Comparative study of preventive protocols in children at high cariogenic risk
The aim of this study was to compare the efficacy of two preventive protocols -fluoride gel (F) alone or combined with chlorhexidine varnishes (CHX) - on sialochemical, clinical and microbiological parameters in a group of children at high cariogenic risk Two therapeutic-preventive protocols were applied in 73 children at high cariogenic risk (average age 6.2 +/- 1.4 years old) and clinical parameters (simplified oral hygiene index: OHI-S; decayed, missing and filled teeth: dmf index; sugar intake and exposure to fluoride), as well as sialochemical parameters (salivary pH and flow, buffer capacity) and microbiological parameters (CFU/mg of dental biofilm of Streptococcus mutans group) were recorded and correlated before and after the protocols. Association was found between parameters that cause deficient control of dental biofilm: high values of OHIS index, CFU/mg dental biofilm, sugar intake and the d component of dmft index, and lower values of salivary flow rate and buffer capacity. After the protocols, a significant decrease was found in OHI-S and CFU/mg dental biofilm. No significant difference was found with children's gender and age. The association observed between OHI-S and cariogenic bacteria emphasizes the importance of prevention, especially regarding the oral health of the most vulnerable children. The early inclusion of F associated with CHX in the initial step of preventive and therapeutic protocols would provide benefits regarding oral microbe control while children acquire new habits of oral hygiene.
Acta Odontol. Latinoam. 2012 ISSN 0326-4815 Vol. 25 Nº 2 / 2012 / 218-223
El objetivo de este trabajo fue evaluar la eficacia de dos pro-
tocolos preventivos - fluoruro (F) sólo o combinando con
clorhexidina (CHX)- sobre indicadores clínicos, sialoquími-
cos y microbiológicos, en una población de niños de alto
Se aplicaron dos protocolos terapéutico-preventivos en 73
niños en edad escolar de alto riesgo cariogénico y se determi-
naron y correlacionaron parámetros clínicos (índice de higiene
oral simplificado IHO-S, ceo-d, consumo de azúcar y exposi-
ción a fluoruros), sialoquímicos (pH y flujo salival, capacidad
amortiguadora) y microbiológicos (UFC/mg de biofilm dental
de Streptococcus grupo mutans) antes y después de la apli-
cación de los tratamientos.
Se observó una asociación entre los parámetros que producen
una deficiente control de placa bacteriana: altos valores de
IHO-S, de UFC/mg biofilm dental, de consumo de azúcar, del
componente c del índice ceo-d y los menores valores de flujo
salival y de capacidad amortiguadora. Luego de la aplicación
de los tratamientos, se observó una disminución significativa
de IHO-S y UFC/mg biofilm dental. No se observaron diferen-
cias significativas con el género y la edad de los niños.
La asociación observada entre los niveles de higiene oral y de
bacterias cariogénicas enfatiza la importancia de la preven-
ción y atención de la salud de los niños más vulnerables. La
incorporación del F asociada a la CHX en la etapa inicial de
los protocolos terapéutico-preventivos ofrecería temprana-
mente beneficios en el control microbiano mientras se
incorporan hábitos de higiene oral.
Palabras clave: niños, susceptibilidad a caries dental, saliva,
The aim of this study was to compare the efficacy of two preventive
protocols - fluoride gel (F) alone or combined with chlorhexidine
varnishes (CHX) - on sialochemical, clinical and microbiological
parameters in a group of children at high cariogenic risk.
Two therapeutic-preventive protocols were applied in 73 chil-
dren at high cariogenic risk (average age 6.2±1.4 years old)
and clinical parameters (simplified oral hygiene index: OHI-
S; decayed, missing and filled teeth: dmf index; sugar intake
and exposure to fluoride), as well as sialochemical parameters
(salivary pH and flow, buffer capacity) and microbiological
parameters (CFU/mg of dental biofilm of Streptococcus mutans
group) were recorded and correlated before and after the pro-
tocols. Association was found between parameters that cause
deficient control of dental biofilm: high values of OHIS index,
CFU/mg dental biofilm, sugar intake and the d component of
dmft index, and lower values of salivary flow rate and buffer
capacity. After the protocols, a significant decrease was found
in OHI-S and CFU/mg dental biofilm. No significant difference
was found with children’s gender and age.
