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Dental Caries is a kind of chronic oral disease that greatly threaten human being’s health. Though dentists and researchers struggled for decades to combat this oral disease, the incidence and prevalence of dental caries remain quite high. Therefore, improving the disease management is a key issue for the whole population and life cycle management of dental caries. So clinical difficulty assessment system of caries prevention and management is established based on dental caries diagnosis and classification. Dentists should perform oral examination and establish dental records at each visit. When treatment plan is made on the base of caries risk assessment and carious lesion activity, we need to work out patient‑centered and personalized treatment planning to regain oral microecological balance, to control caries progression and to restore the structure and function of the carious teeth. And the follow-up visits are made based on personalized caries management. This expert consensus mainly discusses caries risk assessment, caries treatment difficulty assessment and dental caries treatment plan, which are the most important parts of caries management in the whole life cycle.

We previously demonstrated that recombinant protein PAc could be administered as an anticaries vaccine. However, the relatively weak immunogenicity of PAc limits its application. In the present study, we investigated the effect of two adjuvant combinations of chitosan plus Pam(3)CSK(4) (chitosan-Pam(3)CSK(4)) and of chitosan plus monophosphoryl lipid A (chitosan-MPL) in the immune responses to the PAc protein in vivo and in vitro. PAc-chitosan-Pam(3)CSK(4) or PAc-chitosan-MPL promoted significantly higher PAc-specific antibody titers in serum and saliva, inhibited Streptococcus mutans colonization onto the tooth surfaces, and endowed better protection effect with significantly less caries activities than PAc alone. Chitosan-Pam(3)CSK(4) and chitosan-MPL showed no statistically significant differences. In conclusion, our study demonstrated that the chitosan-Pam(3)CSK(4) and chitosan-MPL combinations are promising for anticaries vaccine development.

Objective To investigate whether the recombinant FimH-S.T protein could modulate immune response to anti-caries vaccine in vitro and in vivo. Design Recombinant FimH protein derived from Salmonella was constructed and purified. The expression of dendritic cell maturation markers and cytokines release were performed by flow cytometry, Real-time PCR and ELISA. In addition, BALB/c mice were administered with anti-caries PAc vaccine plus FimH-S.T, antibody responses were evaluated by ELISA. Splenocytes of immunized mice were detected for their proliferative ability in response to in vitro retreatment with PAc antigen by flow cytometry. Caries protection against dental caries formation was also investigated. Results The purified FimH-S.T induced phenotypic maturation of DC2.4 by up-regulating the expression of costimulatory molecules and MHC II, provoked the production and secretion of cytokines via TLR4-dependent signaling pathway in vitro. Furthermore, the mice immunized with the mixture of FimH-S.T and PAc significantly enhanced the PAc-specific antibodies in the serum along with saliva and promoted splenocyte proliferation. Our results also confirmed that PAc+FimH-S.T decreased the caries lesions formation which provided high protective efficacy against dental caries. Conclusion Our study demonstrates that recombinant FimH-S.T could enhance specific IgA responses and protection of anti-caries vaccine, possessing mucosal adjuvant ability by activating DC2.4 via TLR4 signaling pathway.

Aims To investigate the effects of the ginsenoside Rh2 on monospecies and multispecies cariogenic biofilms and explore the mechanism of the antibiofilm effect of Rh2 in vitro. Methods and results Streptococcus mutans, Streptococcus sobrinus and Streptococcus sanguinis were chosen to form the monospecies or multispecies biofilms. Crystal violet staining and laser scanning confocal microscopy were used to observe the effect of Rh2 on biofilms in vitro. Cytotoxicity was examined by the Cell Counting Kit-8. The effects of Rh2 on bacterial membranes were observed via transmission electron microscopy (TEM). The isobaric tags for relative and absolute quantification (iTRAQ) method were used to profile the common differentially expressed proteins. Gene expression was analysed by reverse transcription quantitative polymerase chain reaction. In general, the treatment of cariogenic biofilms with Rh2 significantly decreased biomass accumulation by inhibiting bacterial growth and extracellular polysaccharide synthesis without any cytotoxic effects. TEM imaging showed that Rh2 could disrupt the cell membranes of these bacteria. The iTRAQ results indicated that the levels of mannose-specific IIC/D and acetaldehyde/alcohol dehydrogenase were substantially down-regulated, while the mRNA expression of the corresponding genes were significantly changed. Conclusions Our data revealed a potential application for Rh2 in the protection against dental caries via the inhibition of cariogenic biofilms. Significance and impact of the study This study describes the first application of a ginsenoside against multispecies cariogenic biofilms. Rh2 may serve as an alternative agent to prevent dental caries by effectively modulating the pathogenic potentials of oral biofilms.

The purpose of this study was to determine the relationships among early childhood caries (ECC), root caries (RC), the quantity of Streptococcus mutans in saliva, and the concentrations of total and specific secretory IgA (sIgA). Saliva samples were collected from 70 children, 3-4 yr of age, with and without ECC, and from 43 adults, ≥60 yr of age, with and without RC. The decayed, missing, and filled teeth (dmft) and decayed, missing, and filled surfaces (dmfs) scores of each child, and the root decayed and filled teeth (RDFT) and root decayed and filled surfaces (RDFS) scores of each elderly subject, were determined. The S. mutans levels, total sIgA, and specific sIgA against two virulence antigens of S. mutans in saliva were analysed using quantitative real-time PCR (qPCR) and ELISAs. The quantity of S. mutans was significantly higher in caries-positive subjects within the two populations than in the caries-free subjects; and a positive correlation was found between the quantity of S. mutans and the dmft, dmfs, RDFT, and RDFS scores. In addition, the salivary total sIgA was significantly higher in children with severe early childhood caries (SECC) and in the elderly subjects with RC. Moreover, although the S. mutans level was significantly higher, the concentrations of specific sIgA against S. mutans antigens were significantly lower in samples from elderly subjects than in samples from children. These results support the concept that S. mutans is positively associated with ECC and RC. Furthermore, the levels of S. mutans-specific antibodies in saliva are too low to prevent infection with cariogenic bacteria and to inhibit development of ECC and RC.

The aim of this study was to test the hypothesis that the conventional restorative treatment (CRT) and the atraumatic restorative treatment (ART) protocols, in comparison with the ultra-conservative treatment (UCT) protocol, would increase the quality of life of children over a period of 1 year. Cavitated primary molars of 302 children 6–7 years of age were treated according to the CRT, ART and UCT protocols at the school compound. Children’s parents completed the Brazilian version of the Early Childhood Oral Health Impact Scale (B-ECOHIS) at baseline and one year later. Paired t-test, Chi-square test and anova were applied in analysing the data. Questionnaires from 277 and 160 children were collected at baseline and after 1 year, respectively. A statistically significant difference in B-ECOHIS scores over the 1-year period was found for domains ‘child symptoms’ (P = 0.03) and ‘child psychology’ (P = 0.02). Treatment protocols did not statistically significantly influence the changes in B-ECOHIS scores over the 1-year period (P = 0.78). It can be concluded that the UCT protocol was as good as the two restorative protocols. All treatment protocols were effective in reducing children’s experience of pain, their sleeping problems and their irritability and/or frustration levels over the 1-year period.

