Transcript Effects of Xylitol on S. mutans

Get the Key for Tomorrow’s
Dentistry
a symposium organized by the
KOREAN CARIES PREVENTION ASSOCIATION
and sponsored by
Danisco Cultor and Lotte Korea
Seoul April 22, 2001
The Influence and Mechanism
of Xylitol
on Mutans Streptococci
Luc Trahan, PhD
Groupe de Recherche en Écologie Buccale
Faculté de médecine dentaire
Université Laval
Québec, Canada
Principal Proposed
Mechanisms of Action of Xylitol
Lack of acid production by plaque bacteria
 Increased saliva secretion and salivary buffer capacity
 Inhibition of demineralization of sound enamel
 Remineralization of decalcified sites
 Inhibition of acid production by MS from dietary sugars
 Inhibition of MS growth
 Selection of a new less virulent MS population
 Reduction of the transmission / implantation of Ms

Aim and Content
Aim: To survey and summarize the influence and
the mechanisms of action of xylitol on
Mutans Streptococci
Background information on the dental plaque ecosystem
and the utilisation of sugars by MS
 Details of the effects of xylitol on MS
 Recent progresses on the mechanisms of action
 Summary and take home messages

Content
Background information on the dental plaque
ecosystem and the utilisation of sugars by MS
 Details of the effects of xylitol on MS

Inhibition
of growth and acid production from sugars
Selection of a new MS population

Recent progresses on the mechanisms of action
Disturbed
protein synthesis and reduced survival
Virulence properties

Summary and take home messages
The Caries Etiological Factors Constitute
a Fragile Ecosystem
Fluoride
Antiseptics
Sealant
Plaque
Tooth
CARIES
Multiple
Factors
Xylitol
Sugar
Saliva
Modification of the Plaque
Ecosystem by Sugar Consumption
SUGARS
SUGARS
(sucrose)
SM
SM
ACIDS
ACIDS
Ecological Plaque Hypothesis
and Prevention of Caries
Fermentable
Sugar
Acid
Neutral
pH
S. oralis
S. sanguis
Environmental
changes
Ecological
shift
Low
pH
MS
Lactobacilli
Marsh, P., 1991
Remineralization
Demineralization
Dietary Sugars: Source of Energy and
Carbon for the Growth of Plaque Bacteria
OD (660 mm)
1,00
Sugar could be: sucrose
0,50
0,40
0,30
glucose
lactose
mannitol
sorbitol
etc..
0,20
0,10
0,05
0,04
0,03
0,02
Xylitol
0,01
0
4
8
Time (hours)
12
24
Acidic End-products Generation
from Dietary Sugars by Plaque Bacteria
Xylitol
7,0
pH
Sugars
Sorbitol
Remineralization
6,0
Critical pH
Demineralization
5,0
4,0
0
10
20
Time (minutes)
30
40
Xylitol: a Caries Preventive
5C- Sugar Alcohol
CH2OH
H-C-OH
HO-C-H
H-C-OH
H-C-OH
CH2OH
SORBITOL
C6H14O6
CH2OH
H-C-OH
HO-C-H
H-C-OH
CH2OH
XYLITOL
C5H12O5
A NATURAL 5 CARBON SUGAR
ALCOHOL (PENTITOL)
CH2OH
C=O
HO-C-H
H-C-OH
H-C-OH
CH2OH
FRUCTOSE
C6H12O6
Sugar
Dietary Sugar Utilization
by Oral Streptococci
P
E
Sugar
ATP
ADP
Sugar~P
ADP+PPi
Polysaccharides
reserves
Sugar
ATP
Glucose~P
ADP
Glucan
Fructan
+
+
Fructose
Glucose
PTS
ATP
Sucrose
PEP
Pyruvate
ADP
ATP
Lactate
Lactic acid
Five-carbon Sugar Alcohols
Utilization by Non-oral Bacteria
Kinase
2-Keto
pentose
5-C alcohol
PERMEASE
5-C alcohol
(xylitol)
2-Keto pentose~P
DH
PTS
5-C alcohol~P
Epimerisation
Carbohydrate pathways
ADP
ATP
Lactic acid
Acetic acid
Ethanol
etc...
}
± acidic
end-products
Incapacity for Oral Bacteria to
Utilize 5-Carbon Sugar Alcohols
Kinase
2-Keto
pentose
5-C alcohol
2-Keto pentose~P
DH
Epimerisation
ADP
5-C alcohol

