[Materials for Selective Inhibition of Streptococcus mutans and Progress in Relevant Research].

Dental caries is a disease in which chronic progressive destruction of the hard dental tissues occurs under the influence of multiple factors, among which, bacterial infection being the most important one. Dental plaque biofilm is a key factor in the pathogenesis of dental caries. Under normal circumstances, microorganisms within the biofilm maintain a dynamic balance through coordination, competition, and antagonism. However, when the environment changes, the balance in the biofilm will be disrupted, and the number of cariogenic bacteria, especially Streptococcus mutans ( S. mutans), will increase significantly, thereby causing the production of large amounts of organic acids on the tooth surface, tooth demineralization, and the formation of dental caries. Therefore, finding ways to restore the dynamic balance of oral microorganisms through selective inhibition of S. mutans is key to the prevention and treatment of dental caries. Herein, we reviewed the research progress of recent years in the development of materials with selective antibacterial effect, intending to provide references for the further development of drugs for the prevention and treatment of dental caries. Future studies should focus on the following aspects, mechanism, clinical efficacy, chemical modification, and safety, to supplement and make improvements on the existing relevant research, and to promote progress in research and development of drugs for the prevention and treatment of dental caries.

[Regulatory Effect of All-Trans Retinoic Acid on the Expression of IL-1ß in Macrophages and the Mechanisms Involved].

Objective: To investigate the regulatory effect of all-trans retinoic acid (ATRA) on the expression interleukin-1ß (IL-1ß) in macrophages and the mechanisms involved. Methods: Macrophages were treated with 1 µmol/L ATRA for 24 h before RNA-Sequence. Differentially expressed genes (DEGs) were screened out and analyzed by Kyoto Encyclopedia of Genes and Genomes (KEGG) pathway enrichment analysis, gene ontology (GO) functional analysis, and protein-protein interaction networks (PPI) analysis. After treatment with different doses of ATRA for 24 h, the expression of IL-1ß was examined with qRT-PCR and Western blot. The activation of NF-κB signaling and caspase-1 was observed by Western blot and immunofluorescence staining. Results: Compared with the blank control group, a total of 71 DEGs of macrophages were upregulated in the ATRA treatment group. KEGG analysis showed that the up-regulated DEGs were involved in IL-17 signaling pathway, tumor necrosis factor (TNF) signaling pathway, etc. GO analysis indicated that the up-regulated DEGs were involved in the biological processes of the production of IL-1ß, response to lipopolysaccharide, etc. PPI analysis revealed that inflammatory cytokines, adhesion molecules, and chemokines were the key genes that ATRA acted on. In vitro experiments showed that ATRA promoted IL-1ß expression in macrophages in a concentration-dependent manner. The expression of p-NF-κB, NF-κB, and caspase-1 were significantly increased by ATRA compared with those of the control group ( P<0.05), and p-NF-κB translocated to the cell nucleus in the ATRA group. Conclusion: ATRA may promote the expression of IL-1ß by activating NF-κB signaling and caspase-1 in macrophages, this study may provide evidence for the immune regulatory function of ATRA on macrophages.

[Application Progress of Dual RNA Sequencing in Microbial Research].

The human body is colonized by densely-populated and structurally complex communities of microorganisms. The microbiota interact not only with their host cells, but also with other microbiota. Dual RNA sequencing (Dual RNA-seq) can be used to conduct simultaneous analysis of the dynamic changes of gene expression of two (or more) interactive species, and to obtain thus, through the interaction model diagram, the inter-species regulatory relationship of genes of different species, and hence the interaction mechanism between species. We herein reviewed the application status and development prospects of Dual RNA-seq in the research of intestinal, respiratory, skin and oral microbes. Since the concept of Dual RNA-seq was first introduced, the technology has been applied to a range of infection models. Direct investigation into the dynamic interactions between species at the molecular level will contribute to the better understanding of the physiological changes of pathogens and hosts during the course of infection, and thus help reveal potential new targets or biomarkers. However, the Dual RNA-seq technology is still in its early stage of development, and there are some limitations in the experimental technology. For example, due to the dynamic nature of the interaction between species, there are urgent problems awaiting solutions, such as the optimal experimental conditions, the selection of sampling sites and how to achieve real-time observation. In addition, due to the large amount of bioinformatics data of Dual RNA-seq, further research is needed to explore for ways to process the interaction information quickly and flexibly.