Anticaries protein vaccines that induce a mucosal immune response are not effective. Therefore, development of effective and convenient anticaries vaccines is a priority of dental research. Here we generated self-assembling nanoparticles by linking the glucan-binding region of Streptococcus mutans glucosyltransferase (GLU) to the N-terminal domain of ferritin to determine whether these novel nanoparticles enhanced the immunogenicity of an anticaries protein vaccine against GLU in rodents. We constructed the expression plasmid pET28a-GLU-FTH and purified the proteins from bacteria using size-exclusion chromatography. BALB/c mice were used to evaluate the ability of GLU-ferritin (GLU-FTH) nanoparticles to induce GLU-specific mucosal and systemic responses. The protective efficiency of GLU-FTH nanoparticles was compared with that of GLU alone or a mixture of GLU and poly(I:C) after administering an intranasal infusion to Wistar rats. The phagocytosis and maturation of dendritic cells (DCs) exposed in vitro to the nanoparticles were assessed using flow cytometry. The GLU-FTH nanoparticle vaccine elicited significantly higher levels of GLU-specific antibodies compared with GLU or a mixture of GLU and poly(I:C). Immunization with GLU-FTH achieved lower caries scores compared with those of the other vaccines. Administration of GLU-FTH nanoparticles enhanced phagocytosis by DCs and their maturation. Thus, self-assembling GLU-FTH is a highly effective anticaries mucosal vaccine that enhanced antibody production and inhibited S. mutans infection in rodents.

To investigate the effects of anti-caries DNA vaccine-induced salivary secretory immunoglobulin A (S-IgA) antibodies on Streptococcus mutans (S. mutans) adherence and biofilms formation in vitro. Adult female Wistar rats were intranasally immunized with the anti-caries DNA vaccine pGJA-P/VAX. Their saliva samples were collected at different times after the immunization, and S-IgA antibody level in the saliva and its inhibition on S. mutans adherence were examined. The effects of S-IgA in the saliva with the strongest inhibitory effects were examined at 3 different stages, ie acquired pellicles, biofilm formation and production of mature biofilms. The number of viable bacteria and depth of the biofilm at 16 h in each stage were determined using counting colony forming units and using a confocal laser scanning microscopy (CLSM). The participation of S-IgA in acquired pellicles and its aggregation with S. mutans were also observed under CLSM. The S-IgA titer in saliva reached its peak and exhibited the strongest inhibition on S. mutans adhesion at 10 weeks after the immunization. The colonies and depth of the biofilm in the saliva-pretreated group were 41.79% and 41.02%, respectively, less than the control group. The colonies and depth of the biofilm in the co-culture group were 27.4% and 22.81% less than the control group. The assembly of S. mutans and S-IgA was observed under CLSM after co-cultivation. In the mature-stage biofilm, no differences were observed between the different groups. These results demonstrate that the anti-caries DNA vaccine induces the production of specific S-IgA antibodies that may prevent dental caries by inhibiting the initial adherence of S. mutans onto tooth surfaces, thereby reducing the accumulation of S. mutans on the acquired pellicles.

We previously demonstrated that an anti-caries DNA vaccine intranasally administered with recombinant flagellin protein as a mucosal adjuvant enhanced salivary IgA response and conferred better protection against caries. However, the relatively weak immunogenicity of DNA vaccines and the necessity for a large quantity of antigens remain significant challenges. Here, we fused the flagellin derived from E. coli (KF) and target antigen PAc containing the A-P fragment of PAc from S. mutans (rPAc) to produce a single recombinant protein (KF-rPAc). The abilities of KF-rPAc to induce rPAc-specific mucosal and systemic responses and protective efficiency against caries following intranasal immunization were compared with those of rPAc alone or a mixture of rPAc and KF (KF + rPAc) in rats. Results showed that KF-rPAc promoted significantly higher rPAc-specific antibodies in serum as well as in saliva than did an equivalent dose of rPAc alone or a mixture of KF + rPAc. Intranasal immunization of 8.5 µg KF-rPAc could achieve 64.2% reduction of dental caries in rats. In conclusion, our study demonstrated that flagellin and PAc fusion strategy is promising for anti-caries vaccine development, and KF-rPAc could be used as an anti-caries mucosal vaccine.

Item does not contain fulltext OBJECTIVES: The aim of the present study was to investigate the caries-preventive effect of sealants produced with a high-viscosity glass-ionomer with an elevated powder-liquid ratio (ART), of having energy added to this glass-ionomer, and that of glass-carbomer, in comparison to that of resin composite sealants. METHODS: The randomized controlled trial covered 407 children, with a mean age of 8 years. At a school compound three dentists placed sealants in pits and fissures of high caries-risk children. Evaluation by two independent evaluators was conducted after 0.5, 1 and 2 years. The Kaplan-Meier survival method, ANOVA and t-test were used in analyzing the data. RESULTS: 1352 first permanent molars were sealed. 6.6% of children and 6.8% of sealants dropped out within 2 years. 27 re-exposed pits and fissures, 20 in occlusal and 7 in smooth surfaces, in 25 children, developed a dentin carious lesion. The cumulative survival of dentin carious lesion-free pits and fissures in the glass-carbomer sealant group was statistically significantly lower (97.4%) than those in the high-viscosity glass-ionomer with energy supplied (99%) and the resin-composite (98.9%) sealant groups. There was no statistically significant difference in the cumulative survival of dentin carious lesion-free pits and fissures, between the high-viscosity glass-ionomer with (99%) and without (98.3%) energy supplied sealant groups, after 2 years. SIGNIFICANCE: The survival of dentin carious lesion-free pits and fissures was high in all sealant types. More dentin carious lesions were observed in the glass-carbomer sealant group. 01 mei 2012

We and others have shown that anti-caries DNA vaccines, including pGJA-P/VAX, are promising for preventing dental caries. However, challenges remain because of the low immunogenicity of DNA vaccines. In this study, we used recombinant flagellin protein derived from Salmonella (FliC) as a mucosal adjuvant for anti-caries DNA vaccine (pGJA-P/VAX) and analyzed the effects of FliC protein on the serum PAc-specific IgG and saliva PAc-specific IgA antibody responses, the colonization of Streptococcus mutans ( S. mutans) on rat teeth, and the formation of caries lesions. Our results showed that FliC promoted the production of PAc-specific IgG in serum and secretory IgA (S-IgA) in saliva of rats by intranasal immunization with pGJA-P/VAX plus FliC. Furthermore, we found that enhanced PAc-specific IgA responses in saliva were associated with the inhibition of S. mutans colonization of tooth surfaces and endowed better protection with significant fewer caries lesions. In conclusion, our study demonstrates that recombinant FliC could enhance specific IgA responses in saliva and protective ability of pGJA-P/VAX, providing an effective mucosal adjuvant candidate for intranasal immunization of an anti-caries DNA vaccine.

