Urease activity as a risk factor for caries development in children during a three-year study period: a survival analysis approach.
ABSTRACT: Recent cross-sectional studies suggest that reduced ability to generate alkali via the urease pathway in dental plaque may be an important caries risk factor, but it has not been assessed prospectively.To evaluate the effect of plaque and saliva urease activity on the risk for developing new caries over a three-year period in children.A panel of 80 children, three to six years of age at recruitment, was followed prospectively for three years. Plaque urease activity, saliva urease activity and dental caries were measured every six months. Survival analysis methodology was used to evaluate the effect of urease on caries development during the study period adjusted for gender, age, baseline caries levels, sugar consumption, amount of plaque, and mutans streptococci levels.The risk for developing new caries increased in a dose-responsive manner with increasing levels of urease activity in saliva (adjusted HR(Q4 vs. Q1): 4.98; 95% CI: 1.33, 18.69) and with decreasing urease activity in plaque (adjusted HR(Q4 vs. Q1): 0.29; 95% CI: 0.11, 0.76). Multiple measurements of urease activity were conducted to overcome the variability of urease activity in this study. Baseline caries and mutans streptococci in saliva were also important predictors of caries risk.Increased urease activity in saliva can be an indicator of increased caries risk in children, whilst increased urease activity in plaque may be associated with reduced caries risk. The reproducibility of urease measurements must be improved before these findings can be further tested and clinically applied.
Project description:Bacterial urease activity in dental plaque and in saliva generates ammonia, which can increase the plaque pH and can protect acid-sensitive oral bacteria. Recent cross-sectional studies suggest that reduced ability to generate ammonia from urea in dental plaque can be an important caries risk factor. In spite of this proposed important clinical role, there is currently no information available regarding important clinical aspects of oral ureolysis in children.The objective of this study was to evaluate the distribution and pattern of urease activity in the dental plaque and in the saliva of children during a three-year period, and to examine the relationship of urease with some important caries risk factors.A longitudinal study was conducted with repeated measures over a three-year period on a panel of 80 children, aged 3-6 years at recruitment. The dynamics of change in urease activity were described and associated with clinical, biological, and behavioural caries risk factors.Urease activity in plaque showed a trend to remain stable during the study period and was negatively associated with sugar consumption (P<0.05). Urease activity in unstimulated saliva increased with age, and it was positively associated with the levels of mutans streptococci in saliva and with the educational level of the parents (P<0.05).The results of this study reveal interesting and complex interactions between oral urease activity and some important caries risk factors. Urease activity in saliva could be an indicator of mutans infection in children.
Project description:Early childhood caries (ECC) recurrence occurs in approximately 40% of treated cases within one year. The association of Streptococcus mutans and Candida albicans with the onset of ECC is well known. Also, S. mutans strains harboring collagen-binding proteins (Cbps) avidly bind to collagen-rich dentin and are linked to increased caries risk. Here, we investigated the presence of Cbp<sup>+</sup> S. mutans and C. albicans in saliva and dental plaque of children with varying caries statuses, and their salivary microbiome. In this cross-sectional study, 143 children who were caries-free (n = 73), treated for ECC with no signs of recurrence after 6 months (n = 45), or treated for ECC and experiencing recurrence within 6 months following treatment (n = 25) were enrolled. Co-infection with C. albicans and S. mutans, especially Cbp<sup>+</sup> S. mutans, was strongly associated with caries recurrence. Subjects of the recurrence group infected with Cbp<sup>+</sup> S. mutans showed a greater burden of Candida spp. and of Mutans streptococci in dentin than those infected with Cbp<sup>-</sup> strains. Salivary microbiome analysis revealed that Streptococcus parasanguinis was overrepresented in the caries recurrence group. Our findings indicate that Cbp<sup>+</sup> S. mutans and C. albicans are intimately associated with caries recurrence, contributing to the establishment of recalcitrant biofilms.
