Yamada et al.

Asian Pac J Dent 2013; 13: 5-10.

Three months variation of the incipient tooth caries in Japanese
school children
Hidenori Yamada, DDS, PhD,a Yoshiaki Nomura, DDS, PhD,a Shinpei Tsuge, DDS,PhD,b and Nobuhiro Hanada, DDS, PhDa
abDepartment of Translational Research, Tsurumi University School of Dental Medicine, Yokohama, andTsuge Dental Clinic, Gifu, Japan

Purpose: To confirm the incipient caries so called white spot can be reversible process of demineralization andremineralization.
Materials and Methods: A total of 128 pupils were selected for the analysis. Dentists conducted oral
examinations under light using dental mirrors. Images of the labial surface of the maxillary right and left central
incisors of all the subjects captured using the quantitative light-induced fluorescence (QLF) systems. Saliva
samples were obtained by having subjects chew a gum base for at least 3 minutes that contained no taste orflavor additives. Salivary pH value was evaluated using pH testing paper.
Results: In this study, we revealed the 3 months variation of the incipient caries in Japanese school children.
Even DMF/DMFT was a low level, over one third of all pupils (37.5%) had incipient caries detected by thequantitative light-induced fluorescence (QLF). After 3 months, 11 pupils (8.6%) had disappeared the incipientcaries in central incisors. However, newly incipient caries were appeared in residuary pupils. Finally, a half ofall pupils had incipient caries detected.
Conclusion: In this study, we revealed the three months variation of the incipient caries in Japanese school
children using the quantitative light-induced fluorescence (QLF). We found that 8.6% of the subjects haddisappeared the incipient caries. This data shows that the incipient caries is a reversible process of toothdemineralization and remineralization. (Asian Pac J Dent 2013; 13: 5-10.)
Key Words: demineralization, incipient caries, quantitative light-induced fluorescence, remineralization

Introduction
With the advances of the knowledge of the dental caries, incipient caries so called white spot or early caries
can be reversible disease process of demineralization and remineralization.1 Dental caries is now consideredbeing as a dynamic process in which multiple factor determine whether process begins, progresses, stops orreverses.2 The demineralization and reminarelization of incipient caries are intimately related and occur based
upon the dental biofilm and the availability of the refined carbohydrates for the bacterial fermentation to organicacid. There are varied biological factors in saliva that protect the tooth surface from caries development.3
It is now accepted that several strategies can be used for facilitate the reminarelization process. It is including
treatment such as the concentrated use of topical fluoride, improved oral hygiene, and reducing the number ofexposures to the fermentable carbohydrate.2,3 The prevention, control, and treatment should be based on theappropriate diagnosis and detection of the disease in its earliest stages. Under these conditions, it has been
advocated that highly sensitive and accurate method for the detection of dental caries is necessary. In theseconcepts, the light or fluorescence applied method had been developed.4,5 Quantitative laser/light-induced
fluorescence (QLF) is a technique for the detection, quantification, and longitudinal monitoring of incipientcaries. This technique is noninvasive and nondestructive. Some studies have shown that QLF is a highly
sensitive method that is suitable for the detection of visually undetected initial caries lesions.6-9 However, theclinical or epidemiological data by the QLF is not sufficient, and few data is available for the evaluation of thedental caries progression especially without any intervention.
Thus, we conducted an observational survey for the school children during 3 months without intervention.
After 3 months, we evaluated the progression or recovery of the incipient caries. We also analyzed the so-called5

Yamada et al.

Asian Pac J Dent 2013; 13: 5-10.

risk factors for the dental caries including salivary cariogenic bacterial levels.

Materials and Methods
Study design
Pupils received a first routine dental examination with QLF and the saliva test. Then, 3 months later thesecond dental examination was performed. The study population was pupils from 4th grade elementary school
children (nine or ten years old) residing in the Gifu Prefecture, Japan. The fluoride concentration of the drinking
water in this area is less than 0.8 ppm. Children and parents were informed of the survey in advance atparent-teacher’s meeting, and we also obtained informed consent. Among the 147 pupils, 128 pupils (71 males,55.5% and 57 females, 45.5%) were selected for the analysis. This study was approved by the ethical committeeof the Tsurumi University School of Dental Medicine (No. 430, 2007).
Clinical examination
Dentists conducted oral examinations under light using dental mirrors. Teeth conditions were scored as sound,decayed or filled. The decayed or filled teeth were identified according to the standard method and criteria ofJapanese Association of School Dentists. Saliva samples were obtained by having subjects chew a gum base forat least 3 minutes that contained no taste or flavor additives. Salivary pH value was evaluated using pH testing
paper (Toyo Roshi, Tokyo, Japan).
Fluorescence image acquisition
Images of the labial surface of the maxillary right and left central incisors of all the subjects captured using
light-induced fluorescence systems. For acquisition of the images, QLF system (QLF/cin 1007, Inspecktor
Research system B.V. Amsterdam, Netherland) was used. The images of the tooth surface were acquired using aminiature CCD camera inside the hand piece through a 520 nm high pass filter, transmitting only light atwavelength less than 520 nm.
Image analysis
Average fluorescence loss (%) and size (cm2) of white spot lesions were determined by QLF in a manner
described by Ando et al. 2001.10 Fluorescence loss (ΔF; %) and area of the lesion (A; mm2) and fluorescenceloss integrated over the lesion area (ΔQ; ΔF x A; % x mm2) were determined. The presence or absence of initialcaries lesions was diagnosed by QLF. Fluorescence radiance level less than 95% of sound surface wereconsidered to be white spot lesions. Another variable ΔQ (%, cm2) that defined as fluorescence loss integrated
over the lesion size was determined.
Microbial procedures
The saliva samples were vortexed for 10 s and inoculated onto Mitis-Salivarius agar (MS agar, Difco, Tokyo,Japan) medium for total streptococci.