Background The present study aimed to enhance the specific anti-caries immunity induced by DNA prime-protein boost strategy for an A-P fragment of a cell-surface protein antigen of Streptococcus mutans (PAc). Methods BALB/c mice were immunized with DNA prime-protein boost, DNA–DNA or protein–protein regimens by the intranasal route, using combinations of plasmid vector (pCIA-P) that express PAc protein and a pure secretec recombinant PAc protein (rPAc). Then, a gnotobiotic mouse model was constructed 2 weeks after the last immunization, and specific immune responses in vivo and their protection against dental caries were observed. Results The present study revealed stronger antibody responses in the DNA prime-protein boost group compared to those elicited by either DNA–DNA vaccination or protein–protein vaccination. In particular, PAc-specific antibody concentrations were improved significantly after boosting the pCIA-P DNA-primed mice with rPAc. Moreover, protection against S. mutans challenge was obtained in the mice treated with the DNA prime-protein boost vaccination, as demonstrated by a significant reduction in S. mutans colonization compared to control mice and animals immunized with the DNA–DNA vaccination or protein–protein vaccination. Conclusions The results obtained in the present study suggest that the intranasal DNA prime-protein boost vaccination regimen is a novel strategy for the practical application of DNA vaccine against dental caries. Copyright © 2009 John Wiley & Sons, Ltd.

Our previously prophylactic studies have proved that the anti-caries DNA vaccine pGJA-P/VAX could generate effective immune response by intramuscular (i.m.) and intranasal (i.n.) administration in rats without caries. In the present, we determine whether it also could produce efficacy in rats with caries. By immunized with pGJA-P/VAX, rats were elicited both significantly higher anti-Streptococcus mutans serum IgG and salivary SIgA responses, compared to those with pVAX1. Correspondingly, rats immunized with pGJA-P/VAX via i.n. displayed significantly fewer enamel, dentinal lesions compared to those with pVAX1 via i.n. However, there was no significant difference in dental caries lesions between pGJA-P/VAX (i.m.) and pVAX1 (i.m.). These findings suggest that DNA vaccination via i.n., with bupivacaine delivery system, could be a promising alternatives for slowing down caries development in rat models.

Item does not contain fulltext This study, conducted from a government program perspective, compared the incremental cost-effectiveness of oral health interventions, in particular their delivery to underserved populations in whom dental sealants constitute an important, high-yielding complement to toothbrushing in dental-caries prevention. The study data concern the relative cost-effectiveness of three sealant materials in four approaches to prevent cavitated dentine carious lesions in permanent molars in a community intervention trial among school-age children in Wuhan, China. The four approaches were high-viscosity glass-ionomer cement without heat application (HVGIC); high-viscosity glass-ionomer cement with heat application [light-emitting diode (LED) thermocured HVGIC]; glass-carbomer; and composite resin. The costs studied were: cost of sealing permanent molars; adverse event costs for restoring cavitated dentine carious lesions developing within 4 yr in study data; and projections of 1,000 sealants per group. Preventing one more cavitated dentine carious lesion cost US$105 for the study data when comparing HVGIC (n = 405) with composite resin (n = 396) and US$59 per 1,000 sealants in the projections; LED thermocured HVGIC compared with composite resin cost US$115 for one more cavitated lesion and US$52 per 1,000 sealants, respectively. Although more expensive than composite resin, LED thermocured HVGIC was identified as the most cost-effective among the sealant materials studied. Ease of application, minimal technical and infrastructure requirements, and cost-effectiveness make glass-ionomers a practicable option for governments making decisions under economic constraints.

Contains fulltext : 53242.pdf (Publisher’s version ) (Closed access) OBJECTIVES: This study aimed to determine whether salivary mutans streptococci (MS) counts in Chinese children had any value in the prediction of new caries in the permanent dentition in the age interval of 6.5-8.5 years. METHODS: Four hundred and thirty-three 6-7-year-old children participated in this 2-year longitudinal study. Salivary MS counts were obtained at baseline with the spatula method. Dentin and enamel caries was recorded at baseline and at the follow-up, 2 years later. Spearman's correlation coefficients were computed. Past caries experience variables at baseline were entered into a multiple regression model with caries increment of the permanent dentition as dependent variable. MS counts were subsequently entered into the model to assess the additional caries predictive value. RESULTS: Spearman's correlation coefficient for MS counts and caries experience of the primary dentition at baseline was 0.48 and for MS counts and caries increment in the permanent dentition 0.12. In the multiple regression model caries increment of the permanent dentition had an adjusted R2 of 0.17 when the past caries experience variables were entered. Adding MS counts in the regression model did not change the R2 value. CONCLUSIONS: Salivary MS counts did not add any caries predictive value when past caries experience variables were used as caries predictors in this Chinese child population.

The levels of Streptococcus (S.) mutans infections in saliva were evaluated and a comparison for specific antibody levels among children with different levels of S. mutans infection was made. The promising epitopic regions of antigen AgI/II (PAc) and glucosyltransferase (GTF) for potential vaccine targets related to S. mutans adherence were screened. A total of 94 children aged 3–4 years were randomly selected, including 53 caries-negative and 41 caries-positive children. The values of S. mutans and those of salivary total secretory immunoglobulin A (sIgA), anti-PAc and anti-Glucan binding domain (anti-GLU) were compared to determine the correlation among them. It was found the level of s-IgA against specific antigens did not increase with increasing severity of S. mutans infection, and the complete amino acid sequence of PAc and GTFB was analyzed using the DNAStar Protean system for developing specific anti-caries vaccines related to S. mutans adherence. A significantly positive correlation between the amount of S. mutans and children decayed, missing, and filled teeth index was observed. No significant difference was detected in specific sIgA against PAc or GLU between any two groups. No significant correlation was found between such specific sIgA and caries index. A total of 16 peptides from PAc as well as 13 peptides from GTFB were chosen for further investigation. S. mutans colonization contributed to early children caries as an important etiological factor. The level of sIgA against specific antigens did not increase with increasing severity of S. mutans infection in children. The epitopes of PAc and GTF have been screened to develop the peptide-based or protein-based anti-caries vaccines.

Aims The aim of this study was to identify new small molecules that can inhibit Streptococcus mutans biofilms by in vitro and in vivo model. Methods and results We evaluated the effect of a small molecule 2-amino-imidazole/triazole conjugate (2-AI/T) on the formation of Strep. mutans biofilms by culturing in 96-well plates. Toxicity was assessed through cell culture and intragastrically administering to mice. The anti-biofilm and anti-caries effects were investigated in vivo. The inhibitive mechanism was detected by isobaric tag for relative and absolute quantification (itraq) and RT-QPCR. In vitro and in vivo study revealed that 2-AI/T significantly inhibited biofilm formation of Strep. mutans and is more so than inhibiting planktonic cells without toxicity. The ribosome and histidine metabolism pathways of Strep. mutans were significantly regulated by this compound. Conclusions These results suggest that the 2-AI/T conjugate is a potent inhibitor that can be potentially developed into a new drug to treat and prevent dental caries. Significance and impact of the study This is the first study to use small molecule from marine natural products, to protect from dental caries in vivo. It has potential broad range application in clinical caries prevention, or as a bioactive ingredient for food applications.

