries experience in humans

1978 B. Khler S. mutans is transmitted from mother to child


window of infectivity

Taxonomy I.
(Bergeys Manual of Determinative Bacteriology 2nd Edition, 1992)
Domain (Bacteria)
Kingdom (Procaryotae /Monera/)
Phylum (Firmicutes)
Class (Bacilli)
Order (Lactobacillales)
Family (Lactobacillaceae)

Genus (Streptococcus)
Species (S.

mutans)

Oral lactobacilli

Heterofermentative
main substrate: starch (glucose,
maltose, dextrins)
slow metabolism
produce only 50% lactic acid and
considerable amounts of ethanol,
acetic acid and carbon dioxide
not so low pH
L. casei
L. fermentum
L. salivarius
L. plantarum
L. brevis
L. cellobiosus
L. buchneri

Homofermentative
main substrate: glucose,
sucrose
rapid metabolism
produce more than 85% lactic
acid from glucose
low pH
L. acidophillus
L. acidophillus sensu
stricto
L. crispatus
L. gasseri
L. rhamnosus

S. mutans ecological
requirements
Presence of solid surfaces in the oral cavity
hard dental tissues, dentures, infant obturator in cleft
children

Repeated infection
Increased frequency of sucrose intake
Ecological niche
mode of existence that a species has within an ecosystem
smaller proportion of other viridans streptococci (showing
alpha haemolysis on blood agar - S. viridans, S. oralis, S.
sanguinis)

S. mutans topography

Pits and fissures


Interdental spaces
Carious defects
SM is not necessarily present on all teeth
SM can be transmitted iatrogenically
Rare on mucous membranes
Proportion in salivary planktonic bacteria
reflects the proportion in plaque saliva as
material for microbial examination
Treatment of carious defects does not decrease
the proportion of SM in plaque

S. mutans colonies morphology


Colonies of S. mutans on
the spoon contaminated
with saliva and dipped for
36 hrs in BHI broth + 5%
sucrose
Colonies of S. mutans on
MSA broth, incubated for
24 hrs, GasPak

Lactobacilli - ecological requirements


Presence of solid surfaces in the oral cavity
hard dental tissues, dentures, infant obturator in
cleft children

Increased proportion of aciduric flora


Increased frequency of sucrose intake
Carious defects
Increased amount of plaque

Lactobacilli topography
Carious defects
Proportion in salivary planktonic bacteria
partially reflects the proportion in plaque
saliva as material for microbial examination
Treatment of carious defects does decrease the
proportion of SM in plaque

Actinomycetes - ecological
requirements
Presence of solid surfaces in the oral cavity
hard dental tissues, dentures, infant obturator in
cleft children

Wide range of plaque flora


Increased frequency of sucrose intake
Carious defects
Increased amount of plaque

Actinomycetes topography

Interdental spaces
Subgingival plaque
Carious defects
Proportion in salivary planktonic bacteria does
not reflect the proportion in plaque
Treatment of carious defects does not decrease
the proportion of A. in plaque
Abundant in the human mouth and induce root
surface caries in hamsters and gnotobiotic
animals

Virulence factors cariogenicity I.


S. mutans
Energy utilisation - ,(1-2) glycosidic bonds in sucrose
Intensive glycolysis under wide range of pH
Production of EPS intermicrobial matrix in plaque
Production of IPS intracellular glycogen (ADPglucose
pyrophosphorylase (GlgC) and glycogen synthase (GlgA)
Specific adherence to acquired pellicle
Tolerance towards high sucrose concentration - up to 40 %
Microaerofilic

Heterofermentative lactobacilli
Optimum pH for metabolism 3.5 5.5
Enzymatic set for intensive glycolysis
Amylase

Virulence factors cariogenicity II.


A. odontolyticus, A. naeslundii
Glycolysis under wide range of pH
Production of IPS intracellular glycogen
Specific adherence to acquired pellicle
Wide range of proteases
Microaerophilic

Relationship of other
microorganisms to dental caries
Mixed flora of dental plaque
Microaerophilic conditions for cariogenic
microorganisms
Aciduricity (tolerance of, and growth at low pH)
Co-aggregation increased cohesion of plaque
proteases destruction of protein matrix

of

enamel/dentin
amylase hydrolysis of starch to oligosaccharides

Kochs postulates
Suspected pathogenic microorganism should be
present in all cases of the disease and absent from
healthy animals
The microorganism should be grown in pure
culture
Cells from pure culture should cause disease in a
healthy animal
The microorganism should be re-isolated and
shown to be the same as the original

Metabolic activity of cariogenic


microorganisms pH of the
environment

Production of polysaccharides by SM
Extracellular polysaccharides ( 1-3, 1-4, 1-6
glucane) adhesion factor
glucosyltranspherase (GTF)