[Microbial Diversity and Community Analysis of Dental Plaques in Orthodontic Patients Wearing Invisible Appliances and Fixed Appliances].

Objective: To explore the microbial diversity and community structure of dental plaques in orthodontic patients with invisible appliances and fixed appliances and to study the differences. Methods: Ten orthodontic patients wearing invisible appliances (I) and ten wearing fixed appliances (F) were recruited. Dental plaques were collected from both buccal (B) and lingual (L) sides. Based on 16S rDNA, 40 dental plaque samples were analyzed after Illumina sequencing. Results: The microbial diversity, abundance and evenness of the FB group were significantly higher than those of the IB and IL groups (P<0.05), while the FL group showed substantial individual differences. The community structures were generally similar among the four groups, but significant differences in the relative abundance of some bacteria were found. The IB group showed higher abundances of Actinomycetes and Rosella (P<0.05), which were considered to be involved in dental caries and periodontal diseases. Some key communities showing significant differences were significantly enriched in the FB group, including Coprobacillus, Bifidobacterium, Enterobacterium, Lactobacillus, etc.. Conclusion: Dental plaques in patients wearing invisible appliances and fixed appliances showed significantly different microbial abundance, diversity and composition, which may be involved in orthodontic complications such as dental caries and periodontal diseases. Orthodontic patients need strengthened measures for oral hygiene maintenance, no matter what kind of appliances they wear.

[The Effect of PAMAM Dendrimer on the Dentin Remineralization and Matrix Metalloproteinases Activity].

OBJECTIVE: The purpose of this study was to investigate the effect of G4.5 carboxyl-terminated poly dendrimer (PAMAM-COOH) on the dentin remineralization and the matrix metalloproteinases (MMPs) activity. METHODS: The dentine samples were averagely divided into four groups: 100 mg/mL PAMAM-COOH group (A group), 10 mg/mL PAMAM-COOH group (B group), 2% (wt) chlorhexidine (CHX) group (C group) and deionized water group (Control group). MMP Activity Assay Kit was used to detect the activity of dentin endogenous MMPs in the four groups. The loss of dry mass of dentin after 30 d were measured. In situ zymography analysis was performed to detect the effects of PAMAM dendrimer in each group (except A group) on gelatinase activity in dentin. After incubation in artificial saliva for 7 and 14 d incubated, the remineralization of each group (except A group) in dentin surfaces were examined using a field emission-scanning electron microscope (FESEM). RESULTS: Compared with the control group, the dentin endogenous MMPs activity in A, B and C groups were all decreased ( P<0.05). The activity of endogenous MMPs in C group was lower than that of A and B groups ( P<0.001), but the difference between A and B groups was not statistically significant. The loss of dry mass in A, B and C groups were lower than that in control group ( P<0.05), but there were no significant difference in A, B and C groups. The in situzymography analysis showed that 48 h later, the dentin gelatinase activity in B group was inhibited compared with the control group, but the inhibitory effect was weaker than that of CHX. After 7 d and 14 d, there were no obvious mineralization in the control group, while distinct mineralization were observed in B group. The mineralization effect in group B was better than group C. CONCLUSION: G4.5 PAMAM-COOH could introduce remineralizationin and demineralizeddentin by effectively inhibiting endogenous MMPs and gelatinase, thus contributes as novel material to enhancing durability of adhesion.

[The Effect of Concentrate Growth Factors on the Survival, Proliferation and Differentiation of Human Dental Pulp Cells].