Targeting antigens to antigen-presenting cells by fusion to cytotoxic T lymphocyte-associated antigen 4 (CTLA4) has been shown to be a highly efficient method to enhance the efficacy of DNA vaccines. The purpose of this study was to determine the immunogenicity and protective efficacy of the targeted fusion DNA construct pGJA-P, which contains the signal peptide and extracellular regions of human CTLA4 gene, the hinge and Fc regions of human Igγ1 gene, the glucan-binding domain of the Streptococcus mutans gtfB gene and the A-P fragment of the S. mutans pac gene, compared with the fusion DNA construct pGLUA-P, which contains only the glucan-binding domain of the S. mutansgtfB gene and the A-P fragment of the S. mutans pac gene. BALB/c mice were immunized with pGJA-P, pGLUA-P, or pCI (vector) by the intramuscular or intranasal route. Specific anti-PAc and anti-GTF-I serum IgG and salivary IgA antibody responses were assessed by an enzyme-linked immunosorbent assay. Wistar rats were orally challenged with S. mutans and immunized with pGJA-P, pGLUA-P, or pCI intramuscularly or intranasally, and caries activity was evaluated by the Keyes method. pGJA-P induced accelerated and increased serum and salivary antibody responses in mice compared with pGLUA-P. Rats immunized with pGJA-P had significantly fewer caries lesions than rats immunized with pGLUA-P (p < 0.01). Thus, this study demonstrates that the targeted DNA construct pGJA-P can enhance both systemic and mucosal immunity and may be a useful strategy for improving the protective efficacy of anticaries DNA vaccines.

Glucosyltransferases (GTFs) and A cell-surface protein (PAc) are two important virulence factors of the cariogenic organism Streptococcus mutans. They may mediate sucrose-independent or sucrose-dependent attachment of Streptococcus mutans to tooth surfaces, respectively. Thus, inhibiting both virulence factors is predicted to provide better protection against caries than inhibiting a single factor. To develop a highly efficient vaccine against caries, we constructed a fusion DNA vaccine, pGLUA-P, by cloning the GLU region of GTF into a DNA vaccine, pCIA-P, which encodes two highly conservative regions of PAc. In this report, we provide evidence that fewer caries lesions were observed in rats following subcutaneous injection of pGLUA-P, compared with pCIA-P, near the submandibular gland. Our findings suggest that a multigenic DNA vaccine may be more caries-preventive than a single-gene DNA vaccine.

The purpose of this study was to investigate whether there was transmission between adults in Chinese families using chromosomal DNA fingerprinting. Plaque samples were obtained from buccal and occlusal surfaces of 11 married couples. The colonization levels of mutans streptococci were estimated as colony-forming units per milliliter, and caries experience was scored by decayed, missing and filled teeth. Information about medical history, diet regimes and age at marriage was obtained. The isolates were serotyped by biochemical test and genotyped using the restriction endonuclease HaeIII. The procedure was repeated after 3 months. The results showed that 1 couple had the same genotype of mutans streptococcus at the first examination, but this could not be repeated for the husband who had lost his mutans streptococci at the second examination. On the contrary, another couple that did not have the same mutans streptococcal genotype at the beginning had the same genotype after 3 months. No matching of genotypes was observed within 8 couples. In 1 male, no mutans streptococci were detected, therefore that couple was not considered. These data indicate that spouses had a chance to be infected by strains of mutans streptococci from another person. The results suggest that there may be transmission between adults in Chinese families, but it may be difficult for mutans streptococci to colonize another mouth permanently.

To investigate the effects of co-delivering IL-6 expressing plasmid pCI-IL-6 on the immunogenicity of the anti-caries DNA vaccine pCIA-P, which encodes the surface protein antigen PAc of Streptococcus mutans.Plasmid pCI-IL-6 was constructed by inserting the murine IL-6 gene into the pCI vector. Expression of IL-6 in vitro was assessed using Western blot analysis. BALB/c mice were intranasally co-immunized with pCIA-P plus pCI-IL-6 on d 0 and 14. Anti-PAc IgG and secretory IgA (sIgA) were assessed by ELISA. Splenocytes from the mice were re-stimulated with the PAc protein, and IFN-γ and IL-4 production was measured using ELISA. Splenocyte proliferation was analyzed with flow cytometry. Rats were similarly immunized, and dental caries scores were determined using the Keyes method.Marked expression of IL-6 was found in COS-7 cells transfected with pCI-IL-6. In the pCI-IL-6 co-immunized mice, the specific IgG antibodies in serum and sIgA antibodies in saliva were significantly higher than those in the control mice at weeks 4 and 8. Moreover, the secretion of IFN-γ from splenocytes in response to re-stimulation with PAc protein was significantly higher in the pCI-IL-6 co-immunized mice than that in the control mice, whereas the secretion of IL-4 had no significant difference. The proliferation of splenocytes from the pCI-IL-6 co-immunized mice was significantly higher than that from the mice immunized with pCIA-P and pCI vector. In the rat caries model, the pCI-IL-6 co-immunization rats displayed lower caries scores than the control rats.Intranasal co-delivery of IL-6 gene significantly enhances the immunogenicity of the anti-caries DNA vaccine.

DNA vaccine plasmids were constructed that encoded two highly-conservative regions of a surface protein, PAc, from the human major cariogenic bacterium, Streptococcus mutans. Antigen expression was evaluated in vitro by immunohistochemical analysis of human endothelial cells following cationic liposome-mediated transient transfection with recombinant plasmid. The results of this study provided a basis for further testing of these recombinant plasmids in primates and for efficacy testing of dental caries DNA vaccines in human volunteers in future.

The objective of this study was to assess the effect of six-monthly professional applications of chlorhexidine varnish on the prevention of dental caries in primary molars in Chinese preschool children. In a double-blinded, randomized, placebo-controlled clinical trial, 334 children aged 4–5 years were randomly divided into two groups. Children in the test group received six-monthly applications of a 40% chlorhexidine varnish, and the control children received a placebo varnish. Caries status of the children was assessed by two calibrated examiners at baseline and after 24 months, according to criteria recommended by the World Health Organization. The two-year mean caries increments in the test and the control group children were 1.0 and 1.6 decayed, missing, or filled molar surfaces (dmfs-molar), respectively, a 37.3% reduction ( t test, p = 0.036). No side-effects were found. It was concluded that six-monthly applications of chlorhexidine varnish were effective in reducing the incidence of dental caries in primary molars.

Item does not contain fulltext OBJECTIVE: The hypotheses tested were: the cumulative survival rates of dentin caries lesion-free pits and fissures of ART conventional high-viscosity glass-ionomer sealants with light-curing (high-intensity LED) and glass-carbomer sealants are higher than those of conventional ART sealants and resin-composite sealants after 4 years. METHODS: The randomized controlled clinical trial covered 405 children (mean age 8-years). Three dentists placed sealants in pits and fissures of high caries-risk children. Evaluation by two independent evaluators was conducted after 0.5, 1, 2, 3 and 4 years. The Kaplan-Meier survival method, ANOVA and t-test were used in data analyses. RESULTS: 1304 first permanent molars were sealed. 12.3% of children and 15.4% of sealants dropped out. 46 re-exposed pits and fissures, 39 (occlusal) 7 (free smooth surfaces), in 42 children developed a dentin carious lesion. The cumulative survival of dentin caries lesion-free occlusal pits and fissures in ART plus LED group (98%) was statistically significantly higher than in the resin-composite group (96.4%) and in the glass-carbomer group (94.5%). The cumulative survival of dentin caries lesion-free occlusal pits and fissures in the glass-carbomer group was statistically significantly lower than that in the conventional ART group (97.3%). For the free smooth surfaces, there was no statistically significantly difference among the four sealant groups. SIGNIFICANCE: Light-cured ART conventional high-viscosity glass-ionomer sealants prevented the occurrence of dentin cavities best.