The association observed between OHI-S and cariogenic bac-
teria emphasizes the importance of prevention, especially
regarding the oral health of the most vulnerable children. The
early inclusion of F associated with CHX in the initial step of
preventive and therapeutic protocols would provide benefits
regarding oral microbe control while children acquire new
habits of oral hygiene.
Key words: dental caries susceptibility, children, saliva, fluo-
COMPARATIVE STUDY OF PREVENTIVE PROTOCOLS
IN CHILDREN AT HIGH CARIOGENIC RISK
María C. Martínez
, Betina Tolcachir
, Alfonsina Lescano de Ferrer
, María A. Bojanich
Silvina R. Barembaum
, Silvia E. Calamari
, Ana I. Azcurra
Social and Preventive Dentistry Department, School of Dentistry, National University of Córdoba, Argentina.
Oral Biology Department, School of Dentistry, National University of Córdoba, Argentina.
ESTUDIO COMPARATIVO DE PROTOCOLOS PREVENTIVOS
EN NIÑOS DE ALTO RIESGO CARIOGÉNICO
Knowledge which has come to light in recent years
regarding dental caries has led to a new paradigm and
comprehensive approach to its treatment. The multi-
factorial etiology and complex pathogenesis of dental
caries involve microbiological, histological, immune
and biochemical aspects, which lead to a diagnostic
criterion and therapeutic reformulation for a preven-
tive approach. In odontopediatric clinical practices, it
is essential to unify the criteria for a certain diagnosis,
identifying individual children or groups of more vul-
nerable children. With this purpose, clinical indexes
have been associated to sialochemical and microbio-
. The former arise from dental medical
ACTA-2-2012-SEGUNDAS:3-2011 19/10/2012 10:41 a.m. Página 218
history, both personal and family histories, cultural
and socio-economic context, and clinical dental-stom-
atological examination. Among sialochemical tests,
salivary buffer capacity, pH and flow rate are consid-
ered as risk factors; while microbiological tests refer
to colony-forming units and species identification
Considering these indicators, a patient can be cate-
gorized as being at high or low cariogenic risk,
based on which it is possible to devise preventive
and therapeutic strategies tending towards the con-
trol each of the factors involved. It is therefore
essential to plan preventive actions according to the
needs of each group of children, thus reducing costs
and improving efficacy
In general, preventive actions in children at high cario-
genic risk tend to improve host resistance, control cari-
ogenic food intake and, above all, control the
. The two latter can be managed
by changing eating habits and oral hygiene. Depend-
ing on their lifestyle and family environment, children
may take some time to assimilate these behavioral
changes, which require intensive interaction among the
child, his/her family and health care professional.
While these changes are being implemented, one ther-
apeutic action to control dental biofilm is the use of a
chemical that could affect adhesion, colonization
and/or bacterial metabolism, such as fluorides (F)
. F has an important inhibitory
action on microbial biofilm, conditioning the neces-
sary bacterial metabolic processes for the development
and maturation of dental biofilm through enzymatic
. CHX controls the microbiological load in
patients, in particular reducing salivary counts for
Streptococcus mutans, which is highly sensitive to it
The aim of this study was to assess and compare the
efficacy of two preventive protocols, one applying only
F and the other combining F with CHX, on sialochemi-
cal, microbiological and clinical parameters and index-
es, in a population of children at high cariogenic risk.
MATERIAL AND METHODS
The study was a randomized clinical trial on children
with clinical diagnosis of high cariogenic risk (n=73,
both sexes, average age 6.2±1.4 years old), who
received care at the Department of Pediatric Dentistry,
School of Dentistry, National University of Cordoba,
Argentina. The study was approved by the Ethics
Committee, School of Medical Sciences, National
University of Cordoba, Argentina, and
parents/guardians of all subjects signed informed con-
sent after a detailed explanation of the protocol. No
patient had any history of systemic illness, and all
underwent standardized detailed examination of the
oral cavity. High cariogenic risk criteria were estab-
lished by means of the following clinical indicators:
dental caries lesions over the past year including white
spot lesions, presence of dental pits and fissures,
Greene and Vermillion’s oral hygiene index simpli-
(OHI-S) ≥ 0.8, sugar intake > 4 per day and low
frequency of visits to the dentist
. The following
parameters were determined in this high caries risk
group: Microbiological parameters: counts of S.