Project description:Saliva protects dental surfaces against cavities (i. e., dental caries), a highly prevalent infectious disease frequently associated with acidogenic <i>Streptococcus mutans</i>. Substantial <i>in vitro</i> evidence supports amylase, a major constituent of saliva, as either protective against caries or supporting caries. We therefore produced mice with targeted deletion of salivary amylase (Amy1) and determined the impact on caries in mice challenged with <i>S. mutans</i> and fed a diet rich in sucrose to promote caries. Total smooth surface and sulcal caries were 2.35-fold and 1.79-fold greater in knockout mice, respectively, plus caries severities were twofold or greater on sulcal and smooth surfaces. In <i>in vitro</i> experiments with samples of whole stimulated saliva, amylase expression did not affect the adherence of <i>S. mutans</i> to saliva-coated hydroxyapatite and slightly increased its aggregation in solution (i.e., oral clearance). Conversely, <i>S. mutans</i> in biofilms formed in saliva with 1% glucose displayed no differences when cultured on polystyrene, but on hydroxyapatite was 40% less with amylase expression, suggesting that recognition by <i>S. mutans</i> of amylase bound to hydroxyapatite suppresses growth. However, this effect was overshadowed <i>in vivo</i>, as the recoveries of <i>S. mutans</i> from dental plaque were similar between both groups of mice, suggesting that amylase expression helps decrease plaque acids from <i>S. mutans</i> that dissolve dental enamel. With amylase deletion, commensal streptococcal species increased from ~75 to 90% of the total oral microbiota, suggesting that amylase may promote higher plaque pH by supporting colonization by base-producing oral commensals. Importantly, collective results indicate that amylase may serve as a biomarker of caries risk.
Project description:Dental caries in pre-school children has significant public health and health disparity implications. To determine microbial risk markers for this infection, this study aimed to compare the microbiota of children with early childhood caries with that of caries-free children. Plaque samples from incisors, molars, and the tongue from 195 children attending pediatricians' offices were assayed by 74 DNA probes and by PCR to Streptococcus mutans. Caries-associated factors included visible plaque, child age, race, and snacking habits. Species were detected more frequently from tooth than tongue samples. Lactobacillus gasseri (p < 0.01), Lactobacillus fermentum, Lactobacillus vaginalis, and S. mutans with Streptococcus sobrinus (all p < 0.05) were positively associated with caries. By multifactorial analysis, the probiotic Lactobacillus acidophilus was negatively associated with caries. Prevotella nigrescens was the only species (p < 0.05) significantly associated with caries by the 'false discovery' rate. Analysis of the data suggests that selected Lactobacillus species, in addition to mutans streptococci, are risk markers for early childhood caries.
Project description:Chewing gum containing xylitol may help prevent caries by reducing levels of mutans streptococci (MS) and lactobacilli in saliva and plaque. Very little is known about other species which are possibly beneficial to oral health. In this study, we employed high-throughput sequencing of the 16S rRNA gene to profile microbial communities of saliva and plaque following short-term consumption of xylitol and sorbitol containing chewing gum. Participants (<i>n</i> = 30) underwent a washout period and were randomly assigned to one of two groups. Each group chewed either xylitol or sorbitol gum for three weeks, before undergoing a second four-week washout period after which they switched to the alternate gum for three weeks. Analysis of samples collected before and after each intervention identified distinct plaque and saliva microbial communities that altered dependent on the order in which gum treatments were given. Neither the xylitol nor sorbitol treatments significantly affected the bacterial composition of plaque. Lactobacilli were undetected and the number of <i>Streptococcus mutans</i> sequence reads was very low and unaffected by either xylitol or sorbitol. However, sorbitol affected several other streptococcal species in saliva including increasing the abundance of <i>S. cristatus</i>, an oral commensal shown to inhibit bacteria associated with chronic periodontitis.
