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RESEARCH REPORT |
1 University of Queensland, School of Dentistry, 200 Turbot Street, Brisbane, Queensland 4000, Australia;
2 Queensland Institute of Medical Research, Population & Clinical Sciences Division, Herston Rd., Brisbane, Queensland 4006, Australia;
3 Oral Biology Laboratory, University of Queensland, St Lucia, Brisbane, Queensland 4072, Australia; and
4 Mater Misericordiae Hospitals, Raymond Tce., South Brisbane, Queensland 4105, Australia;
*corresponding author, k.seow{at}uq.edu.au
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ABSTRACT |
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 TOP ABSTRACT INTRODUCTIONMATERIALS & METHODSRESULTSDISCUSSIONREFERENCES |
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KEY WORDS: Streptococcus mutans • pre-term • full-term • oral colonization
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INTRODUCTION |
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TOPABSTRACTINTRODUCTIONMATERIALS & METHODSRESULTSDISCUSSIONREFERENCES |
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MATERIALS & METHODS |
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TOPABSTRACTINTRODUCTIONMATERIALS & METHODSRESULTSDISCUSSIONREFERENCES |
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Data Collection
The subjects were recalled every 3 mos from birth until they were 24 mos of age. Interviews and dental examinations were conducted by one investigator (AKLW). Medical, dental, social, and feeding histories were recorded and updated at each recall visit, with the use of previously validated questionnaires (Seow et al., 1999). In addition, medical information provided by the mothers during interviews was verified by hospital records. Dental examinations of infants and their mothers were performed at the University Dental School, with the use of dental mirrors. Intra-examiner consistency for dental examination was established at over 95% (Wan et al., 2001b).
Collection of Microbiological Samples
We obtained microbiological samples from mothers and infants by swabbing the dorsum of the tongue and all surfaces of the teeth with sterile cotton tips. Each swab, which held 0.1 mL of saliva, was placed into a sterile vial containing phosphate-buffered saline, transported to the laboratory at 4°C, and processed within 4 hrs.
Isolation of Streptococcus mutans from Saliva
The methods used for S. mutans isolation and enumeration have been published previously (Wan et al., 2001a,b, 2002). In brief, after samples were vortexed for 30 sec to disperse bacteria, 50-µL aliquots of ten-fold dilutions were plated onto S. mutans-selective tryptone-yeast-cysteine-sucrose-bacitracin agar (TYSCB; Microdiagnostic, Brisbane, Australia). Plates in triplicate were incubated at 37°C under anaerobic conditions for 72 hrs. Colonies were enumerated by means of a colony-counter, and the mean colony-forming units/mL saliva (CFUs/mL) were calculated for S. mutans. Control plates with known concentrations of S. mutans (NCTC 10449) were incubated with the sample plates. Bacterial identification was verified on random colonies by Gram stains and biochemical analyses (Rapid Strep ID32 API, bioMerieux Vitek, Marcy-l’Etoile, France). Colonization was considered positive when 2 consecutive saliva samples showed S. mutans.
Statistical Analysis
All statistical analyses were performed with SPSS version 10.0.1. Non-parametric statistical analyses were performed with Chi-square, Mann-Whitney, Kruskal-Wallis, and Friedman tests and Spearman’s rank-order correlation. We used logistic regression to determine independent predictors for S. mutans colonization.
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RESULTS |
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TOPABSTRACTINTRODUCTIONMATERIALS & METHODSRESULTSDISCUSSIONREFERENCES |
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). The overall mean birthweight and gestational age of pre-term infants were 1.8 + 0.7 kg and 32.7 + 3.6 wks compared with 3.6 + 0.5 kg and 39.8 + 1.0 wks in full-term infants (p < 0.01).