mutans group in dental biofilm collected from the lin-
gual face of mandibular molars with a periodontal
probe, which was immediately placed in sterile physi-
ological solution in pre-weighed tubes. Samples were
immediately cultivated (mg of dental biofilm collect-
ed) in duplicate, in mitis salivarius agar with bacitracin
, Becton Dickinson, France), incubated for
48 hr at 37ºC in microaerophilic environment;
colonies were counted (CFU/mg of dental biofilm).
Biochemical identification was performed by means
of morphological characteristics and conventional bio-
chemical tests (Diatabs
, Doughnut Diagnoses, Den-
. Sialochemical parameters: Saliva samples
(stimulated by chewing a piece of 3x3 cm2 paraffin
film) were collected by direct salivation between 9 and
11 a.m. for 5 min in iced graduated tubes. Patients had
been advised not to eat anything for at least 2 hours
before the sample collection. Salivary flow rate, buffer
capacity and pH were determined immediately
pH meter, USA)
All children received initial preventive therapy con-
sisting of oral hygiene techniques, dietary advice and
caries inactivation. After that, they were randomly
divided into two groups for the application of preven-
tive protocols under evaluation: Group I: (n=36)
received a weekly topical application of fluoride gel
for four weeks (F: acidulated gel 1.23% NaF, Gelato,
Deepak, USA), and Group II: (n=37) received com-
bined therapy, alternating topical acidulated fluoride
application with 1% chlorhexidine varnish, for the
same period and frequency (F + CHX: Periobacter
Prof, NAF, Argentina). Clinical, sialochemical and
microbiological parameters were assessed again one
week after the final application of the different treat-
ments. The analysis of correspondence association
required operationalization of variables and determi-
nation of levels, considering the median for the quan-
titative variables: level 1: lower or equal to the
Prevention in children and cariogenic risk 219
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median; level 2: above the statistical median. For
OHI-S, ≥ 0.8 values were included in level 2, while
level 1 corresponded to < 0.8 values.
Categorical variables were summarized as propor-
tions, and comparisons of groups were made by
Student’s t test for paired samples, ANOVA and cor-
respondence analysis, considering a significance
level of p<0.05 (Infostat Professional, version 2007).
The sample was made up of 61.8% female and
39.2% male participants. Clinical, sialochemical
and microbiological variables at baseline (pretreat-
ment) are shown in Table 1.
Association between the parameters studied and clini-
cal indexes before the application of the treatment was
carried out by multivariate correspondence analysis.
Association was found between deficient levels of
parameters that control cariogenic dental biofilm: high
values for OHI-S, CFU/mg of dental biofilm, and sugar
intake, and poor buffer capacity; as well as high levels
of active caries with the d component of the dmf-t index
and lower values of salivary flow rate. Association was
also found between lower levels of OHI-S and
CFU/mg of dental biofilm and better buffer capacity.
Lower levels of variable active caries, the d compo-
nent of the dmf-t index and higher sugar intake values
were associated with higher values of salivary flow rate
(inertia 37.85%). No association was observed
between salivary pH and children’s gender; neverthe-
less, lower buffer capacity was found in female
patients. Regarding children’s age, a tendency towards
lower salivary flow rate was observed in younger chil-
dren (≤ 5 years old, p=0.08), probably related to the
difficulty in salivating and collecting saliva.
After both treatments, the levels of CFU/mg dental
biofilm decreased significantly in relation to pretreat-
ment values (F: p=0.004; F + CHX: p=0.001). Com-
paring both treatments, the decrease
in CFU/mg dental biofilm was greater
in Group II (F + CHX treatment)
(p <0.0001) (Fig. 1). Moreover, OHI-
S decreased significantly with treat-
ment (treatment F: p=0.0002,
treatment F + CHX: p<0.0001) (Fig.
2). No significant difference was
found between treatments. Higher
values of CFU and OHI-S were
observed in children over 6 years old,
with more significant differences. No
significant difference was found
between girls and boys.