We used modified MSB agar medium, which is MS agar (Difco)

supplemented with 20% sucrose (Wako Pure Chemicals Co., Osaka, Japan), 20 mg/mL Yeast Extract (BectonDikinson MD, USA), 0.25 U bacitracin (Sigma, Inc., St. Louis, MO, USA), 10 mg/mL Colistin (Wako Pure
Chemicals Co., Osaka, Japan), 10 mg/mL nalidixic acid (Wako), 4 mg/mL gramicidin (Sigma), and 1% tellurite
solution for the mutans streptococci. We used Rogosa SL (Nippon Becton Dickinson Company, Ltd. Tokyo,Japan) agar medium for lactobacilli. These media were inoculated using an EDDY JET spiral system (GunzeSangyo, Inc., Tokyo, Japan). MS agar and modified MSB agar media were incubated for 48 hours anaerobically.
After the anaerobic incubation, we counted the colonies to determine the number of bacteria per ml whole-saliva

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Asian Pac J Dent 2013; 13: 5-10.

on each agar medium using a spiral systems counting grid.
Statistical Analysis
The descriptive analysis of the baseline characteristic of the subjects participated in this study were calculated
separately by male and female. The p-values were calculated by Mann-Whitney U test. The statisticallysignificance of the changes of ΔF and ΔQ values was calculated by Wilcoxon signed-rank test. To investigate
the co-relation of the salivary cariogenic bacterial levels and changes of the ΔF values, the subjects wereclassified into four groups by the changes of ΔF values. The p-values were calculated by Kruscal-Wallis test.
These analyses were carried out by SPSS 14.0 (SPSS, Tokyo, Japan).

Results
The baseline characteristics of the oral conditions and the salivary levels of the cariogenic bacteria of thesubjects were given in Table 1. The decayed or filled teeth in Table 1 were identified by school dentists. Caries
prevalence and mean DMFT in the permanent dentition between male (DMFT 0.23) and female (DMFT 0.26)
were remarkably low. The subjects who had at least one DMF were 18 pupils (14.0%).
Table 1. Baseline characteristics of the subjects participated in this study
Female (n=57)

Male (n=71)

p-value*

MeanSDMeanSD

Number of remaining teeth

15.51

3.69

14.13

2.44

0.02

Number of sound teeth

15.25

3.51

13.90

2.49

0.02

Number of decayed teeth

0.05

0.23

0.04

0.20

0.78

Number of filled teeth

0.21

0.73

0.18

0.64

0.90

Number of missing teeth

0.00

0.00

0.00

0.00

1.00

DMF

0.26

0.74

0.23

0.70

0.51

Total streptococci (log10 CFU/mL)

6.56

0.30

6.420.84

0.28

Mutans streptococci (log10 CFU/mL)

3.25

2.00

3.29

1.83

0.66

Lactobacilli (log10 CFU/mL)

2.39

1.63

2.45

1.68

0.85

5 min stimulated saliva volume

5.49

2.90

6.88

3.79

0.06

Salivary pH

7.49

0.32

7.60

0.30

0.05

*Mann-Whitney U test
Table 2. The cross tabulation of incipient caries at baseline and after 3 months by QLF evaluation
After 3 months
Baseline

None

Right central
incisor (+)

Left central
incisor (+)

Both incisors (+)

TotalNone58 (45.3%)

9 (7.0%)9 (7.0%)4 (3.1%)80 (62.5%)

Right central incisor (+)

4 (3.1%)23 (18.0%)

0(0%)

0(0%)

27 (21.1%)

Left central incisor (+)

1 (0.8%)0(0%)

9 (7.0%)0(0%)

10 (7.8%)

Both incisor (+)

1 (0.8%)2 (1.6%)3 (2.3%)5 (3.9%)11 (8.6%)

Total

64 (50%)34 (26.5%)

21 (16.4%)

9 (7.0%)128 (100%)

The cross tabulation of with or without incipient caries at baseline and after 3 months were shown in Table 2.
Among 128 subjects, eleven pupils (8.6%) had incipient caries in both central incisors which examined, and 37pupils (28.9%) had incipient caries either in central incisors and 80 pupils (62.5%) had no incipient caries incentral incisors. Even DMF/DMFT was a low level, over one third of all pupils (in right 21.1% and in left 7.8%,

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Asian Pac J Dent 2013; 13: 5-10.

in both 8.6%: total 37.5%) had incipient caries detected by QLF.
After 3 months, second dental examination was performed.

