Journal: The Scientific World Journal

Article Title: A New gcrR-Deficient Streptococcus mutans Mutant for Replacement Therapy of Dental Caries

doi: 10.1155/2013/460202

Figure Lengend Snippet: (a) To confirm that gcrR was deleted, chromosomal DNA was isolated from the selected transformants as well as UA159 progenitor and used as a template for PCR amplification with the P1/P2 and RT-gcrRF/RT-gcrRR primers. The resulting amplicons (the gcrR upstream-kanamycin resistance (KanR) cassette junction and the part of gcrR gene) were analyzed via agarose gel electrophoresis. Nucleotide sequencing of the purified amplicons with the same primer set was conducted to confirm the gcrR knockout and KanR cassette insertion. (b) Lane 1: the resulting amplicons (chromosomal DNA of MS-gcrR-def as template and RT-gcrRF/RT-gcrRR as primers); lane 2: the resulting amplicons (chromosomal DNA of UA159 as template and RT-gcrRF/RT-gcrRR as primers); lane 3: 1 kb marker (Fermentas). (c) Lanes 1, 2, 3, and 4: the resulting amplicons (chromosomal DNA of MS-gcrR-def as template and P1/P2 as primers); Lane 5, 1 kb marker (Fermentas); Lane 6, the resulting amplicons (chromosomal DNA of UA159 as template and P1/P2 as primers).

Article Snippet: Standard curves were generated for each primer-template set with a series of standard S. mutans UA159 genomic DNAs (ATCC, USA) that were diluted 10-fold.

Techniques: Isolation, Amplification, Agarose Gel Electrophoresis, Sequencing, Purification, Knock-Out, Marker

Journal: The Scientific World Journal

Article Title: A New gcrR-Deficient Streptococcus mutans Mutant for Replacement Therapy of Dental Caries

doi: 10.1155/2013/460202

Figure Lengend Snippet: The adhesion ability of Streptococcus mutans UA159 as well as the MS-gcrR-def was determined based on the absorbance of the cells on the plate. *Significant differences of adhesion between UA159 and MS-gcrR-def ( P < 0.05) were found, while there was no variance in cell growth.

Article Snippet: Standard curves were generated for each primer-template set with a series of standard S. mutans UA159 genomic DNAs (ATCC, USA) that were diluted 10-fold.

Techniques:

Journal: The Scientific World Journal

Article Title: A New gcrR-Deficient Streptococcus mutans Mutant for Replacement Therapy of Dental Caries

doi: 10.1155/2013/460202

Figure Lengend Snippet: (a) IOD (integrated optical density) of the green and red fluorescence from biofilm cells. (b) Biofilm thickness. The Z section was used to record the biofilm thickness. Vertical lines were chosen randomly to analyze each image. The red and green fluorescence intensities of the dead cells and living cells, respectively, were also quantified using Image Pro-plus. *Significant difference was found between green fluorescence of UA159 and MS-gcrR-def ( P < 0.05).

Article Snippet: Standard curves were generated for each primer-template set with a series of standard S. mutans UA159 genomic DNAs (ATCC, USA) that were diluted 10-fold.

Techniques: Fluorescence

Journal: The Scientific World Journal

Article Title: A New gcrR-Deficient Streptococcus mutans Mutant for Replacement Therapy of Dental Caries

doi: 10.1155/2013/460202

Figure Lengend Snippet: For in vitro acid production, the initial pH of the culture was measured at 7.3 by using a pH meter. Each sample was repeated three times. *Significant differences were found between UA159 and MS-gcrR-def ( P < 0.05).

Article Snippet: Standard curves were generated for each primer-template set with a series of standard S. mutans UA159 genomic DNAs (ATCC, USA) that were diluted 10-fold.

Techniques: In Vitro

Journal: The Scientific World Journal

Article Title: A New gcrR-Deficient Streptococcus mutans Mutant for Replacement Therapy of Dental Caries

doi: 10.1155/2013/460202

Figure Lengend Snippet: The enamel and dentinal caries scores of rats infected with UA159, MS-gcrR-def, and the mixture of both. The caries levels were scored by Keyes' method. The results are expressed as mean ± S.D. values. Symbols for statistical significance: *significantly different from the group UA159 infection alone ( P < 0.05).

Article Snippet: Standard curves were generated for each primer-template set with a series of standard S. mutans UA159 genomic DNAs (ATCC, USA) that were diluted 10-fold.

Techniques: Infection

Journal: Frontiers in Microbiology

Article Title: A GntR Family Transcription Factor in Streptococcus mutans Regulates Biofilm Formation and Expression of Multiple Sugar Transporter Genes

doi: 10.3389/fmicb.2018.03224

Figure Lengend Snippet: Deletion of stsR decreases S. mutans biofilm formation at early stage. S. mutans was cultured in BM supplemented with 1% sucrose for 6, 12, 24, and 48 h. The biofilm biomass was determined by CV staining method. Data from three biological replicates were averaged, and the statistical significance between the stsR mutant, wild-type, and complement strain was determined by Student’s t -test. Error bars represent standard deviations based on results from at least three biological replicates. ∗∗ Indicates a significance of P < 0.01.