220 M. C. Martínez, et al.
Acta Odontol. Latinoam. 2012 ISSN 0326-4815 Vol. 25 Nº 2 / 2012 / 218-223
Table 1: Clinical, sialochemical and microbiological parameters
of children before treatments.
Pre- Treatment Values
5.2 ± 2.4
component 3.7 ± 2.3
component 0.9 ± 1.4
OHI-S 1.5 ± 0.5
Moments of sugar per day 4.5 ± 1.5
Sialochemistry and Salivary flow 0.78 ± 0.36
microbiological Salivary pH 7.51 ± 0.35
parameters Salivary buffer capacity 5.92 ± 0.67
CFU / mg biofilm (2.06 ± 3.80) x 10
Data are expressed as mean ± standard deviation
Fig. 2: Effect of therapeutic preventive protocol treatment on
the oral hygiene index (OHI-S).
Fig. 1: Effect of therapeutic preventive protocol treatment on
S. mutans levels (CFU/mg biofilm).
ACTA-2-2012-SEGUNDAS:3-2011 19/10/2012 10:41 a.m. Página 220
Although there is much research on cariogenic risk
employing clinical, sialochemical and microbiolog-
ical parameters as risk indicators at baseline and
after treatment, there is very little information about
young school-age population in our region. The high
prevalence of caries among children warrants fur-
ther research on the subject, which encouraged us to
contribute this study. Clinical, microbiological and
sialochemical variables were selected from biblio-
and our clinical experience
allows us to establish points of reference. Although
there are many indexes for quantifying the presence
of oral biofilm, in odontopediatric practices, the
Green and Vermillion Simplified allows biofilm in
primary teeth to be measured quickly and easily, and
to be associated to oral hygiene status
Like other researchers, we have established assess-
ment protocols and treatments for children at high car-
iogenic risk adapted to our context. Such protocols are
based on oral hygiene, history of dental caries, dietary
habits, state of oral health, oral microbial load, sali-
vary flow rate and pH
. In this regard, Zero et al.
found that the model for the assessment of risk factors
depends on the population studied; Peterson et al.
reinforces this idea, suggesting that geographical area
can be considered as a predictor, as well as social fac-
tors, lifestyles, and cultural and economical aspects.
Many protocolized clinical trials associate the pro-
motion and education of oral health with the applica-
tion of chemical substances, combining mainly
fluoride and chlorhexidine
, as well as studying
the relationship between S. mutans levels and differ-
ent stages of early childhood caries
and in agreement with Rijkmo et al.
, it is difficult
to compare results because of the diversity of
methodological conditions and heterogeneity of pop-
ulations. This is the case for S. mutans counts, report-
ed in our study as CFU/mg dental biofilm, which are
not comparable with some published values, usually
reported as CFU/mL saliva
. In this study we have
considered the fact that CFU associated to dental
biofilm are much more closely related to cariogenic
risk than are planktonic bacteria in saliva.
Our study found an improvement in the level of OHI-
S and CFU/mg dental biofilm after treatments with
more statistically significant levels when the sample
was stratified according to age, which could be relat-
ed to a greater efficacy of educational protocols and
greater motor ability in the older group of children.
Regarding the observed dmf-t index values, the high
prevalence of the d component observed with a low
level of the f component indicates the lack of atten-
tion to oral health among these children; moreover,
high values of the d component determine, together
with other parameters, a future prognosis of high
. These authors noted how powerful
the d component is as a predictor of caries on per-
manent dentition and therefore, how important edu-
cational programs to recover oral health are.
In our study, salivary flow rate values corresponded
to high and medium risk values according to Bordoni
, medium risk values observed by Sánchez-
Pérez et al.
and were lower than those reported by
Tukia-Humala et al.
. These findings reinforce the
need for local studies describing the special charac-
teristics of vulnerable children in our region.