Item does not contain fulltext OBJECTIVES: Data showed that among 5-year-old Chinese, 96.7% of cavitated primary teeth were left untreated. The study aimed to report on the course of cavitated primary teeth within the Chinese health-care system over a period of 3.5 years. METHODS: Selection of high caries risk children for inclusion in a sealant comparison study was based on the presence of cavitated dentine lesions in their primary teeth. At the 6-month sealant evaluation point many of these cavitated dentine lesions had not been treated. This necessitated monitoring these cavitated teeth 6-monthly for those exfoliated, restored, with a cavity left open, having caused toothache (symptom) and having (or having had) an abscess or fistulae (symptom). Care-seeking instruction was given at every evaluation point. anova and t-test were used in analysing the data. RESULTS: A total of 1012 cavitated primary teeth in 305 children (7.6 to 9.3 years old), were followed for 3.5 years. A total of 92.9% of cavitated primary teeth were left open, while 7.1% were restored at some stage during the observation period; 98.5% of restored teeth and 95.5% of cavitated teeth left open exfoliated and 93.9% of restored teeth and 81.5% of cavitated primary teeth left open exfoliated without any symptoms. Having (or having had) toothache was the symptom most frequently related to exfoliated restored teeth and to exfoliated cavitated teeth left open. Restored primary teeth survived statistically significantly longer than cavitated primary teeth left open: 1.99 +/- 0.07 years and 1.68 +/- 0.03 years, respectively. CONCLUSION: The large majority of cavitated primary teeth in this child population exfoliated without symptoms.

To investigate how co-delivery of the gene encoding C-C chemokine ligand-19 (CCL-19) affected the systemic immune responses to an anti-caries DNA vaccine pCIA-P in mice.Plasmid encoding CCL19-GFP fusion protein (pCCL19/GFP) was constructed by inserting murine ccl19 gene into GFP-expressing vector pAcGFP1-N1. Chemotactic effect of the fusion protein on murine dendritic cells (DCs) was assessed in vitro and in vivo using transwell and flow cytometric analysis, respectively. BALB/c mice were administered anti-caries DNA vaccine pCIA-P plus pCCL19/GFP (each 100 μg, im) or pCIA-P alone. Serum level of anti-PAc IgG was assessed with ELISA. Splenocytes from the mice were stimulated with PAc protein for 48 h, and IFN-γ and IL-4 production was measured with ELISA. The presence of pCCL19/GFP in spleen and draining lymph nodes was assessed using PCR. The expression of pCCL19/GFP protein in these tissues was analyzed under microscope and with flow cytometry.The expression level of CCL19-GFP fusion protein was considerably increased 48 h after transfection of COS-7 cells with pCCL19/GFP plasmids. The fusion protein showed potent chemotactic activity on DCs in vitro. The level of serum PAc-specific IgG was significantly increased from 4 to 14 weeks in the mice vaccinated with pCIA-P plus pCCL19/GFP. Compared to mice vaccinated with pCIA-P alone, the splenocytes from mice vaccinated with pCIA-P plus pCCL19/GFP produced significantly higher level of IFN-γ, but IL-4 production had no significant change. Following intromuscular co-delivery, pCCL19/GFP plasmid and fusion protein were detected in the spleen and draining lymph nodes. Administration of CCL19 gene in mice markedly increased the number of mature DCs in secondary lymphoid tissues.CCL19 serves as an effective adjuvant for anti-caries DNA vaccine by inducing chemotactic migration of DCs to secondary lymphoid tissues.

Background.gcrRgene acts as a negative regulator related to sucrose-dependent adherence inS. mutans. It is constructive to test the potential capacity of mutans withgcrRgene deficient in bacteria replacement therapy.Methods. In this study, we constructed the mutant by homologous recombination. The morphological characteristics of biofilms were analyzed by confocal laser scanning microscopy.S. mutansUA159 and the mutant MS-gcrR-def were inoculated, respectively, or together for competitive testing in vitro and in rat model.Results. Adhesion assay showed that the adhesion ability of the mutant increased relative to the wild type, especially in the early stage. MS-gcrR-def out-competedS. mutansUA159 in vitro biofilm, and correspondingly coinfection displayed significantly fewer caries in vivo. The former possessed both a lower level of acid production and a stronger colonization potential thanS. mutansUA159.Conclusion. These findings demonstrate that MS-gcrR-def appears to be a good candidate for replacement therapy.

The design of optimized nanoparticles offers a promising strategy to enable DNA vaccines to cross various physiological barriers for eliciting a specific and protective mucosal immunity via intranasal administration. Here, we reported a new designed nanoparticle system through incorporating anionic liposomes (AL) into chitosan/DNA (CS/DNA) complexes. With enhanced cellular uptake, the constructed AL/CS/DNA nanoparticles can deliver the anti-caries DNA vaccine pGJA-P/VAX into nasal mucosa. TEM results showed the AL/CS/DNA had a spherical structure. High DNA loading ability and effective DNA protection against nuclease were proved by gel electrophoresis. The surface charge of the AL/CS/DNA depended strongly on pH environment, enabling the intracellular release of loaded DNA via a pH-mediated manner. In comparison to the traditional CS/DNA system, our new design rendered a higher transfection efficiency and longer residence time of the AL/CS/DNA at nasal mucosal surface. These outstanding features enable the AL/CS/DNA to induce a significantly (p

Item does not contain fulltext The aim of the present investigation was to assess the impact of dental caries prevalence and the consequences of untreated cavitated dentine lesions on quality of life of 6- and 7-year-old Brazilian children. A total of 826 schoolchildren were assessed using ICDAS and pufa (to score consequences of dental caries on soft tissues) indices. History of extraction and toothache was recorded. Oral health-related quality of life was assessed using the Brazilian version of the Early Childhood Oral Health Impact Scale (B-ECOHIS). A multiple logistic regression model was used to analyze the relationship between the prevalence of dentine carious lesions, pufa, history of extraction and toothache with the B-ECOHIS scores. A total of 587 questionnaires were analyzed. The prevalence of cavitated dentine lesions and pufa was 74.8 and 26.2%, respectively. Some 21.8% of children reported toothache and 9.2% had had at least one tooth extraction. The chance (OR) for children with cavitated dentine lesions, pufa >/=1, history of extraction and toothache of having higher B-ECOHIS scores than those not affected was 1.90 (95% CI: 1.18-3.06), 6.26 (95% CI: 3.63-10.83), 6.87 (95% CI: 2.75-17.16) and 3.68 (95% CI: 2.12-6.39), respectively. Children's quality of life was negatively influenced by untreated cavitated dentine lesions and their consequences.