(xylitol)
PTS
5-C alcohol~P
Carbohydrate pathways

PERMEASE

ATP
Lactic acid
Acetic acid
Ethanol
etc...
}

± acidic
end-products
Inducible Six-carbon Sugar
Alcohols Utilization by
Mutans Streptococci
Mannitol~P
Sorbitol~P
ADP+PPi
Polysaccharides
reserves
Mannitol
PTS mtl
ATP
Glucose~P
ADP
PTS sor
ATP
Sorbitol
PEP
Pyruvate
ADP
ATP
Lactate
Lactic acid
Content
Background information on the dental plaque
ecosystem and the utilisation of sugars by MS
 Details of the effects of xylitol on MS

Inhibition
of growth and acid production from sugars
Selection of a new MS population

Recent progresses on the mechanisms of action
Disturbed
protein synthesis and reduced survival
Virulence properties

Summary and take home messages
Growth Inhibition of
S. mutans Strains by Xylitol
Strains
ATCC 27352
GS5-2
Ingbritt
NTCC 10449
6715
8E3 (fresh isolate)
67.3 (fresh isolate)
% inhibition
82
0
76
85
84
68
33
µ mole H+ / 16 mg cells (dry weight)
Effect of Xylitol on Acid Production
Rate by Resting Cells of S. mutans
40
from glucose
30
Strain % inhibition
27352 XR
0
49
27352 XS
68
6715
38
Ingbritt
20
10
+ xylitol
from xylitol
0
0
10
20
30
40
M IN U T E S
Vadeboncoeur, C., 1983
Definition of MS Sensitivity
to Xylitol
Xs wild type strains : xylitol-sensitive
Strains that are inhibited by xylitol
Xr natural mutants : xylitol-resistant
Strains that are not inhibited by xylitol
None of these strains ferment xylitol
and none produce acid from it.
Effect of the Addition of Xylitol to Growing
XS and XR S. mutans
X (± xyl)
R
+ xylitol
XS (+ H2O)
0 ,7 0
0 ,6 0
XS (+ xyl)
0 ,5 0
0 ,3 0
120
14C-xylitol accumulated
OD (660 nm)
0 ,4 0
0 ,2 0
0 ,1 0
0
1
2
Trahan, L., 1985
XS
100
80
60
40
20
XR
0
3
4
T im e ( h o u r s )
5
6
S. mutans Constitutive Fructose-PTS
also Transports Xylitol
Enzyme I
~
PEP
IIA IIB
~
Constitutive
fructose-PTS
FRUCTOSE-6~P
XYLITOL~P
P
P
IIC
FRUCTOSE
XYLITOL
HPr(His~P)
FRUCTOSE
Pyruvate
Inducible
fructose-PTS
Benchabane, H., 1997
IIA
IIB
~
HPr
~
Enzyme I~P
P
P
FRUCTOSE-1-~P
IIC
Sugar
Xylitol Transport
and Expulsion in
S. mutans
Sugar~P
ADP
ATP
PEP
ADP
FRUCTOSE
PTS
Xylitol
ATP
Pyruvate
Xylitol~P
Lactate
EXPULSION
Pi
Xylitol
Trahan L. 1996
Lactic
acid
Sugar
Establishment and
Consequences of a
Xylitol Futile Cycle
Sugar~P
ADP
ATP
PEP
ADP
FRUCTOSE
PTS
Xylitol
ATP
Pyruvate
Xylitol~P
E
Lactate
Pi
Xylitol
EXPULSION
Lactic
acid
Sugar
Absence of Constitutive
Fructose PTS Activity
in Xr Mutants
Sugar~P
ADP
ATP
PEP
ADP
Xylitol
FRUCTOSE
ATP
Pyruvate
Lactate
PTS
Lactic
acid
Effect of Xylitol and
Xylitol-5-phosphate on Selected
Glycolytic Enzymes in Vitro
Enzyme
PGI
6-PFK
1-PFK
PK
LDH
% inhibition by
xylitol
xylitol 5-P
0
0
0
0
0
>90
50
10
>90
0
Proposed Model of the Effect
of Xylitol on S. mutans
Glycogen
6-PFK
Fructose-6-P
PMF
Glucose
PGI
Fructose-1,6-di-P
DHAP
Glucose
-
Glucose-6-P
PTSman
GA3P
+
PEP
1,3-DPG
3-PG
2-PG
PK
Pyruvate
LDH
-
Lactic
acid
Lactate
PTSfru
Xylitol
Xylitol-P
Trahan L., 1996
Xylitol
Content
Background information on the dental plaque
ecosystem and the utilisation of sugars by MS
 Details of the effects of xylitol on MS