OBJECTIVE: To explore the cytotoxicity of concentrate growth factors (CGF) and the effects on the apoptosis, proliferation and differentiation of human dental pulp cells (hDPCs), which were closely correlated with future application of CGF in the treatment of dental pulpal and periapical diseases. METHODS: hDPCs were isolated from permanent teeth extracted for orthodontic purpose, and expanded in vitro. hDPCs were treated with CGF and mineral trioxide aggregate (MTA) respectively. The cell apoptosis, proliferation, cell cycle and ALP activity were analyzed after 1, 3 and 7 days. RESULTS: Compared with the MTA group, CGF significantly promoted cell proliferation, increased the proportion of S-phase cells and ALP activity on days 3 and 7 (P<0.01). Besides, hDPCs apoptotic rates decreased in CGF group. CONCLUSION: CGF has a good ability to promote the proliferation of dental pulp cells, resist apoptosis and induce osteogenic/odontogenic differentiation in vitro.

Knockout fth1b affects early mineralization of zebrafish pharyngeal teeth.

OBJECTIVES: A study was conducted to explore the expression pattern and function of ferritin heavy polypeptide gene (fth1b) in zebrafish pharyngeal teeth development and lay the foundation for subsequent research on teeth development and mineralization. METHODS: The zebrafish embryos were harvested at 56, 72, 96, and 120 h after fertilization. The expression of fth1b in zebrafish pharyngeal teeth development was detected by whole embryo in situ hybridization and compared with the known pharyngeal teeth marker dlx2b. The specific knockout of fth1b gene was performed using clustered regularly interspaced short palindromic repeats (CRISPR)/Cas9 gene editing technology. The development of zebrafish pharyngeal teeth was detected in the fth1b-/- mutant. RESULTS: The expression pattern of fth1b gene was very similar to that of the known zebrafish pharyngeal teeth marker dlx2b and was specifically expressed in the zebrafish pharyngeal teeth during development. After the specific knockout of the gene fth1b, the earliest gene that can be detect in zebrafish pharyngeal teeth-pitx2 was expressed normally during early development. The dlx2b expression was not significantly different from that of wild type zebrafish, but the mineralization of pharyngeal teeth in the mutant was weaker than that of wild type zebrafish. CONCLUSIONS: The gene fth1b is specifically expressed in zebrafish pharyngeal teeth and acts on their early mineralization.

[Research progress on the biomimetic remineralization of hard tooth tissues based on polyamide-amine dendrimer].

Polyamide-amine (PAMAM) dendrimer, a new hyperbranched macromolecular polymer, is considered an "artificial protein" by many scholars on account of its excellent chemical and biological characteristics. PAMAM has internal cavities and a large number of reactive terminal groups. These structures allow the polymer to be used as a bionic macromoleculethat could simulate the biomimetic mineralization of the natural organic matrix on the surface of tooth tissue. Specifically, PAMAM can beused as an organic template to regulate mineral nucleation and crystal growth; thus, the polymerisa more ideal dental restoration material than traditional allogenic materials. This article reviews research progress on thePAMAM-induced biomimetic mineralization of hard tooth tissues.

[Oral microbiological diversity in patients with salivary adenoid cystic carcinoma].

OBJECTIVE: The aim of this study was to identify the differences in microbial diversity and community in patients with salivary adenoid cystic carcinoma (SACC). METHODS: Saliva was collected from 13 patients with SACC confirmed by histopathological diagnosis and 10 healthy control subjects. Total metagenomic DNA was extracted. The DNA amplicons of the V3-V4 hypervariable regions of the 16S rRNA gene were generated and subjected to high-throughput sequencing. Microbial diversity and community structure were analyzed with Mothur software. RESULTS: A total of 16 genera of dominant bacteria in the SACC group were found, including Streptococcus (36.68%), Neisseria (8.55%), Prevotella_7 (7.53%), and Veillonella (6.37%), whereas 15 dominant bacteria in the control group were found, including Streptococcus (18.41%), Neisseria (18.20%), Prevotella_7 (8.89%), Porphyromonas (6.20%), Fusobacterium (5.86%) and Veillonella (5.82%). The statistically different phyla between the two groups were Firmicutes, Proteobacteria and Fusobacterium (P<0.05). The statistically different genera between the two groups were Streptococcus, Neisseria and Porphyromonas (P<0.05), and Capnocytophaga was only detected in patients with SACC. CONCLUSIONS: Significant differences were observed in the oral microorganisms between the two groups.