To prepare a clinical-grade anti-caries DNA vaccine pGJA-P/VAX and explore its immune effect and protective efficacy against a cariogenic bacterial challenge. A large-scale industrial production process was developed under Good Manufacturing Practices (GMP) by combining and optimizing common unit operations such as alkaline lysis, precipitation, endotoxin removal and column chromatography. Quality controls of the purified bulk and final lyophilized vaccine were conducted according to authoritative guidelines. Mice and gnotobiotic rats were intranasally immunized with clinical-grade pGJA-P/VAX with chitosan. Antibody levels of serum IgG and salivary SIgA were assessed by an enzyme-linked immunosorbent assay (ELISA), and caries activity was evaluated by the Keyes method. pGJA-P/VAX and pVAX1 prepared by a laboratory-scale commercial kit were used as controls. The production process proved to be scalable and reproducible. Impurities including host protein, residual RNA, genomic DNA and endotoxin in the purified plasmid were all under the limits of set specifications. Intranasal vaccination with clinical-grade pGJA-P/VAX induced higher serum IgG and salivary SIgA in both mice and gnotobiotic rats. While in the experimental caries model, the enamel (E), dentinal slight (Ds), and dentinal moderate (Dm) caries lesions were reduced by 21.1%, 33.0%, and 40.9%, respectively. The production process under GMP was efficient in preparing clinical-grade pGJA-P/VAX with high purity and intended effectiveness, thus facilitating future clinical trials for the anti-caries DNA vaccine.

As we know, the catalytic region of glucosyltransferases (GTFs) is a key region responsible for sucrose-dependent adherence of cariogenic bacteria to teeth. In this study, we evaluate the potential of the catalytic region to enhance the immunogenicity of the anti-caries DNA vaccine. We construct two new anti-caries DNA plasmids pGJGAC/VAX and pGJGA-5C/VAX by cloning different styles of the catalytic regions of GTFs into the previous plasmid pGJA-P/VAX. One is the 1.1 kb full length catalytic region of S. sobrinus GTF-I, the other is its catalytic core sequence which is conserved in the GTFs from mutans streptococci and plays a central role in the enzymatic activities of sucrose splitting and glucan synthesis. The results of caries protection experiment indicate that compared to pGJA-P/VAX, immunization with both new plasmids provides more effective protection against cariogenic bacteria, especially against S. sobrinus. The plasmid encoding full length catalytic region could provide more effective protection against cariogenic bacteria than that encoding catalytic core conserved sequence even though the differences are not very dramatic.

(1) To evaluate trends in dental caries of children and adolescents of Hubei Province and Wuhan Municipality over the last two decades. (2) To conduct a situational analysis of existing oral health care and to recommend changes to improve this care for children.An analysis was performed on data from national oral health surveys carried out in China at the provincial level, and data on 12-year-old children of Wuhan Municipality. A review of the literature was conducted on available and utilised oral health care facilities by children.In 1983, the mean decayed, missing and filled teeth (DMFT) of 12-year-olds in Hubei Province was 1.0, and in 1995 it was 0.6. The studies in Wuhan Municipality in 1993-1995 among 12-year-olds reported DMFT values ranging from 0.7-1.5, however different methods of caries assessment were performed in the various studies making comparisons difficult. The caries prevalence and the mean dmft score of 5-year-old children in 1995 in Hubei Province were 69% and 3.6, respectively. The percentage of untreated caries for all age groups in Hubei Province was very high.Analysis of epidemiological information over the last two decades does not provide any conclusive evidence supporting either an increasing or decreasing trend of dental caries in the child and adolescent population of Hubei Province and Wuhan Municipality. There is, however, a clear need to prevent the incidence of dental caries from increasing and to provide care to address the treatment needs of pre-school and school-age children. The promotion of twice-daily tooth brushing with fluoride toothpaste at maternal and child health care centres, and the establishment of daily tooth brushing with pea-size amounts of fluoride toothpaste in the pre-school and primary school setting are recommended preventive strategies. Establishment of school-based oral health promotion should be accompanied with the provision of Atraumatic Restorative Treatment (ART) sealants and ART restorations which can be provided by dental nurses at dramatically lower costs than the traditional high-tech approach.

Objective To study the biodistribution and expression of anti-caries DNA vaccine pGJA-P/VAX in mice models following intranasal (i.n.) and intramuscular (i.m.) immunization. Methods pGJA-P/VAX and pVAX1 were administrated i.n. and i.m. to mice, respectively. Blood and organs were harvested and then genomic DNA was extracted. mRNA in the original tissues and draining lymph nodes was isolated. The in vivo fate of plasmid DNA in these samples was evaluated with real-time quantitative polymerase chain reaction (PCR). Results The ratio of the area under the curve (AUC) and maximum concentration (Cmax) after i.n. immunization were higher than that after i.m. injection. At 1 day post i.n. immunization, the localization of plasmid to the heart, spleen and lung was notable compared with that to the nasal mucosa. At 2 days, for i.n. immunization, the concentration value of plasmid in the mucosa-associated lymphoid tissues such as lung, kidney and gastrointestine had increased and was greater than that via i.m. injection. At later time points, the plasmid was primarily in the original tissues and draining lymph nodes. mRNA expression was observed in the topical tissues up to 7 days and in the draining lymph nodes for at least 3 days. Conclusions These results indicate that the distribution of pGJA-P/VAX was widespread in vivo, and the expression was significant in the inoculation tissue and draining lymph nodes after i.n. immunization and i.m. injection. This data provides evidence which supports the long-term immune responses and the basis of further research on safety issues. Copyright © 2007 John Wiley & Sons, Ltd.

Contains fulltext : 70426.pdf (Publisher’s version ) (Closed access) OBJECTIVES: To investigate the role of SecA in protein secretion, and to evaluate the effect of biofilm formation on protein secretion in Streptococcus mutans. DESIGN: S. mutans strains UA159 and GS-5 were used in this study. Cells grown in biofilm and planktonic conditions were observed using immunogold electron microscopy. The mRNA levels of ftf, gtfB, gtfC, gtfD, Pac and secA were analysed in different growth conditions using real-time quantitative polymerase chain reaction. The levels of wall proteins and whole-cell protein extracts were examined using Western blot analysis. RESULTS: A microdomain colocalised with SecA and virulence factors such as Pac (AgI/II) and glucosyltransferase (GTF) was observed. The mRNA level of secA was upregulated in the biofilm condition. The level of protein expression of SecA and wall protein levels of GTF, fructosyltransferase (FTF) and Pac (AgI/II) in the biofilm condition were significantly higher than in the planktonic condition. CONCLUSIONS: These data suggest that S. mutans utilises the Sec pathway to secrete virulence factor proteins such as Pac (AgI/II), GTF and FTF, and protein secretion occurred at a distinct microdomain. The level of SecA, the key factor in the Sec pathway, was influenced significantly by biofilm formation in S. mutans.