Article Snippet: According to the National Center for Biotechnology Information (NCBI) S. mutans genome annotation, the function of the majority of the differentially expressed genes (DEGs) are unknown.

Techniques: Cell Culture, Staining, Mutagenesis

Journal: Frontiers in Microbiology

Article Title: A GntR Family Transcription Factor in Streptococcus mutans Regulates Biofilm Formation and Expression of Multiple Sugar Transporter Genes

doi: 10.3389/fmicb.2018.03224

Figure Lengend Snippet: Determination of growth curves of S. mutans UA159, S. mutans Δ stsR , and Δ stsR/pDL278-stsR . S. mutans UA159, S. mutans Δ stsR , and Δ stsR/pDL278-stsR strains were cultivated in BHI to mid-exponential phase and then diluted into (A) fresh BHI broth or (B) TV base medium supplemented with either 10 mM (limiting) or 100 mM (excess) sucrose, glucose, or lactose. Growth curves were monitored with a Multiskan Spectrum (Thermo, Multiskan Go, United States), and the OD 600 was measured in 1 h intervals. (A) For CFU counts, after diluted, the bacteria were cultured at 37°C for 1, 4, 8, and 12 h. Then bacterial suspension was serially diluted in BHI and plated on BHI agar plates. CFU values were calculated after the plates were incubated anaerobically at 37°C for 48 h. Error bars represent standard deviations based on results from at least three biological replicates.

Article Snippet: According to the National Center for Biotechnology Information (NCBI) S. mutans genome annotation, the function of the majority of the differentially expressed genes (DEGs) are unknown.

Techniques: Bacteria, Cell Culture, Suspension, Incubation

Journal: Frontiers in Microbiology

Article Title: A GntR Family Transcription Factor in Streptococcus mutans Regulates Biofilm Formation and Expression of Multiple Sugar Transporter Genes

doi: 10.3389/fmicb.2018.03224

Figure Lengend Snippet: Deletion of stsR decreases the amount of glucans in S. mutans biofilm. The amounts of (A) water insoluble glucans and (B) water soluble glucans in the biofilms of S. mutans UA159, S. mutans Δ stsR , and complement strain were quantified using the phenol-sulfuric acid method and calculated according to the standard curve. Error bars represent standard deviations based on results from at least three biological replicates. ∗∗ Indicates a significance of P < 0.01.

Article Snippet: According to the National Center for Biotechnology Information (NCBI) S. mutans genome annotation, the function of the majority of the differentially expressed genes (DEGs) are unknown.

Techniques:

Journal: Frontiers in Microbiology

Article Title: A GntR Family Transcription Factor in Streptococcus mutans Regulates Biofilm Formation and Expression of Multiple Sugar Transporter Genes

doi: 10.3389/fmicb.2018.03224

Figure Lengend Snippet: Scanning electron microscopy analysis reveals altered biofilm morphology and decreased biofilm extracellular matrix of S. mutans Δ stsR . Biofilms formed by S. mutans UA159, S. mutans Δ stsR , and complement strain were grown for 6 h and then scanned by scanning electron microscopy (SEM) under (A) 1000× magnification, (B) 5000× magnification, and (C) 20000× magnification.

Article Snippet: According to the National Center for Biotechnology Information (NCBI) S. mutans genome annotation, the function of the majority of the differentially expressed genes (DEGs) are unknown.

Techniques: Electron Microscopy

Journal: Frontiers in Microbiology

Article Title: A GntR Family Transcription Factor in Streptococcus mutans Regulates Biofilm Formation and Expression of Multiple Sugar Transporter Genes

doi: 10.3389/fmicb.2018.03224

Figure Lengend Snippet: Biofilm structure and EPS distribution of S. mutans strains observed by confocal microscopy. (A) Double-labeling of 6 h S. mutans biofilms. Green indicates bacteria (SYTO 9), and red indicates EPS (Alexa Fluor 647). Images were taken at 60× magnification. The three-dimensional reconstruction of the biofilms and the quantification of EPS/bacteria biomass were performed with IMARIS 7.0.0. (B) The ratio of EPS to bacteria at different heights was quantified with COMSTAT. Results are the average of five randomly selected positions of each sample and are presented as mean ± standard deviation. (C–E) Quantification of S. mutans UA159 (C) , S. mutans Δ stsR (D) , and Δ stsR/pDL278-stsR (E) biofilms. EPS biomass was performed with COMSTAT at different heights. Results are the average of five randomly selected positions of each sample and are presented as mean ± standard deviation.

Article Snippet: According to the National Center for Biotechnology Information (NCBI) S. mutans genome annotation, the function of the majority of the differentially expressed genes (DEGs) are unknown.