Intracellular polysaccharides (intracellular


glycogen-like) energy source
ADPglucose pyrophosphorylase (GlgC
glycogen synthase (GlgA)

Extracellular polysaccharides

Microorganism

Extracellular polysaccharides

S. mutans

1-4, 1-3, glucane


1-6, glucane

mutane
dextrane

lactobacilli

2-6, fructane

levane

actinomycetes

2-6, fructane
1-6, glucane

levane
dextrane

Density of plaque
Fermentable
polysaccharides

EPS

Cariogenic
plaque

Noncariogenic
plaque

Chemo-parasite theory
W.D.Miller 1892
Dental caries occurs as a result
of hard dental tissues
demineralization with organic
acids end products of sucrose
metabolism of oral
microorganisms.

Ecologic factors in dental caries


S. Socransky 1992
S. mutans

S. sanguis

lactobacilli

actinomycetes

Selective ecological pressure (nutrition)


Shortage of
sucrose
neutral pH

Enough of
sucrose acidic
pH

Healthy tooth
caries

Genotype hypothesis of SM cariogenicity


M. W. Russell 1994

Microbial
agent

Defending
mechanisms
Microbial agent

Defending
mechanisms

specific
pathogen
or virulent
genotype

Defending
mechanisms

Hypotheses integration
Non-cariogenic
spectrum of
plaque
Cariogenic
spectrum of
plaque
virulent
virulent SM
SM and
and
lactobacilli
lactobacilli
genotypes
genotypes

Selective genetic
pressure
acidophilic
acidophilic
flora
flora
(lactobacilli)
(lactobacilli)

Selective
ecological
pressure
aciduric
aciduric flora
flora
(S.
(S. mutans)
mutans)

Selective physicalchemical pressure

Window of infectivity for cariogenic


microorganisms
Conditions suitable for transmission of microorganisms
to planktonic flora
Conditions suitable for adhesion of microorganisms to
dental surface/obturator/dentures
Conditions suitable for repeated infection
Contributing factors
Frequent presence of sucrose in the oral cavity
Decreased salivary secretion (during sleep)
Quantity of infection

Window of infectivity
C.W. Caulfield 1993
Beginning of the
eruption of
temporary teeth

Beginning of the
eruption of
permanent teeth

Repeated
infection

Repeated
infection

Window of
infectivity

0
6
age

12

18
24
months

Succedaneous
window of
infectivity?
30

36

6
years

Mechanism of oral infection


Transmission vector
Infection source
Transmission way
Persistent
infection

Infection entry

Repeated transmission

Contributing factors

window of infectivity

Contributing factors of oral infection


Risk behaviour of mother (siblings, father,
grandparents, carers)
Untreated carious dentition of mother/others
High levels of cariogenic microorganisms in saliva
High frequency of sucrose intake especially before
sleeping (decreased salivary flow)
Later start of tooth brushing
Administration of sweetened medications
Prolonged nursing/nursing as tranquilizing practice

Evidence of SM transpher from


mother to child
Li and Caufield - 1995: 34 pairs mother - child;
70.6 % children had SM genotype identical with
mother
De Soet et al. - 1998, 21 pairs mother child (with
lip or palate cleft); 38 % had SM genotype
identical with mother
Kozai et al. - 1999: 20 families, 51 % had SM
genotype identical with mother, 31 % with
father, 19 % other type of SM genotype
Dukov a Broukal 1997: 40 families, 60 % had
SM genotype identical with mother, 8 % with
father, 32 % other type of SM genotype

Proportions of SM and lactobacillli


in fissure plaque
Caries affected

% z TVC (total viable count)

Tooth

Caries
experience

S. mutans

S. sanguinis

lactobacilli

With caries

low

2.0

With caries

high

25

Caries free

intact
dentition

<1

13

Caries free

low

2.0

21

<1

Caries free

high

8.0

<1

Hereditary fructose intolerance


autosomal recessive disorder of fructose
metabolism first described in 1956
incidence 1:22 000 of live births
deficiency of fructose-1-phosphate aldolase (EC
2.1.2.13) activity
accumulated fructose-1-phosphate inhibits
glycogen breakdown and glucose synthesis,
thereby causing severe hypoglycaemia
following ingestion of fructose
patients develop a strong distaste for sweet
food
almost all affected persons have intact dentition
and do not have S. mutans present in the mouth

E-learning resources
www.textbookofbacteriology.net
www.db.od.mah.se/car/carhome.html
www.uic.edu/classes/peri/peri343/
http://www.dent.ucla.edu/ce/caries/index.html
http://www.dentistry.leeds.ac.uk/OROFACE/PAGE
S/micro/micro.html
http://dentistry.ouhsc.edu/intranetweb/Courses/DSA8212/CarHomePage.html
http://hebw.cf.ac.uk/oralhealth/index.html