Mucosal immune responses act as the first line of defense against dental caries. In this study, an optimal vaccination strategy was developed to enhance anti-caries mucosal immune responses. Mice and rats were vaccinated intranasally firstly with plasmid pCIA-P encoding PAc antigen of Streptococcus mutans and then with rPAc, or with pCIA-P for twice, or with rPAc protein for twice, respectively. The potential of inducing mucosal and systemic immune responses to special antigens was measured by ELISA. In addition, antibody type, cytokine production and protection effectiveness against dental caries were also evaluated. Although all immunized groups developed immune responses, the antibody responses in the DNA prime–protein boost group were stronger compared with those elicited by either the DNA vaccine or the protein vaccine. In particular, the Th1-biased response that was established by the DNA immunization was diverted to Th1/Th2-mixed response following the rPAc protein boost. Moreover, protection against S. mutans challenge was obtained in the rats treated with the DNA prime–protein boost regimen, as shown by a significant reduction in dental caries lesion, compared with the control groups immunized with the DNA or protein only. All these findings may provide useful information about effective mucosal vaccines against dental caries.

Aim: To evaluate the comparative immunogenicity and protective efficacy of the cytotoxic T-lymphocyte-associated antigen 4 (CTLA-4) fusion anti-caries DNA vaccines pGJA-P/VAX1, pGJA-P, and non-fusion anti-caries DNA construct pGLUA-P in hamsters. In addition, the ability of CTLA-4 to target pGJA-P/VAX1-encoding antigen to dendritic cells was tested in vitro . Methods: All DNA constructs contain genes encoding the A-P regions of a cell surface protein (PAc) and the glucan binding (GLU) domain of glucosyltransferases (GTFs) of cariogenic organism Streptococcus mutans. Human dendritic cells were mixed with the CTLA-4-Ig-GLU-A-P protein expressed by pGJA-P/VAX1-transfected cells and analyzed by flow cytometry. Gnotobiotic hamsters were immunized with anti-caries DNA vaccines by intramuscular injection or intranasal administration. Antibody responses to a representative antigen PAc were assayed by ELISA, and caries protection was evaluated by Keyes caries scores. Results: A flow cytometric analysis demonstrated that CTLA-4-Ig-GLU-A-P protein was capable of binding to human dendritic cells. pGJA-P/VAX1 and pGJA-P induced significantly higher specific salivary and serum anti-PAc antibody responses than pGLUA-P. Significantly fewer caries lesions were also observed in hamsters immunized with pGJA-P/VAX1 and pGJA-P. There was no significant difference in the anti-PAc antibody level or caries scores between pGJA-P/VAX1 and pGJA-P-immunized groups. Conclusion: Antigen encoded by CTLA-4 fusion anti-caries DNA vaccine pGJA-P/VAX1 could specifically bind to human dendritic cells through the interaction of CTLA-4 and B7 molecules. Fusing antigen to CTLA-4 has been proven to greatly enhance the immunogenicity and protective efficacy of anti-caries DNA vaccines.

Enhancement of mucosal and systemic immune responses is still a challenge for the application of DNA vaccine. Here, we show anti-caries DNA vaccines, pGJA-P and pGJA-P/VAX, encoding Streptococcus mutans antigens fused to cytotoxic T lymphocyte antigen-4 (CTLA4), which binds to B7 molecule expressed on the surfaces of antigen-presenting cells. Rabbits and monkeys were immunized via intranasal or intramuscular routes. The fusion vaccine induced accelerated and increased specific antibody responses in serum and saliva compared with non-fusion DNA vaccine in rabbits. Significant specific serum IgG and salivary IgA levels could be detected in fusion vaccine-immunized monkeys. Therefore, this study demonstrates that fusing antigens to CTLA4 results in enhancing immune efficacy and strongly suggests that it may represent a promising approach to prevent dental caries or other mucosal infectious diseases. These findings also suggest that CTLA4 fusion anti-caries DNA vaccine may be effective immunogen in primates.

Objectives: To assess oral health outcomes of a school-based oral health education (OHE) programme on children, mothers and schoolteachers in China, and to evaluate the methods applied and materials used. Design: The WHO Health Promoting Schools Project applied to primary schoolchildren in 3 experimental and 3 control schools in Hongshan District, Wuhan City, Central China, with a 3-year follow-up. Data on dental caries, gingival bleeding and behaviour were collected. Participants: 803 children and their mothers, and 369 teachers were included at baseline in 1998. After three years, 666 children and their mothers (response rate 83%), and 347 teachers (response rate 94%) remained. Results: DMFT/DMFS increments were comparable but the f/F components were higher among children in experimental schools than in control schools and the gingival bleeding score was, similarly, significantly lower. More children in experimental schools adopted regular oral health behaviour such as toothbrushing, recent dental visits, use of fluoride toothpaste, with less frequent consumption of cakes/biscuits compared to controls. In experimental schools, mothers showed significant beneficial oral health developments, while teachers showed higher oral health knowledge and more positive attitudes, also being satisfied with training workshops, methods applied, materials used and involvement with children in OHE. Conclusions: The programme had positive effects on gingival bleeding score and oral health behaviour of children, and on oral health knowledge and attitudes of mothers and teachers. No positive effect on dental caries incidence rate was demonstrated by the OHE programme.

To investigate the effect of specific anti-streptococcus mutans IgY against streptococcus mutans on dental caries development in rats.35 wistar rats were divided into 5 groups: group A received IgY gargle; group B received IgY lyophilized powder; group C received sterilized water as control; group D and E received egg yolk food with or without specific IgY individually. They were all fed with caries-inducing diet 2000#. The number of caries scores was counted by the procedure of Keyes'.There was a significant lower mean of caries scores in groups treated with IgY lyophilized powder and gargle. By treating with egg-yolk food contained specific IgY, the mean of caries scores decreased comparing with no treatment group.Local passive immunization with specific anti-streptococcus mutans IgY may be an effective way to prevent the development of dental caries.

Salivary secretory immunoglobulin A (S-IgA) antibodies act as the first line of defense against dental caries by blocking of adherence of Streptococcus mutans to tooth surfaces. This study focused on finding proper mucosal immunization route and delivery system to induce higher level of specific anti-S. mutans saliva S-IgA and inhibit dental caries in animal model. By immunizing rats with an anti-caries DNA vaccine, pCIA-P, via different mucosal routes, we found that intranasal (i.n.) immunization with pCIA-P/bupivacaine DNA complexes elicited the highest specific anti-S. mutans saliva S-IgA mucosal antibody responses compared with naked DNA and other routes. Correspondingly, rats immunized with pCIA-P/bupivacaine DNA complex via i.n. displayed the least carious lesions. Our findings suggested that DNA vaccination via intranasal immunization with bupivacaine delivery system be a promising approach against dental caries.

To assess the efficacy of plasmid DNA encoding pac gene of Streptococcus mutans (S. mutans) intranasally immunized in gnotobiotic rats and to compare the effect of two different delivery systems.Sprague Dawley rats, infected with S. mutans at 20 days of age, were intranasally immunized with plasmid pCIA-P (group A), Dosper-DNA complex (group B), Bupivacaine-DNA complex (group C). Control rats were either immunized with plasmid pCI (group D), distilled water (group E) or immunized intramuscularly (group F). All the rats were boosted 2 weeks later. ELISA determined the antibodies against the vaccines. Keyes caries score was used to evaluate the anti- caries effectiveness of the vaccines at the terminal study.As for the antibody reactions, there were significantly (P0.01) differences between rats immunized with DNA vaccine and non-immunized rats. And rats in group B and C had the significantly (P0.01) higher level of specific salivary anti-PAc IgA antibodies and rats (group B, C, F) had the significantly (P0.01) higher specific serum anti-PAc IgG responses to DNA vaccine. Keyes scores of rats (group B and C) were significantly (P0.01) lower than others.Intranasal immunization with plasmid pCIA-P encoding pac gene successfully reduces the caries and appears to be a promising approach against dental caries. Cationic liposome Dosper and local anesthetic bupivacaine could enhance the efficacy of DNA vaccine.