Inhibition
of growth and acid production from sugars
Selection of a new MS population

Recent progresses on the mechanisms of action
Disturbed
protein synthesis and reduced survival
Virulence properties

Summary and take home messages
R
X
In vitro Selection of
Natural Mutants
Following Growth in the Presence of
Xylitol
XYLITOL
XYLITOL
In vivo Selection of XR Natural Mutants in
Saliva and Plaque of Xylitol Consumers
XYLITOL
XYLITOL
Proportion of salivary natural XR
MS strains in patients
Number of strains
Total
XR
XS
%
XR
20
2
18
10
Xylitol-consumers 54
47
7
87
Patients
Controls
R
X
Occurrence in 1988 of
Salivary
Strains in Patients from the
Ylivieska Xylitol Study (1982-1984)
Group
N
Controls
16
Active consumers 14
Former consumers 13
% XR
35.0 ± 6.4
82.5 ± 5.7
74.6 ± 8.6
Effect of Xylitol Consumption
on the Plaque Ecosystem
SM
SM
XYLITOL
Content
Background information on the dental plaque
ecosystem and the utilisation of sugars by MS
 Details of the effects of xylitol on MS

Inhibition
of growth and acid production from sugars
Selection of a new MS population

Recent progresses on the mechanisms of action
Disturbed
protein synthesis and reduced survival
Virulence properties

Summary and take home messages
Effect of a Heat or Xylitol Stress on
14C-labelled Protein Profile of S. mutans
Densitometric Analysis of Protein Profiles of
Xr and Xs MS Strains After a Xylitol Exposure
Xs strain
Xr strain
Protein ID
Control Xylitol
exposure
Control
Xylitol
exposure
HSP-70
HSP-60
1.00
1.00
0.53
0.54
1.00
1.00
1.08
0.82
71 kDa
63 kDa
57 kDa
53 kDa
50 kDa
40 kDa
1.00
1.00
1.00
1.00
1.00
1.00
0.58
0.57
1.37
0.53
1.48
1.53
1.00
1.00
1.00
1.00
1.00
1.00
1.16
1.11
1.03
0.95
0.95
1.09
Effect of a Xylitol Shock on the Survival
Capacity of S. mutans 123.1
Number of surviving cells (CFU /mL)
S. mutans 123.1 XS
S. mutans 123.1 XR
Control
Xylitol
X/C
Control
Xylitol
X/C
3.51±0.13 x1010
1.51±0.47 x1010
0.43 1
2 .62±0.68 x109
2.78±0.38 x109
1.06 4
1.45±0.38 x109
0.654±0.09 x109
0.45 2
8.15±0.34 x108
8.44±0.42 x108
1.03 5
3.07±0.60 x109
2.19±0.05 x109
0.71 3
1.60±0.39 x109
1.91±0.43 x109
1.19 6
1, 2, 3 p<0.0003
4, 5, 6 p>0.11
Content
Background information on the dental plaque
ecosystem and the utilisation of sugars by MS
 Details of the effects of xylitol on MS