[Expression patterns of ectodysplasin and ectodysplasin receptor during early dental development in zebrafish].

OBJECTIVE: This study aims to study the expression patterns of ectodysplasin (EDA) and ectodysplasin receptor (EDAR) during the early development of zebrafish and provide a foundation for further research of the Eda signaling pathway in tooth development. METHODS: Total RNA was extracted from zebrafish embryos at 48 hours postfertilization (hpf) and then reverse transcribed for cDNA library generation. The corresponding RNA polymerase was selected for the synthesis of the digoxin-labeled antisense mRNA probe of zebrafish pharyngeal tooth specific marker dlx2b and Eda signaling-associated genes eda and edar in vitro. The three sequences were ligated into a pGEMT vector with a TA cloning kit, and polymerase chain reaction (PCR) was applied to linearize the plasmid. The resultant PCR sequences were used as templates for synthesizing Dig-labeled mRNA probe dlx2b, eda, and edar. Zebrafish embryos were collected at 36, 48, 56, 60, 72, and 84 hpf, then whole mount in situ hybridization was performed for the detection of eda and edar expression patterns. Then, their expression patterns at 72 hpf were compared with the expression pattern of dlx2b. RESULTS: The mRNA antisense probes of dlx2b, eda, and edar were successfully obtained. The positive signals of eda and edar were observed in zebrafish pharyngeal tooth region at 48-72 hpf and thus conform to the signals of dlx2b in the positive regions. CONCLUSIONS: The ligand eda and edar, which are associated with the Eda signaling pathway, are strongly expressed only at the pharyngeal tooth region in zebrafish from tooth initiation to the morphogenesis stage. Thus, the Eda signaling pathway may be involved in the regulation of the early development of zebrafish pharyngeal teeth.

[An in vitro study of the angiogenic effects of concentrate growth factor on human umbilical vein endothelial cells].

OBJECTIVE: This study aimed to explore the effects of concentrate growth factor extracts (CGFe) on human umbilical vein endothelial cells (HUVECs) in vitro. METHODS: Concentrate growth factor (CGF) were prepared from the peripheral blood of healthy donors, followed by CGFe. Four groups were designed based on cell culture medium, as follows: 2%CGFe, 5%CGFe, 10%CGFe, and control. The proliferation activity of HUVECs was detected by cell cycle and CCK-8 assays. The migration of HUVECs was detected by scratch assay. The mRNA expression levels of vascular endothelial growth factor (VEGF), chemokine receptor 4 (CXCR4), and platelet derived growth factor (PDGF) were examined by quantitative real time polymerase chain reaction (qRT-PCR). RESULTS: Results of CCK-8 and cell cycle assays showed that CGFe promoted the proliferation capability of HUVECs in a dose-dependent manner, and the data had statistical significance among four groups (P<
0.05). The cell migration assay indicated that CGF accelerated wound closure in a dose-dependent manner after 12 h of culture (P<0.05). The results of qRT-PCR showed that CGF upregulated the expression levels of VEGF, CXCR4, and PDGF in HUVECs. CONCLUSIONS: CGFe can promote the proliferation, migration, and angiogenic differentiation of HUVECs. Thus, CGF might be an appropriate cure for dental pulp revascularization.

[A study of genetic diversity in lactate dehydrogenase of Streptococcus mutans from clinical isolates].