To construct a fusion anti-caries DNA vaccine pGLUA-P carrying GLU fragment from gtfB gene of Streptococcus mutans GS-5 and A-P fragment including the A region and P region of PAc protein from a DNA anti-caries vaccine pCIA-P, and to investigate its expression in prokaryotic and eukaryotic cells.The sequence of GLU fragment in pGLU plasmid was testified by DNA sequencing. The fusion anti-caries DNA vaccine was constructed by ligating A-P fragment from pCIA-P to pGLU. The expression of GLUA-P fusion protein in E. coli BL21 (DE3) was induced by IPTG and checked by SDS-PAGE electrophoresis. pGLUA-P was transfected in vitro to cultured rat primary muscle cells by cation liposome Dosper, and immunohistochemical method was used to test the expression of GLUA-P fusion protein in cells.GLU sequence was identical with relative sequence of GTF-I (GS-5 strain) in Gene Bank. Recombinant eukaryotic expression plasmid pGLUA-P was confirmed to have both GLU and A-P fragment. After pGLUA-P was transferred into E. coli (DE3), it could express a new 115 000 protein by the induce of IPTG. Specific brown products could be found in the cytoplasm of cultured rat primary muscle cells transfected by pGLUA-P.Fusion anti-caries DNA vaccine pGLUA-P is successfully constructed and confirmed by sequencing and enzymes digestion. Fusion GLUA-P protein can be correctly expressed in prokaryotic and eukaryotic cells.

A cell-surface protein antigen (PAc) of Streptococcus mutans is considered a virulence factor because it may mediate initial attachment of Streptococcus mutans to tooth surfaces. Thus, inhibiting PAc is predicted to provide protection against caries. To develop vaccines against dental caries, we constructed a DNA vaccine, pCIA-P, which encodes two high-conservative regions of PAc. Expression of the recombinant protein was obtained in eukaryotic cells in vitro and in vivo. In this report, we provide evidence that fewer caries lesions, and high levels of PAc-specific salivary IgA antibody and serum IgG antibody, were observed in gnotobiotic rats following targeted salivary gland (TSG) administration of pCIA-P. This study shows that the recombinant DNA vaccine pCIA-P could induce protective anti-caries immune responses and that TSG immunization is a promising strategy for the inhibition of dental caries.

Streptococcus mutans and Streptococcus sobrinus are associated with the development of dental caries. The purpose of this study was to differentiate these bacteria by morphology, biochemical characteristics and PCR, and to compare their occurrence with the prevalence of dental caries in the Mosuo people. Plaque samples were collected from the permanent first molar in 126 Mosuo people (83 females, 43 males, aged 25-55 years, average age 36.1 +/- 7.73). Dental status was recorded as DMFT by WHO caries diagnostic criteria. Males had a significantly lower prevalence of caries and DMFT than females: 11.4 vs. 86.9% and 1.65 vs. 6.95, respectively (p0.001). Morphological and biochemical tests gave unreliable results. The prevalence of S. mutans and S. sobrinus was 75.4 and 57.1%, respectively. 26.5% of females and 53.5% of males were positive for S. mutans alone, 18.1% of females and 16.3% of males were positive for S. sobrinus alone, while 50.6% of females and 18.6% of males were positive for both S. mutans and S. sobrinus and 4.8% of females and 11.6% of males were negative for both species. The DMFT scores of subjects positive for both S. mutans and S. sobrinus were significantly higher than of those positive for either S. mutans or S. sobrinus alone. These results indicate that subjects harboring both S. mutans and S. sobrinus have a significantly higher prevalence of dental caries than those with S. mutans or S. sobrinus alone.

Protein of Streptococcus mutans is considered as one of the virulence factors due to its ability to mediate the initial attachment of Streptococcus mutans to tooth surface. In this study, an anticaries DNA vaccine pCIA-P was used to immunize rats. The expression of PAc in different tissues in vivo, specific immune response and protection effects against dental caries were observed.Plasmid pCIA-P was injected into rats by two different routs: intramuscular injection (i.m.) and targeted salivary gland immunization (TSG). Immunohistochemistry technique was used to detect the expression of PAc. Gnotobiotic rats were vaccinated with pCIA-P by three different approaches: TSG, intramuscular injection and buccal mucosal injection (i.o.). The specific immune responses were evaluated by ELISA and their anticaries effects were evaluated by Keyes caries scores.PAc was expressed in the sarcoplasm and sarcolemma of muscle fibers and submandibular glands, especially strongly positive in duct regions. The levels of serum specific anti-PAc IgG and salivary specific anti-PAc IgA in TSG immunization and buccal mucosal immunization group were significantly higher than those of other groups. The Keyes caries scores of those two groups were significantly lower than those of other groups.The plasmid pCIA-P could provoke specific immune responses as a novel immunogen. Mucosal immunization with pCIA-P appears to be an effective genetic immunization method against dental caries.

The rationale of bacterial replacement therapy against dental caries is that relatively avirulent strains of Mutans Streptococi (MS) are most likely to occupy the same ecological niche in plaque as their more cariogenic counterparts. As known, lactate dehydrogenase (LDH) deficiency has been proposed as one aspect of a strategy to construct an effector strain because LDH-deficient mutants could affect acid production by MS so as to reduce their cariogenicity. Glucan-binding lectin (GBL) plays a very important role in the formation of dental plaque biomembrane and the adherence of S. mutans to teeth surface. The gcrR gene acts as a negative transcriptional regular of gbpc gene which encoded S. mutans GBL. We presume an LDH-deficient S. mutans mutants which harbours an insertion-deletion mutation in gcrR gene can result in overexpression of GBL and higher adherence to teeth than the wild-type S. mutans and will possess of both low-level acid production and strong colonization potential.

The purpose of this study was to evaluate the changes of oral health knowledge and behaviour among Wuhan citizens after six years of 'Love Teeth Day' campaigns. Representative samples of citizens aged 10-62 years were identified and interviewed in 1987 (n = 1180) and in 1995 (n = 1014). Data on oral health knowledge and practises were collected by structured questionnaires in both surveys and high response rates were obtained (87-94 per cent). In general, improvements in oral health knowledge and toothbrushing behaviour were observed in 1995 as compared with 1987. The younger age group (10-19-year-olds) showed significantly higher improvements of positive answers to knowledge items and toothbrushing behaviour. The findings also indicated that less than half of the interviewees were aware of the anti-caries effect of fluoride and about 30 per cent of the participants brushed their teeth performing the recommended method (vertical). In China, community based oral health education programmes should be implemented to further improve regular self-care practises, especially in relation to the adult population.