gingivalis omz314  (ATCC)


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    Structured Review

    ATCC gingivalis omz314
    Adhesion/invasion of Ca9-22 cells by P . gulae D049 as compared with P . <t>gingivalis</t> strains. Antibiotic protection invasion assay of P . gulae D049, and P . gingivalis 33277 and <t>OMZ314.</t> Ca9-22 cells were infected with bacteria at an MOI of 100 for 90 min. The numbers of adherent and/or intracellular bacteria were determined by counting viable cell lysates and are expressed as percentage of input bacterial cell number. Values are shown as the mean ± SD of three independent experiments and were analyzed with a t test. * P
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    Images

    1) Product Images from "Adhesion and invasion of gingival epithelial cells by Porphyromonas gulae"

    Article Title: Adhesion and invasion of gingival epithelial cells by Porphyromonas gulae

    Journal: PLoS ONE

    doi: 10.1371/journal.pone.0213309

    Adhesion/invasion of Ca9-22 cells by P . gulae D049 as compared with P . gingivalis strains. Antibiotic protection invasion assay of P . gulae D049, and P . gingivalis 33277 and OMZ314. Ca9-22 cells were infected with bacteria at an MOI of 100 for 90 min. The numbers of adherent and/or intracellular bacteria were determined by counting viable cell lysates and are expressed as percentage of input bacterial cell number. Values are shown as the mean ± SD of three independent experiments and were analyzed with a t test. * P
    Figure Legend Snippet: Adhesion/invasion of Ca9-22 cells by P . gulae D049 as compared with P . gingivalis strains. Antibiotic protection invasion assay of P . gulae D049, and P . gingivalis 33277 and OMZ314. Ca9-22 cells were infected with bacteria at an MOI of 100 for 90 min. The numbers of adherent and/or intracellular bacteria were determined by counting viable cell lysates and are expressed as percentage of input bacterial cell number. Values are shown as the mean ± SD of three independent experiments and were analyzed with a t test. * P

    Techniques Used: Invasion Assay, Infection

    2) Product Images from "Impact of Porphyromonas gingivalis Peptidylarginine Deiminase on Bacterial Biofilm Formation, Epithelial Cell Invasion, and Epithelial Cell Transcriptional Landscape"

    Article Title: Impact of Porphyromonas gingivalis Peptidylarginine Deiminase on Bacterial Biofilm Formation, Epithelial Cell Invasion, and Epithelial Cell Transcriptional Landscape

    Journal: Scientific Reports

    doi: 10.1038/s41598-018-32603-y

    Identification and functional annotation of TIGK genes significantly dependent on PPAD activity. ( A ) Scatterplot of log2 fold-change (log2FC) in gene expression of TIGKs infected with P. gingivalis WT vs. control (y-axis), and P. gingivalis PPAD C351A vs. P. gingivalis WT (x-axis). Genes altered exclusively in the P. gingivalis WT vs. control comparison are displayed in red when significantly up-regulated (log2FC > 0.5, FDR q
    Figure Legend Snippet: Identification and functional annotation of TIGK genes significantly dependent on PPAD activity. ( A ) Scatterplot of log2 fold-change (log2FC) in gene expression of TIGKs infected with P. gingivalis WT vs. control (y-axis), and P. gingivalis PPAD C351A vs. P. gingivalis WT (x-axis). Genes altered exclusively in the P. gingivalis WT vs. control comparison are displayed in red when significantly up-regulated (log2FC > 0.5, FDR q