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shows the overall prevalence of S. mutans in the study cohort of 312 infants. By the end of the study at age 24 mos, 79% of the 312 infants were colonized by S. mutans. The cumulative prevalence of S. mutans colonization in all children was 5%, 18%, 49%, 53%, 62%, 68%, 70%, 74.5%, and 79% at 0, 6, 9, 12, 15, 18, 21, and 24 mos of age, respectively. As reported in our previous paper, the overall prevalence of S. mutans colonization in infants without teeth was 64% (153/238) (Wan et al., 2001b). The prevalence increased to 84% (93/111) after the eruption of teeth in this part of the study.
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, the overall prevalence of S. mutans colonization in all children after tooth eruption was 1%, 12%, 37%, 54%, 58%, 71%, and 84% at 6, 9, 12, 15, 18, 21, and 24 mos of age, respectively. Cumulative colonization is represented by these prevalence rates. At 12 mos of age, the prevalence of S. mutans infection was similar between pre-term and full-term infants (p > 0.1) (Fig. 2). Before 12 mos of age, there was a higher prevalence in full-term compared with pre-term children, and after 12 mos of age, a higher prevalence in the pre-term infants, but the difference was not statistically significant (p > 0.1) (Fig. 2). In contrast, at 24 mos of age, the prevalence in pre-term infants was significantly higher, at 94%, compared with that in full-term infants, at 79% (p < 0.042; OR = 4.4) (95%CI = 1.0-20.3) (Fig. 2). The pre-term infants had 4.4 times (95%CI = 1.0-20.3) higher odds of being colonized by S. mutans compared with full-term infants.
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The differences in colonization ages between pre-term and full-term infants disappeared when comparisons with pre-term infants were made by corrected age rather than by chronological age. The mean corrected age of colonization for pre-term infants was 15.0 + 4.7 mos vs. 15.4 + 5.3 mos in full-term infants (p > 0.1), and the median corrected age of colonization in pre-term infants was 14.3 mos vs. 15.0 mos in full-term infants (p > 0.1).
Mean Levels of S. mutans in Saliva
At all ages when colonization was first detected, the mean S. mutans levels in pre-term infants (707 + 1400 CFU/mL) were similar to those in full-term infants (716 + 1695 CFU/mL) (p > 0.1). However, the levels in both groups of children increased significantly with increasing numbers of erupted teeth (p < 0.001).
Dental Caries
By 24 mos of age, 9% of all infants (four pre-term, four full-term) colonized with S. mutans showed dental caries. In contrast, none of the infants without the bacteria had dental caries (Table
). The mean age of colonization in infants who developed dental caries was higher (19.7 + 5.4 mos) compared with that of those who remained caries-free at 24 mos (15.3 + 5.0 mos, p < 0.03).
Factors Associated with S. mutans Colonization in Pre-term Infants
Two major factors were identified in pre-term children which contributed to their increased predisposition to S. mutans colonization. These include a 26.2 times increased risk of enamel hypoplasia (95%CI = 7.8-88.1), and a 7.9 times (95%CI = 3.4-18.7) higher risk of frequent sugar consumption (> 3 times/day) in pre-term compared with full-term infants. Other factors associated with S. mutans colonization were generally similar in all infants, and are listed below.
General Infant Factors Associated with S. mutans Colonization
The risk factors associated with S. mutans colonization are summarized in Web appendix (www.dentalresearch.org).
Demographic Factors
S. mutans colonization was higher in infants from families of low total annual income, and infants with mothers who have only a primary school education, at 9 mos (p < 0.01), 12 mos (p < 0.03), and 15 mos of age (p < 0.05). However, at 24 mos of age, this trend was reversed, and the S. mutans colonization rate was higher in infants of high socio-economic status (p < 0.03).
At 9 Mos of Age
The most significant factors associated with colonization of S. mutans at 9 mos of age were being habitually kissed on the lips (OR = 6.4, 95%CI = 3.0-13.4) and having food pre-tasted (OR = 6.4, 95%CI = 2.9-14.5). Other significant factors were foods or utensils shared with other individuals (OR = 4.6, 95%CI = 2.3-9.5) and total sugar exposures of > 3/day (OR = 4.6, 95%CI = 2.3-9.2).