Techniques: Confocal Microscopy, Labeling, Bacteria, Standard Deviation

Journal: Frontiers in Microbiology

Article Title: A GntR Family Transcription Factor in Streptococcus mutans Regulates Biofilm Formation and Expression of Multiple Sugar Transporter Genes

doi: 10.3389/fmicb.2018.03224

Figure Lengend Snippet: Quantitative RT-PCR assays for the relative expression levels of gtfs and ftf genes in S. mutans UA159, S. mutans Δ stsR , and Δ stsR/pDL278-stsR . The experiments were carried out as described in Experimental procedures. All target genes were amplified using specific primers. Different gene expressions were normalized to the levels of 16S rRNA gene transcripts, and the folds of expression change were calculated.

Article Snippet: According to the National Center for Biotechnology Information (NCBI) S. mutans genome annotation, the function of the majority of the differentially expressed genes (DEGs) are unknown.

Techniques: Quantitative RT-PCR, Expressing, Amplification

Journal: Frontiers in Microbiology

Article Title: A GntR Family Transcription Factor in Streptococcus mutans Regulates Biofilm Formation and Expression of Multiple Sugar Transporter Genes

doi: 10.3389/fmicb.2018.03224

Figure Lengend Snippet: The differentially expressed sugar transport system operons in S. mutans Δ stsR . The genetic organization of differentially expressed gene clusters that were associated with sugar transport systems in S. mutans Δ stsR . Upregulated genes are colored red, and downregulated genes are colored green.

Article Snippet: According to the National Center for Biotechnology Information (NCBI) S. mutans genome annotation, the function of the majority of the differentially expressed genes (DEGs) are unknown.

Techniques:

Journal: Applied and Environmental Microbiology

Article Title: Raffinose Induces Biofilm Formation by Streptococcus mutans in Low Concentrations of Sucrose by Increasing Production of Extracellular DNA and Fructan

doi: 10.1128/AEM.00869-17

Figure Lengend Snippet: The function of the polysaccharide synthase genes in biofilm formation in TSBr. (A) The amount of biofilm formed by the deletion mutants of the gene encoding polysaccharide synthase in TSBr. (B) CLSM analysis of the biofilm formed in TSBr. The cells in the biofilm were stained with the LIVE/DEAD BacLight bacterial viability kit for 30 min. Representative images from three independent experiments are presented. (C) An aggregation assay of the bacterial cells. The roles of eDNA and fructan in cell aggregation were examined by mimicking the extracellular matrix synthesized in TSBr. Five micrograms per milliliter genomic DNA of UA159 and/or 250 μg/ml commercial inulin was added to 3 ml of cell suspension, which was grown in BHI and adjusted to an OD600 of 1.5 with aggregation buffer. The data are presented as the means ± SD of the results from three independent experiments. The asterisks indicate a significant difference between two groups (Student's t test, P < 0.05).

Article Snippet: We used genomic DNA isolated from S. mutans UA159 using the DNeasy blood and tissue kit (Qiagen, Venlo, The Netherlands) and inulin (Nacalai Tesque, Kyoto, Japan) to mimic the main components of the biofilm matrix formed in TSBr.

Techniques: Staining, Synthesized, Suspension

Journal: Infection and Immunity

Article Title: The Collagen Binding Protein Cnm Contributes to Oral Colonization and Cariogenicity of Streptococcus mutans OMZ175

doi: 10.1128/IAI.03022-14

Figure Lengend Snippet: Streptococcus mutans strains used in this study

Article Snippet: The cariogenic potentials of OMZ175, OMZ175Δ cnm , UA159, UA159- cnm , and UA159-pBGE were assessed in Sprague-Dawley rats by methods described elsewhere ( 36 , 37 ).

Techniques: Expressing

Journal: Infection and Immunity

Article Title: The Collagen Binding Protein Cnm Contributes to Oral Colonization and Cariogenicity of Streptococcus mutans OMZ175

doi: 10.1128/IAI.03022-14

Figure Lengend Snippet: Attachment to, and invasion of, human gingival fibroblasts (HGF-1) by OMZ175 (Cnm+), OMZ175Δcnm (cnm knockout), or UA159 (Cnm−). (A and B) Antibiotic protection assay to assess S. mutans attachment to, and invasion of, human gingival fibroblasts (A) and human oral keratinocytes (B). Medians and interquartile ranges for eight replicates were normalized to the average results for OMZ175 infections. Asterisks indicate significant differences (P < 0.05). (C) Transmission electron microscopy of HGF-1 cells coincubated with OMZ175 for 2 h revealed S. mutans bacteria attached to HGF-1 cells, enveloped by lamellipodium-like extensions (black arrows), and intracellular S. mutans bacteria within membrane-bound vacuoles (white arrows). (D) Detail of transmission electron micrograph. Projections from the surfaces of OMZ175 cells were visible (black arrows) and appeared to be in contact with the HGF-1 vesicle wall (white arrows).

Article Snippet: The cariogenic potentials of OMZ175, OMZ175Δ cnm , UA159, UA159- cnm , and UA159-pBGE were assessed in Sprague-Dawley rats by methods described elsewhere ( 36 , 37 ).

Techniques: Knock-Out, Transmission Assay, Electron Microscopy, Bacteria, Membrane