The salivary buffer capacity values observed in our
study did not correlate with those reported in other
papers according to caries risk and age
papers found that salivary buffer capacity and flow
rate improved after tooth brushing techniques were
taught, emphasizing the importance of implement-
ing preventive and educational programs. Our
results showed higher values of salivary flow rate
than those reported in the literature according to the
cariogenic risk observed
The decrease in S. mutans counts and improvement
in oral hygiene index after fluoride application com-
bined with CHX varnishes, added to the educational
motivation and preventive and educational actions,
were similar to those reported by Petersson et al.
confirmed the benefits of applying
chemical compounds in order to reduce the bacterial
load and control caries, emphasizing the benefits of
the application of fluoride combined with CHX var-
nishes in patients at high cariogenic risk.
showed the antimicrobial properties of
CHX, although its effectiveness on long-term preven-
tion was not proved. Experimental studies with both
antimicrobial agents also showed a greater effective-
ness in the prevention of biofilm development, and
thus of enamel demineralization
. The association
observed between oral hygiene index and cariogenic
bacteria counts in this study supports the need to
emphasize preventive actions for maintaining oral
health in the most vulnerable children; this could also
demonstrate that OHI-S reflects pathogenic Strepto-
coccus levels, although it must be remembered that
dental caries is a multifactorial disease, and other fac-
tors such as oral hygiene and access to fluoridated
products are also determinants of the carious process.
In this sense, prevention should be based on diet con-
trol, teaching oral hygiene techniques and reinforcing
Prevention in children and cariogenic risk 221
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ACTA-2-2012-SEGUNDAS:3-2011 19/10/2012 10:41 a.m. Página 221
host resistance. Nevertheless, including CHX during
the initial phase of clinical odontopediatric actions in
children at high cariogenic risk offers benefits in
microbial control while dietary and oral hygiene habits
are being promoted. This study shows that the appli-
cation of CHX together with fluoride varnishes is an
important complement to initial therapy in children at
high cariogenic risk and provides better control of
some indicators than does using only fluoride. Never-
theless, and considering that our work cannot be inter-
preted as an epidemiological study, further studies are
needed to assess the efficacy on long-term prevention.
222 M. C. Martínez, et al.
Acta Odontol. Latinoam. 2012 ISSN 0326-4815 Vol. 25 Nº 2 / 2012 / 218-223
This work was supported by the Secretary of Science and Tech-
nology (SeCyT) of the National University of Cordoba, Nº 159/07.
Dra. Ana I. Azcurra
Oral Biology Department, Faculty of Dentistry, National
University of Córdoba. Haya de la Torre SN, Pabellón Argentina
Ciudad Universitaria- 5016, Córdoba, Argentina
1. Pitts NB. How the detection, assessment, diagnosis and
monitoring of caries integrate with personalized caries
management. Monogr Oral Sci 2009;21:1-14.
2. Battellino LJ, Cornejo LS, Dorronsoro de Cattoni ST, Luna
Maldonado de Yankilevich ER, Calamari SE, Azcura
AI, Virga C. Oral health status evaluation of pre-school
children: longitudinal epidemiologic study (1993-1994),
Córdoba, Argentina. Rev Saude Publica 1997;31:272-81.
3. Cornejo LS, Brunotto M, Hilas E. Salivary factors associ-
ated to the prevalence and increase of dental caries in rural
schoolchildren. Rev Saude Publica 2008;42:19-25.
4. Ekstrand J. Oliveby A. Fluoride in the oral environment.
Acta Odontol Scand 1999;57:330-3.
5. Okada M, Soda Y, Hayashi F, Doi T, Suzuki J, Miura K,
Kozai K. Longitudinal study of dental caries incidence asso-
ciated with Streptococcus mutans and Streptococcus sobrinus
in pre-school children. J Med Microbiol 2005;54:661-5.
6. Duckworth R. Fluoride in saliva and plaque following use of
fluoride-containig mouthrinses. J Dent Res 1987;66:1730-4.
7. Forgie A, Paterson M, Pine C. A randomized controlled trial
of caries-preventive efficacy of a chlorhexidine-containing
varnish in high-caries-risk adolescents. Caries Res
8. Edelstein B, Tinanoff N. Screening preschool children for den-
tal caries using a microbial test. Pediatr Dent 1989;11:129-32.
9. Matthijs S, Adriens P. Chlorhexidine varnishes: a review. J
Clin Periodontol 2002;29:1-8.