OBJECTIVES: Lactate dehydrogenase (LDH) is shown to be an important virulence factor resulting in acid production of Streptococcus mutans (S. mutans), on which the cariogenic potential of S. mutans depends. Differences in cariogenic abilities of S. mutans isolates may be determined by genetic heterogeneity from virulent factors. The relationship between LDH activity or genetic diversity and cariogenicity of S. mutans (serotype c) isolates was studied in this research. METHODS: The genome DNA of S. mutans isolates were isolated and LDH gene (ldh) were amplified with specific primers. These isolates came from 34 caries-active individuals and 36 caries-free ones, in which 24 strains showed the high LDH activity and 21 strains showed the low LDH activity. Then genetic diversity of PCR products were analyzed or assessed by restriction fragment-length polymorphism (RFLP). Some of amplified LDH genes from different group were sequenced and assayed. RESULTS: It is testified that two genotypes A and B of ldh-RFLP were revealed when LDH genes were digested with Mse I, but Hph I, Mnl I, Dde I, Nla III and Alu I digesting fragments of Idh gene did not show different pattern. Furthermore, Fisher Exact one-Tail Test showed that the proportion of genotype B among strains from caries-free individuals was higher than that from caries-active ones (P = 0.033), while the distribution of genotypes with different LDH activity was different between two groups (two-Tail Test P = 0.017). The sequencing DNA testified that the specific base mutation would lead to multiple kind of genotype resulted. CONCLUSION: This study indicated that LDH gene of S. mutans clinical strains from different individuals is conservative, while there still is the gene mutation in. The ldh genetic diversity may be related to the low caries sensitivity, and closely correlated with the differences in LDH enzyme activity of S. mutans strains.

[Relationship of the genetic diversity within the V-region, P-region and C-terminus of surface protein of Streptococcus mutans to dental caries].

OBJECTIVE: To gain an understanding of the relationship between the genetic diversity within the V-region, P-region, C-terminus of surface protein of Streptococcus mutans (serotype c) and their adherent abilities. METHODS: The clinical isolates of S. mutans (serotype c) in two groups with different adherent abilities (cpm>2000, cpm<1000) had been prepared in our experimental laboratory. The genome DNA was extracted, and the spaP-pv (2060-3157 bp) and spaP-c (4003-4851 bp) were amplified by PCR, respectively. Genetic diversity was assessed by restriction fragment-length polymorphism (RFLP) with restriction endonucleases Alu I. RESULTS: (1) Two different patterns of spaP-pv RFLP among strains were revealed after the amplified product being digested with Alu I. The distributions of two genotypes (a and b) in the clinical strains with different adherent abilities differed significantly (P<0.05). The proportion of genotype b in the strains with higher adherent abilities was higher than that in the strains with lower adherent abilities. (2) Two different patterns (c and d) of spaP-c RFLP among strains were also revealed after the spaP-c being digested with Alu I. No statistically significant difference was observed in the distribution of the two genotypes in the clinical strains. CONCLUSION: The genetic diversity within spaP-pv of clinical isolates of S. mutans might be related to the differences of their adherent abilities.

[Construction of a red fluorescent shuttle vector controlled by recA operon promoter of Streptococcus mutans].

OBJECTIVE: To construct a red fluorescent shuttle vector controlled by recA operon promoter to transform Streptococcus mutans. METHODS: The promoter of recA was amplified from Streptococcus mutans UA159, and connected to plasmid pDsRed2-N1 to construct pRred with a red fluorescent coding gene, which was then inserted into the shuttle vector pDL276 to construct pLRred. RESULTS: pLRred was successfully constructed, and Escherichia coli transformed with the pLRred plasmid could express reporter gene DsRed. CONCLUSIONS: The recombination plasmid pLRred can be used in the further research of the expression of cariogenic virulence factor gene by Streptococcus mutans in biofilm.

V-ATPase subunit ATP6AP1 (Ac45) regulates osteoclast differentiation, extracellular acidification, lysosomal trafficking, and protease exocytosis in osteoclast-mediated bone resorption.