At 12 Mos of Age
At 12 mos of age, the most significant factors were: irregular toothbrushing habits of < 1 day (OR = 2.1, 95%CI = 1.8-2.6), spending > 10 hrs per week at care facilities (OR = 3.7, 95%CI = 1.2-7.3), presence of enamel hypoplasia (OR = 5.0, 95%CI = 2.8-9.1), snacking > 3 times/day (OR = 5.6, 95%CI = 2.8-11.1), sharing food with others (OR = 5.7, 95%CI = 2.4-13.4), sugar exposure from fluids or solids > 3 times/day (OR = 6.1, 95%CI = 2.7-13.5), and formula feeding on-demand (OR = 8.9, 95%CI = 1.9-41.6).
At 15 Mos of Age
Sharing utensils with others (OR = 15.5, 95%CI = 4.1-58.4), plaque covering > 50% of teeth (OR = 8.1, 95%CI = 1.7-38.2), use of child care facilities before 12 mos of age (OR = 6.0, 95%CI = 1.4-26.2), sweetened fluids taken to bed (OR = 5.2, 95%CI = 1.9-14.3), presence of enamel hypoplasia (OR = 4.8, 95%CI = 2.7-8.7), and drinking only from bottles (OR = 4.8, 95%CI = 1.6-14.7) were the most significant risks associated with S. mutans colonization at 15 mos of age.
At 18 Mos of Age
Regular consumption of sticky snacks (OR = 10.4, 95%CI = 3.7-29.2), snacking > 3/day (OR = 9.3, 95%CI = 3.1-28.1), total sugar exposure > 3/day (OR = 6.2, 95%CI = 1.2-5.3), taking sweetened fluids to bed (OR = 4.4, 95%CI = 1.5-12.4), use of care facilities before 12 mos of age (OR = 3.9, 95%CI = 2.1-7.0), and sharing foods with others (OR = 3.1, 95%CI = 1.2-8.0) were the most significant risk factors associated with S. mutans colonization at 18 mos of age.
At 21 Mos of Age
At 21 mos of age, significant factors associated with S. mutans colonization were total sugar exposure of > 3/day (OR = 26.0, 95%CI = 6.1-111.5), sharing food or utensils (OR = 15.2, 95%CI = 3.9-58.7), feeding at night and sleeping beside mother (both OR = 7.3, 95%CI = 2.7-20.1), and snacking > 3/day (OR = 10.3, 95%CI = 2.1-50.3).
At 24 Mos of Age
The most significant factors at 24 mos of age were feeding at night (OR = 62.5, 95%CI = 7.8-94.9), drinking only from bottles (OR = 14.1, 95%CI = 4.4-45.3), on-demand feeding (OR = 11.6, 95%CI = 3.1-43.3), sleeping beside mother (OR = 7.8, 95%CI = 2.9-20.9), and total sugar exposure from > 3/day (OR = 3.6, 95%CI = 1.3-10.4).
Maternal Factors Associated with Infant Streptococcus mutans Colonization
Presence of S. mutans at levels > 105 CFU/mL (OR = 2.1-8.5, 95%CI = 1.2-27.6), snacking > 3/day (OR = 3.3-30.3, 95%CI = 1.2-233.2), plaque covering > 50% of dentition (OR = 3.8-18.8, 95%CI = 0.9-84.0), and periodontal pocketing/CPI > 2 (OR = 1.6-5.3, 95%CI = 0.3-30.1) were identified as important maternal factors associated with S. mutans colonization in infants aged 9-24 mos.
Factors Associated with Non-colonization of S. mutans in Dentate Infants
Non-colonization of S. mutans was associated with parent-assisted toothbrushing (OR = 1.3-2.7, 95%CI = 1.1-5.6) and multiple courses of antibiotics taken between 13 and 24 mos of age (OR = 1.4-2.6, 95%CI = 1.1-11.6).