10. Greene J, Vermillion J. The simplified oral hygiene in dex.
J Am Dent Assoc 1964;68:7-13.
11. Maltz M, Jardim JJ, Alves LS Health promotion and dental
caries. Braz Oral Res 2010;24:18-25.
12. de Carvalho FG, Silva DS, Hebling J, Spolidorio LC, Spoli-
dorio DM. Presence of mutans streptococci and Candida
spp. in dental plaque/dentine of carious teeth and early
childhood caries. Arch Oral Biol 2006;51:1024-8.
13. Larmas M. Saliva and dental caries: diagnostic tests for
normal dental practice. Int Dent J 1992;42:199-208.
14. Zero D, Fontana M, Lennon A. Clinical applications and
outcomes of using indicators in caries management. J Dent
15. Peterson L, Twetman S. Efficacy of a chlorhexidine and a
chlorhexidine –fluoride varnish mixture to decrease interden-
tal levels of mutans streptococci. Caries Res 1997;31:361-5.
16. Peterson LG, Magnusson K, AnderssonH, Almquist B, Tweat-
man S. Effect of quarterly treatments with a chlorhexidine
and fluoride varnish on proximal caries in caries-susceptible
teenagers: a 3 years clinical study. Caries Res 2000;34:140-3.
17. Twetman S, Peterson L. Effect of different chlorhexidine
varnish regimens on mutans streptococci levels in interden-
tal plaque and saliva. Caries Res 1997;31:189-93.
18. Parisotto TM, Steiner-Oliveira C, Duque C, Peres RC,
Rodrigues LK, Nobre-dos-Santos M. Relationship among
microbiological composition and presence of dental plaque,
sugar exposure, social factors and different stages of early
childhood caries. Arch Oral Biol 2010;55:365-73.
19. van Rijkmo H, Truin G, van’t Hof A. A meta analysis of
clinical studies on the caries inhibiting effect of chlorhexi-
dine treatment. J Dent Res 1996;75:790-5.
20. Habibian M, Roberts G Lawson M, Stevenson R, Harris S
Dietary habits and dental health over the first 18 months of
life. Community Dent Oral Epidemiolog 2001;29:239-46.
21. Tinanoff N, Palmer C. Dietary determinants of dental caries
and dietary recommendations for preschool children. J Pub-
lic Health Dent 2000;60:197-206.
22. Bordoni N. Programas preventivos. Propuesta para la orga-
nización de la atención clínica . Rev Asoc Odontol Argent
23. Sánchez-Pérez L, Saénz-Martínez L, Irigoyen-Camacho E,
Zepeda-Zepeda M, Acosta-Gío AE, Méndez-Ramírez I.
Stimulated saliva flow rate patterns in children: A six-year
longitudinal study. Arch Oral Biol 2009;54:970-5.
24. Tukia-Kulmala H, Tenovuo J. Intra- and inter-individual
variation in salivary flow rate, buffer effect, lactobacilli,
and mutans streptococci among 11- to 12-year-old school-
children. Acta Odontol Scand 1993;51:31-7.
25. Ligtenberg AJ, Brand HS, Bots CP, Nieuw Amerongen AV.
The effect of toothbrushing on secretion rate, pH and buffer
ing capacity of saliva. Int J Dent Hyg 2006;4:104-5.
26. Scheutz F, Matee MI, Poulsen S, Frydenberg M. Caries risk
factors in the permanent dentition of Tanzanian children: a
cohort study (1997-2003). Community Dent Oral Epidemi-
27. Splieth C, Steffen H, Rosin M, Welk A. Caries prevention
with chlorhexidine-thymol varnish in high risk schoolchild-
ren. Community Dent Oral Epidemiol 2000; 28:419-23.
28. Twetman S. Antimicrobials in future caries control? A
review with special reference to chlorhexidine treatment.
Caries Res 2004 38: 223-9.
29. Ccahuana-Vásquez RA, Cury JA. S. mutans biofilm model
to evaluate antimicrobial substances and enamel deminer-
alization. Braz Oral Res 2010;24:135-41.
ACTA-2-2012-SEGUNDAS:3-2011 19/10/2012 10:41 a.m. Página 222