Lysosomal trafficking and protease exocytosis in osteoclasts are essential for ruffled border formation and bone resorption. Yet the mechanism underlying lysosomal trafficking and the related process of exocytosis remains largely unknown. We found ATP6ap1 (Ac45), an accessory subunit of vacuolar-type H(+)-ATPases (V-ATPases), to be highly induced by receptor activator for nuclear factor kappa B ligand (RANKL) in osteoclast differentiation. Ac45 knockdown osteoclasts formed normal actin rings, but had severely impaired extracellular acidification and bone resorption. Ac45 knockdown significantly reduced osteoclast formation. The decrease in the number of osteoclasts does not result from abnormal apoptosis; rather, it results from decreased osteoclast precursor cell proliferation and fusion, which may be partially due to the downregulation of extracellular signal-regulated kinase (ERK) phosphorylation and FBJ osteosarcoma oncogene (c-fos), nuclear factor of activated T-cells, cytoplasmic 1 (NFATc1), and "transmembrane 7 superfamily member 4" (Tm7sf4) expression. Notably, Ac45 knockdown osteoclasts exhibited impaired lysosomal trafficking and exocytosis, as indicated by the absence of lysosomal trafficking to the ruffled border and a lack of cathepsin K exocytosis into the resorption lacuna. Our data revealed that the impaired exocytosis is specifically due to Ac45 deficiency, and not the general consequence of a defective V-ATPase. Together, our results demonstrate the essential role of Ac45 in osteoclast-mediated extracellular acidification and protease exocytosis, as well as the ability of Ac45 to guide lysosomal intracellular trafficking to the ruffled border, potentially through its interaction with the small guanosine-5'-triphosphatase (GTPase) Rab7. Our work indicates that Ac45 may be a novel therapeutic target for osteolytic disease.

[Construction and expression of spap/A eukaryotic expression plasmid of Streptococcus mutans in mammalian cells].

PURPOSE: The study is aimed to evaluate the expression of recombinant plasmid pVAX1-spap/A of surface protein antigen A of Streptococcus mutans in mammalian cells COS-7. METHODS: The eukaryotic plasmid carrying encoding gene of spap/A of Streptococcus mutans was constructed and the plasmid introduced into COS-7 cells by lipofectamine reagent. The transient expressed protein was detected by immunochemistry technique in COS-7 cells. RESULTS: Positive expression was detected in plasma of the cells which were transfected with recombinant plasmid pVAX1-spap/A. The cells which were transfected with pVAX1 were negative. CONCLUSIONS: Spap/A can translate and express in COS-7 cells after transfected with recombinant plasmid pVAX1-spap/A. The expressed protein locates in the plasma and the protein is able to combine with anti-spap/A antibody. The expressed protein has the antigenicity and recombinant plasmid pVAX1-spap/A is a candidate vaccine.

[Construction of gtfB/CAT eukaryotic expression plasmid of Streptococcus mutans in mammalian cells].

PURPOSE: The study aimed to evaluate the expression of recombinant plasmid pVAX1- gtfB/CAT in mammalian COS-7 cells. METHODS: The eukaryotic plasmid carrying encoding gene of gtfB/CAT of Streptococcus mutans was constructed and introduced into COS-7 cells by lipofectamine reagent. The transient protein expression was detected by immunochemistry technique in COS-7 cells. RESULTS: The positive expression of gefB/CAT was detected in plasma of the cells which were transfected with recombinant plasmid pVAX1- gtfB/CAT. The cells which were transfected with pVAX1 were negative for gtfB/CAT expression. CONCLUSIONS: GtfB/CAT can be translated and expressed in COS-7 cells after transfected with recombinant plasmid pVAX1- gtfB/CAT. The expressed protein is located in the plasma and the protein is able to combine with anti- gtfB/CAT antibody. The expressed protein has the antigenicity and recombinant plasmid pVAX1- gtfB/CAT is a candidate vaccine.Supported by Key Research Project of Bureau of Education of Guizhou Province (Grant No.2004119).