    Techniques Used: Functional Assay, Activity Assay, Expressing, Infection

    The effect of P. gingivalis PPAD on bacterial abundance ( a ), species composition ( b ) and ( c ) the biofilm structure in multispecies biofilms. A biofilm consisting of T. forsythia , F. nucleatum , A. naeslundii , S. gordonii , and one of three P. gingivalis strains, wild type (WT), ppad deletion strain ( Δppad ), or a strain harboring inactivated PPAD (C351A), was cultured for 48 h, following which ( a,b ) the bacterial DNA was extracted and assessed via qPCR, ( c ) biofilm was fixed and observed via SEM under 1500x magnification. The ( a ) bacterial load and ( b ) species composition of each biofilm model are presented as mean ± SD from three independent experiments. Data were plotted on a logarithmic scale.
    Figure Legend Snippet: The effect of P. gingivalis PPAD on bacterial abundance ( a ), species composition ( b ) and ( c ) the biofilm structure in multispecies biofilms. A biofilm consisting of T. forsythia , F. nucleatum , A. naeslundii , S. gordonii , and one of three P. gingivalis strains, wild type (WT), ppad deletion strain ( Δppad ), or a strain harboring inactivated PPAD (C351A), was cultured for 48 h, following which ( a,b ) the bacterial DNA was extracted and assessed via qPCR, ( c ) biofilm was fixed and observed via SEM under 1500x magnification. The ( a ) bacterial load and ( b ) species composition of each biofilm model are presented as mean ± SD from three independent experiments. Data were plotted on a logarithmic scale.

    Techniques Used: Cell Culture, Real-time Polymerase Chain Reaction

    3) Product Images from "Identification of the Linkage between A-Polysaccharide and the Core in the A-Lipopolysaccharide of Porphyromonas gingivalis W50"

    Article Title: Identification of the Linkage between A-Polysaccharide and the Core in the A-Lipopolysaccharide of Porphyromonas gingivalis W50

    Journal: Journal of Bacteriology

    doi: 10.1128/JB.02562-14

    Genomic organization of the PG0129 locus in P. gingivalis W83 (PG_0242 in ATCC 33277). *, Orf56 is not annotated in the J. Craig Venter Institute sequence; **, gene marked as hypothetical.
    Figure Legend Snippet: Genomic organization of the PG0129 locus in P. gingivalis W83 (PG_0242 in ATCC 33277). *, Orf56 is not annotated in the J. Craig Venter Institute sequence; **, gene marked as hypothetical.

    Techniques Used: Sequencing

    4) Product Images from "RgpA-Kgp Peptide-Based Immunogens Provide Protection against Porphyromonas gingivalis Challenge in a Murine Lesion Model"

    Article Title: RgpA-Kgp Peptide-Based Immunogens Provide Protection against Porphyromonas gingivalis Challenge in a Murine Lesion Model

    Journal: Infection and Immunity

    doi:

    Maximum lesion size of mice challenged with either  P. gingivalis  ATCC 33277 (A) or W50 (B). BALB/c mice were immunized s.c. with the RgpA-Kgp proteinase complex (50 μg), formalin-killed (FK)  P. gingivalis  cells (2 × 10 8  of either strain ATCC 33277 or strain W50), formalin-killed  E. coli  cells (2 × 10 8 ), or PBS administered in IFA for both the primary and secondary doses. All mice were challenged 12 days after the secondary immunization with either  P. gingivalis  strain ATCC 33277 (7.5 × 10 9  viable cells) or W50 (3 × 10 9  viable cells) and were weighed, and the lesion sizes were measured daily over 14 days. Lesion sizes were statistically analyzed using Mann-Whitney U-Wilcoxon rank sum test. ∗, Groups significantly different ( P
    Figure Legend Snippet: Maximum lesion size of mice challenged with either P. gingivalis ATCC 33277 (A) or W50 (B). BALB/c mice were immunized s.c. with the RgpA-Kgp proteinase complex (50 μg), formalin-killed (FK) P. gingivalis cells (2 × 10 8 of either strain ATCC 33277 or strain W50), formalin-killed E. coli cells (2 × 10 8 ), or PBS administered in IFA for both the primary and secondary doses. All mice were challenged 12 days after the secondary immunization with either P. gingivalis strain ATCC 33277 (7.5 × 10 9 viable cells) or W50 (3 × 10 9 viable cells) and were weighed, and the lesion sizes were measured daily over 14 days. Lesion sizes were statistically analyzed using Mann-Whitney U-Wilcoxon rank sum test. ∗, Groups significantly different ( P