Inhibition
of growth and acid production from sugars
Selection of a new MS population

Recent progresses on the mechanisms of action
Disturbed
protein synthesis and reduced survival
Virulence properties

Summary and take home messages
Preliminary Results on the Attenuated
Virulence Properties of XR MS Strains
Selected by Xylitol ...
When XR strains are compared with corresponding XS
strains the accumulated evidence suggest:
a
reduced acid production from sugars
a modified extracellular polysaccharides synthesis
a modified bacterial agglutination/aggregation capacity
an elevated glucan-binding protein C elevated 20 folds
Preliminary Results on the Attenuated
Virulence Properties of XR MS Strains
Selected by Xylitol
an
increased dextran-dependent aggregation capacity
a reduced adherence to saliva coated hydroxyapatite
and glass surfaces
a reduced cariogenicity in a rat model
an absence of DnaK and GroEL stress proteins
response to a xylitol exposure
a larger bacterial cells shedding from plaque to saliva
Content
Background information on the dental plaque
ecosystem and the utilisation of sugars by MS
 Details of the effects of xylitol on MS

Inhibition
of growth and acid production from sugars
Selection of a new MS population

Recent progresses on the mechanisms of action
Disturbed
protein synthesis and reduced survival
Virulence properties

Summary and take home messages
Summary of the Effects of Xylitol on
Mutans Streptococci ...


A variable intensity inhibition of growth on most
fermentable sugars resulting from:
 the
establishment of an energy spending futile cycle
 the
strong inhibition of glycolytic pathway enzymes
An inhibition of acid production from dietary sugars
also resulting from the above mentioned mechanisms
Summary of the Effects of Xylitol on
Mutans Streptococci

A selection of XR MS strains with apparently less
virulent properties

A disturbance of protein synthesis, including stress
proteins DnaK and GroEL, resulting in a reduced
capacity to survive further exposure to xylitol

A reduced capacity to be transmitted from mother to
child
Clinical Benefits for Patients
Chewing Xylitol Gum
The accumulated evidence suggests:
a reduced caries incidence via:
an increased saliva secretion and salivary buffer capacity
an inhibition of demineralization of sound enamel
a remineralization of decalcified sites
an inhibition of acid production from dietary sugars
 a less acidogenic and less aciduric plaque ecosystem

Additional Clinical Benefits from the
Inhibitory Action of Xylitol on MS
The accumulated evidence suggests:

a reduction of:
plaque indices
the amount of plaque accumulated
the number of MS in plaque and saliva
the capacity to transmit MS strains

a selection of Xr MS strains with:
apparently less virulent properties
a loss of MS strains from plaque to saliva
Take Home Message # 1

Xylitol influences positively the plaque microflora in
establishing ecological conditions less appropriate for
the low-pH bacteria producing acidic end-products
and thus beneficial for the patients

Long and/or frequent exposure to xylitol does not
result, for oral bacteria, in adaptation to metabolize
the sugar-alcohol
Take Home Message # 2

Xylitol is not only unfermentable by oral bacteria but
it also inhibits most mutans streptococci

Biochemical mechanisms explaining the inhibition of
the growth of, and acid production by, mutans
streptococci have been well known for a few years

Knowledge on other mechanisms involving disturbed
proteins synthesis by xylitol and reduced survival of
MS to repeated exposures to xylitol is progressing
Take Home Message # 3

The occurrence of xylitol-resistant strains in xylitol
consumers is explained by a simple selection of
natural mutants

Xylitol-resistance of MS does not impair its use in
caries prevention and is most probably beneficial for
the patients

Knowledge of the hypothesized less virulent
properties of xylitol-resistant strains is progressing
This presentation is the result of various
studies conducted with numerous
graduate students and colleagues from
Québec and Finland
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
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