[Dynamic study on saccharomyces albicans drug efflux pumps gene expression during the biofilm formation].

OBJECTIVE: To observe the drug resistance and drug efflux pumps gene mRNA of Saccharomyces albicans, including CDR1 gene and MDR1 gene, at different stage of biofilm formation in chemostat, furthermore to analysis the relationship between the drug efflux pump gene expression and the biofilm related drug resistance. METHODS: To form the mature biofilm in vitro in chemostat, then collect the biofilm strains at different development stages (2, 12, 24, 48 h) to semi-quantified mRNA amount of CDR1 gene and MDR1 gene by one step RT-PCR method. Using XTT reduction method to test the dynamic change of Saccharomyces albicans drug resistance in biofilm. RESULTS: Antifungal resistance of biofilm-grown cells increased conjunction with the biofilm maturation. Compared with earth stage of biofiom strains, the amount of CDR1 mRNA gene in mature biofilm strains increased, while MDR1 gene did not. CONCLUSION: There is positive correlation between drug resistance and biofilm maturation of Saccharomyces albicans. Biofilm related drug resistance appears to be partially associated with the upregulation of drug efflux pumps, although the variation is not shown coincidence. During the biofilm formation, CDR1 gene expression is actively up-regulated, but MDR1 gene expression is stable.

[Genetic diversity and mRNA expression of F-ATPase subunit uncG gene of streptococcus mutans clinical isolates].

OBJECTIVE: To study the genetic diversity and the gene expression of membrane-bound proton-translocating ATPase (F-ATPase) subunit gene uncG derived from Streptococcus mutans (S. mutans) clinical isolates. METHODS: 38 S. mutans strains derived from caries-active and caries-free individuals including 18 strains displaying high acid tolerance and 20 strains displaying low acid tolerance. Gene uncG was amplified with specific primers from S. mutans genomic DNA, then the PCR product was analyzed by RFLP and sequenced. The relative expression quantity of uncG gene against the housekeeping gene recA was determined by using RT-PCR method. A gel documentation system and QUANTITY ONES software were used to analyze the data results. RESULTS: It was testified that four genotypes A, B, C and D of PCR-RFLP were revealed when respectively digested with Alu I and Bsr I, but the distributions of the four genotype strains showed no difference (P > 0.05). The differences of uncG gene transcript quantities derived from different genotype or different aciduranc strains had no significance (P > 0.05). CONCLUSION: This study indicated that uncG gene of F-ATPase obviously displayed genetic diversity and existed polymorphism at mRNA expression level, while the Alu I-RFLP genotypes and the expression levels would not be responsive to different acid tolerance of S. mutans strains.

[Genetic diversity of F-ATPase subunits gene uncEBF amplified from Streptococcus mutans clinical isolates].

OBJECTIVE: The purpose of this research was to study the genetic diversity of F-ATPase subunit gene uncEBF derived from Streptococcus mutans (S. mutans) clinical isolates, furthermore to investigate the relationship between the genetic diversity of F-ATPase and S. mutans aciduric ability. METHODS: 38 S. mutans strains included 18 high acid tolerance strains and 20 low acid tolerance strains. Gene uncEBF of these isolates were amplified with specific primers from S. mutans genomic DNA, and the PCR products were analyzed by RFLP and sequenced. SPSS 11.0 statistic software assayed the results. RESULTS: It was testified that two genotypes A and B of PCR-RFLP were revealed when digested with Alu I and Dde I digested fragments of uncEBF displayed two different patterns C and D. Fisher exact two-tail test showed that the distributions of A and B genotype strains with different acidurance were different (P < 0.05), and the proportion of A genotype strains from high acidurance group was higher than that from low acidurance one. Some of these amplified uncEBF genes from different genotype were sequenced and testified that there existed variation of Alu I and Dde I recognized sites. CONCLUSION: This study indicated that uncEBF gene of S. mutans F-ATPase obviously exhibited genetic diversity.