Project description:Lactoferrin is one of a number of multifunctional proteins that are present in or on all mucosal surfaces throughout the body. Levels of lactoferrin are consistently elevated in inflammatory diseases such as arthritis, inflammatory bowel diseases, corneal disease, and periodontitis. Single-nucleotide polymorphisms (SNPs) in lactoferrin have been shown to be present in individuals susceptible to Escherichia coli-induced travelers' diarrhea and in tear fluid derived from virally associated corneal disease. Here, we review data showing a lactoferrin SNP in amino acid position 29 in the antimicrobial region of lactoferrin that acts against caries associated bacteria. This SNP was initially discovered in African American subjects with localized aggressive periodontitis (LAP) who had proximal bone loss but minimal proximal caries. Results were confirmed in a genetic association study of children from Brazil with this same SNP who showed a reduced level of caries. In vitro data indicate that lactoferrin from whole saliva derived from subjects with this SNP, recombinant human lactoferrin containing this SNP, or an 11-mer peptide designed for this SNP kills mutans streptococci associated with caries by >1 log. In contrast, the SNP has minimal effect on Gram-negative species associated with periodontitis. Moreover, periodontally healthy subjects homozygous for this lysine (K) SNP have lactoferrin in their saliva that kills mutans streptococci and have reduced proximal decay. The review summarizes data supporting the ecologic plaque hypothesis and suggests that a genetic variant in lactoferrin with K in position 29 when found in saliva and crevice fluid can influence community biofilm composition. We propose that, for caries, this SNP is ethnicity independent and protective by directly killing caries-provoking bacteria (reducing proximal decay). However, the clinical effect of this SNP in LAP is ethnicity dependent, destructive (increases LAP incidence), and complex with mechanisms still to be determined.
Project description:<h4>Background</h4>Xylitol is a naturally occurring sugar substitute that has been shown to reduce the level of mutans streptococci in plaque and saliva and to reduce tooth decay. It has been suggested that the degree of reduction is dependent on both the amount and the frequency of xylitol consumption. For xylitol to be successfully and cost-effectively used in public health prevention strategies dosing and frequency guidelines should be established. This study determined the reduction in mutans streptococci levels in plaque and unstimulated saliva to increasing frequency of xylitol gum use at a fixed total daily dose of 10.32 g over five weeks.<h4>Methods</h4>Participants (n = 132) were randomized to either active groups (10.32 g xylitol/day) or a placebo control (9.828 g sorbitol and 0.7 g maltitol/day). All groups chewed 12 pieces of gum per day. The control group chewed 4 times/day and active groups chewed xylitol gum at a frequency of 2 times/day, 3 times/day, or 4 times/day. The 12 gum pieces were evenly divided into the frequency assigned to each group. Plaque and unstimulated saliva samples were taken at baseline and five-weeks and were cultured on modified Mitis Salivarius agar for mutans streptococci enumeration.<h4>Results</h4>There were no significant differences in mutans streptococci level among the groups at baseline. At five-weeks, mutans streptococci levels in plaque and unstimulated saliva showed a linear reduction with increasing frequency of xylitol chewing gum use at the constant daily dose. Although the difference observed for the group that chewed xylitol 2 times/day was consistent with the linear model, the difference was not significant.<h4>Conclusion</h4>There was a linear reduction in mutans streptococci levels in plaque and saliva with increasing frequency of xylitol gum use at a constant daily dose. Reduction at a consumption frequency of 2 times per day was small and consistent with the linear-response line but was not statistically significant.
Project description:PURPOSE:Genotypic strains of mutans streptococci (MS) may vary in important virulence properties and be differentially affected by specific components of full-mouth caries restorative therapy. The purpose of this pilot study was to identify mutans streptococci strains that predominate following caries restorative therapy. METHODS:Plaque from 7 children with severe early childhood caries was collected before and following therapy. MS isolates (N=828) were subjected to polymerase chain reaction (PCR) and arbitrarily primed-PCR (AP-PCR) for assignment within MS strains. Determining the longitudinal changes in MS strain distribution over time within each patient required the isolation of larger numbers of isolates per patient, but from fewer patients. RESULTS:Up to 39 genotypic strains of Streptococcus mutans and Streptococcus sobrinus, and 7 genotypic strains of non-MS streptococci were identified by AP-PCR and 16S ribosomal rRNA gene sequencing. The number of MS strains isolated from each patient were 3 to 7 prior to treatment, diminishing to 1 to 2 dominant MS strains in most patients 6 months following therapy. CONCLUSIONS:Caries restorative therapy resulted in shifts of specific mutans streptococcus and non-mutans streptococcus strains. The implications are that caries restorative therapy affects the distribution of MS strains, and that well-accepted practices for caries prevention should be more closely examined for efficacy.