Eleven pupils (8.6%) had disappeared theincipient caries in central incisors. However, newly incipient caries were appeared in residuary pupils. Finally,a half of all pupils (in right 26.5% and in left 16.4%, in both 7.0%) had incipient caries detected by QLF.
The distribution of the ΔF and ΔQ values were given in Table 3. The distributions of the both values werehighly skewed. The changes of the ΔF and ΔQ values when compared with the baseline and after 3 months, thechanges were statistically significant by Wilcoxon signed-rank test.
Table 3. The distribution of ΔF and ΔQ
ΔF0nΔQn197

0197

67

<0.510727110813210923810-2054-1011<20510<10Table 4. The relationship between incipient caries and salivary bacteria
Number ofbacteria

Cured incipientcaries(n=4)

No incipientcaries(n=60)

With incipientcaries(n=44)

Incidence ofincipient caries(n=22)

Total(n=130)p-value

Lactobacilli(log10 CFU/mL)

1.63 +/- 1.88

2.62 +/- 1.51

2.38 +/- 1.72

2.11 +/- 1.92

2.42 +/- 1.66

0.469

Mutansstreptococci(log10 CFU/mL)

1.60 +/- 1.94

3.65 +/- 1.64

3.18 +/- 1.89

2.80 +/- 2.31

3.28 +/- 1.890.072

Fig. 1.
abThree months variation of the incipient tooth caries. In some cases, drastically reduction of the ΔF and ΔQ values
were observed. a, QLF image photographed in a first dental examination. b, QLF image photographed in asecond dental examination.

The changes of the individual data were photographed and reserved by QLF computer apparatus. In somecases, drastically reduction of the ΔF and ΔQ values were observed (Fig. 1). Then the subjects classified into
four groups: the incipient caries cured in either teeth, no incipient caries in the baseline and after 3 months, with8

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Asian Pac J Dent 2013; 13: 5-10.

incipient caries at baseline in either teeth but not cured, and new incidence of incipient caries in either teeth. The
effects of the salivary levels of cariogenic bacteria for the changes of the ΔF values were investigated. Table 4
showed the relationship between incipient caries and salivary bacteria. Statistically significant effects of themutans streptococci and lactobacilli were not observed for the changes of ΔF value.

Discussion
The demineralization process of enamel starts when the acids are generated by biofilm bacteria. These acids
leaded to lower the biofilm pH to below pH 5.5. Then, begin the loss of calcium and phosphates from thesurface enamel, which creating incipient caries, so called a white spot lesion. According to the school dental
health criteria of Japanese Association of School Dentists, a visible white spot lesion is identified as caries forobservation (CO). The incipient caries (white spot or CO) is characterized by low calcium and phosphate
content and is the initial detectable lesion of enamel demineralization in the surface or subsurface of the tooth.
The incipient caries will progress into cavitation if the biofilm is not removed from the tooth surface for a longtime.11 The rate of carious progression is relatively slow, with enamel lesions confined to outer enamel taking
5-6 years to progress through to dentine.12 Another research shows that progression of caries from the surface ofthe enamel into dentine, sometimes taking up to four years, and that preventive efforts can reduce the carious
lesion progression.13
However, the demineralization process is reversible.

Saliva, a supersaturated solution of calcium andphosphates, plays a critical role in the process of remineralization. Remineralization of the incipient caries
effectively occurs when the biofilm is removed. After removal of biofilm, supersaturated saliva can be easily
access to the damaged enamel surface. Treatment of the incipient caries by professional mechanical toothcleaning14 or dental drug delivery system (3DS)15 and their subsequent remineralization has the potential tosignificantly advance noninvasive clinical management of dental caries.
In this study, we revealed the three months variation of the incipient caries using QLF. We found that 8.6% of
the subjects had disappeared the incipient caries within three months. Certainly, fluoride supplies strong andsignificant effects of the remineralization. In Japan, more than 85% of commercially available toothpaste
contained the fluoride.

Therefore, without special intervention, proper number of incipient caries will berepaired.
However, new incipient caries were also observed, and number of incipient caries were higher than before. In
order to control the incipient caries, three-monthly dental check-ups with QLF inspection will be preferable.

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Correspondence to:
Dr. Nobuhiro Hanada
Department of Translational Research, Tsurumi University School of Dental Medicine
2-1-3 Tsurumi, Tsurumi-ku, Yokohama 230-8501, Japan
Fax: +81-45-573-2473 E-mail: hanada-n@tsurumi-u.ac.jp
Accepted February 9, 2013.
Copyright ©2013 by the Asian Pacific Journal of Dentistry.
Online ISSN 2185-3487, Print ISSN 2185-3479

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