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DISCUSSION |
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TOPABSTRACTINTRODUCTIONMATERIALS & METHODSRESULTSDISCUSSIONREFERENCES |
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In contrast to the concept of a discrete "window of infectivity" for the timing of S. mutans colonization, we found instead a steady increase in the rate of infection as the infants’ ages and numbers of erupted teeth increased. The cumulative prevalence of S. mutans colonization in all infants was 5%, 18%, 49%, 53%, 62%, 68%, 70%, 74.5%, and 79% at 0, 6, 9, 12, 15, 18, 21, and 24 mos of age, respectively. Colonization after tooth eruption showed the same trend: At 6 mos of age, the infection rate was approximately only 1%, rising to 12% at 9 mos, 37% at 12 mos, 54% at 15 mos, 58% at 18 mos, 71% at 21 mos, and 84% at 24 mos of age in those infants who were colonized by S. mutans after tooth eruption. Compared with previous studies, colonization rates at the various ages in this study are generally higher than those reported by other investigators (Caufield et al., 1993; Karn et al., 1998). The differences are likely to be related to cohort population differences, or to the relatively large numbers of subjects in the present study. In addition, increased recovery potential of S. mutans-selective TYCSB agar used in the present investigation compared with other media for isolation of S. mutans (Wan et al., 2002) may have increased the culture sensitivity of the samples in the present study.
As is the case before tooth eruption, S. mutans colonization after tooth eruption is influenced by both maternal and infant factors. It is now well-recognized that the mother is usually the primary source of S. mutans for infection of her child (Kohler and Bratthall, 1978; Li and Caufield, 1995), and poor maternal oral hygiene and dietary habits increase the likelihood of transmission of the infection from mother to child. Hence, in this study, greater numbers of mothers of infected infants showed higher S. mutans levels, less frequent toothbrushing, greater plaque levels, and higher daily frequencies of snacking and sugar exposures compared with mothers who had uninfected infants.
Furthermore, since successful colonization by S. mutans probably requires repeated exposures, our data showed that child-rearing habits which facilitate saliva transfer from adults to the child, such as sharing of food and utensils, and habits which involve close contact, such as breast feeding and sleeping beside the mother, were also significantly associated with colonization of S. mutans.
Several infant factors contribute significantly to the colonization of S. mutans. First, although S. mutans may colonize the mouth before tooth eruption, the emergence of teeth increases the non-shedding surfaces for adherence of S. mutans. Thus, with tooth eruption, the colonization rate of the infants increases as their ages increase. Second, infant dietary and oral hygiene habits also facilitate the colonization of S. mutans. As in our previous studies on predentate infants, frequent exposure to sugars, especially from snacking and sweetened pacifiers, correlated significantly with S. mutans colonization (Wan et al., 2001b). In the case of pre-term infants, this habit is worsened by the increased sugar content in diets that are recommended to boost calorie intake (Sauve and Geggie, 1991).
The importance of regular toothbrushing in the prevention of S. mutans infection in infants is also demonstrated in this study. Our data thus support and extend the work of others (Habibian et al., 2002), who reported that infants who have their teeth brushed by 12 mos of age are less likely to harbor S. mutans compared with those who had not commenced brushing.
Previous studies suggested that the earlier the colonization of S. mutans, the higher the caries risk (Kohler et al., 1988). In the present investigation, we found that in the eight infants who developed caries, S. mutans was first detected at a median age of 18 mos. Although these results relating age of colonization to caries development are preliminary, they suggest that age of colonization on its own may not be the most important aspect in caries development, since other factors, such as sugar consumption and oral hygiene, are also likely to play significant roles in determining caries risk (Loesche, 1986).
In conclusion, data from our longitudinal studies have established the timing of S. mutans colonization from birth to 24 mos of age, and provided insight into infant and maternal factors which facilitate the colonization. These data would be highly relevant in the institution of preventive strategies for early childhood caries.
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ACKNOWLEDGMENTS |
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FOOTNOTES |
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Received October 14, 2002; Last revision March 31, 2003; Accepted April 4, 2003
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REFERENCES |
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TOPABSTRACTINTRODUCTIONMATERIALS & METHODSRESULTSDISCUSSIONREFERENCES |
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