Project description:Antimicrobial peptides (AMPs) are among the repertoire of host innate immune defenses. In the oral cavity, several AMPs are present in saliva and have antimicrobial activities against oral bacteria, including Streptococcus mutans, a primary etiological agent of dental caries. In this study, we hypothesized that unique S. mutans strains, as determined by DNA fingerprinting from sixty 13-year-old subjects with or without experience of caries, would have different susceptibilities to ?-defensins-1-3 (HNP-1-3), ?-defensins-2-3 (HBD-2-3) and LL-37. The salivary levels of these peptides in subjects were also measured by enzyme-linked immunosorbent assays. We found that S. mutans strains from children with active caries showed greater resistance to salivary HNP-1-2, HBD-2-3 and LL-37 at varying concentrations than those from caries-free subjects. In addition, combinations of these peptides increased their antimicrobial activity against S. mutans either additively or synergistically. The salivary levels of these peptides were highly variable among subjects with no correlation to host caries experience. However, the levels of a number of these peptides in saliva appeared to be positively correlated within an individual. Our findings suggest that the relative ability of S. mutans to resist host salivary AMPs may be considered a potential virulence factor for this species such that S. mutans strains that are more resistant to these peptides may have an ecological advantage to preferentially colonize within dental plaque and increase the risk of dental caries.
Project description:<h4>Background</h4>Supragingival plaque and saliva are commonly used for microbiome analysis. Many epidemiological studies have identified deciduous teeth caries as a risk factor for caries development in first permanent molar (FPM); nevertheless, to the best of our knowledge, there are no reports on the effects of deciduous teeth caries on the microbiome of healthy FPM. Additionally, it remains unclear whether saliva can be used instead of supragingival plaque for caries microbial studies. Therefore, we aimed to elucidate this issue, and to characterize and compare the oral microbiome of healthy FPMs in children with different caries statuses and that from children with and without caries in a similar microhabitat, by PacBio sequencing. Currently, few studies have investigated the oral microbiome of children using this technique.<h4>Methods</h4>Thirty children (aged 7-9 years) with mixed dentition were enrolled; 15 had dental caries, and 15 did not. Supragingival plaques of deciduous molars and maxillary FPMs, and non-stimulating saliva samples were collected. DNA was extracted and the v1-v9 regions of 16S rRNA were amplified. Subsequently, PacBio sequencing and bioinformatic analyses were performed for microbiome identification.<h4>Results</h4>The microbial alpha diversity of the saliva samples was lower than that of the supragingival plaque (p < 0.05); however, no differences were detected between deciduous teeth and FPMs (p > 0.05). In addition, the alpha and beta diversity of children with and without caries was also similar (p > 0.05). Nonmetric multidimensional scaling and Adonis analyses indicated that the microbial structure of salivary and supragingival plaque samples differ (p < 0.05). Further analysis of deciduous teeth plaque showed that Streptococcus mutans, Propionibacterium acidifaciens, and Veillonella dispar were more abundant in children with caries than in those without (p < 0.05); while in FPMs plaque, Selenomonas noxia was more abundant in healthy children (p < 0.05). No differences in microorganisms abundance were found in the saliva subgroups (p > 0.05).<h4>Conclusion</h4>We have determined that supragingival plaque was the best candidate for studying carious microbiome. Furthermore, S. mutans, V. dispar, and P. acidifaciens were highly associated with deciduous teeth caries. S. noxia may be associated with the abiding health of FPM; however, this requires additional studies.