    Techniques Used: Mouse Assay, Immunofluorescence, MANN-WHITNEY

    5) Product Images from "RgpA-Kgp Peptide-Based Immunogens Provide Protection against Porphyromonas gingivalis Challenge in a Murine Lesion Model"

    Article Title: RgpA-Kgp Peptide-Based Immunogens Provide Protection against Porphyromonas gingivalis Challenge in a Murine Lesion Model

    Journal: Infection and Immunity

    doi:

    Schematic representation of the RgpA and Kgp polyproteins showing the location and amino acid sequence of the synthesized peptides. Amino acid numbering is from the initial Met of the polyprotein. The downward arrows indicate the processing site for the proteinase and adhesin domains. The numbers above the arrows indicate the N-terminal residues of that domain. Similarly hatched areas at other locations correspond to repeat sequences (sometimes with substitutions) of the same motif.
    Figure Legend Snippet: Schematic representation of the RgpA and Kgp polyproteins showing the location and amino acid sequence of the synthesized peptides. Amino acid numbering is from the initial Met of the polyprotein. The downward arrows indicate the processing site for the proteinase and adhesin domains. The numbers above the arrows indicate the N-terminal residues of that domain. Similarly hatched areas at other locations correspond to repeat sequences (sometimes with substitutions) of the same motif.

    Techniques Used: Sequencing, Synthesized

    Maximum lesion size of mice challenged with either P. gingivalis ATCC 33277 (A) or W50 (B). BALB/c mice were immunized s.c. with the RgpA-Kgp proteinase complex (50 μg), formalin-killed (FK) P. gingivalis cells (2 × 10 8 of either strain ATCC 33277 or strain W50), formalin-killed E. coli cells (2 × 10 8 ), or PBS administered in IFA for both the primary and secondary doses. All mice were challenged 12 days after the secondary immunization with either P. gingivalis strain ATCC 33277 (7.5 × 10 9 viable cells) or W50 (3 × 10 9 viable cells) and were weighed, and the lesion sizes were measured daily over 14 days. Lesion sizes were statistically analyzed using Mann-Whitney U-Wilcoxon rank sum test. ∗, Groups significantly different ( P
    Figure Legend Snippet: Maximum lesion size of mice challenged with either P. gingivalis ATCC 33277 (A) or W50 (B). BALB/c mice were immunized s.c. with the RgpA-Kgp proteinase complex (50 μg), formalin-killed (FK) P. gingivalis cells (2 × 10 8 of either strain ATCC 33277 or strain W50), formalin-killed E. coli cells (2 × 10 8 ), or PBS administered in IFA for both the primary and secondary doses. All mice were challenged 12 days after the secondary immunization with either P. gingivalis strain ATCC 33277 (7.5 × 10 9 viable cells) or W50 (3 × 10 9 viable cells) and were weighed, and the lesion sizes were measured daily over 14 days. Lesion sizes were statistically analyzed using Mann-Whitney U-Wilcoxon rank sum test. ∗, Groups significantly different ( P

    Techniques Used: Mouse Assay, Immunofluorescence, MANN-WHITNEY

    Immunoblot analysis of the RgpA-Kgp complex using murine anti-RgpA-Kgp complex sera. The RgpA-Kgp complex was separated by SDS-PAGE and transferred onto PVDF membrane. The RgpA-Kgp complex was probed with sera (1:50 TN buffer) from mice immunized with the RgpA-Kgp proteinase complex. Molecular mass markers are indicated in kilodaltons.
    Figure Legend Snippet: Immunoblot analysis of the RgpA-Kgp complex using murine anti-RgpA-Kgp complex sera. The RgpA-Kgp complex was separated by SDS-PAGE and transferred onto PVDF membrane. The RgpA-Kgp complex was probed with sera (1:50 TN buffer) from mice immunized with the RgpA-Kgp proteinase complex. Molecular mass markers are indicated in kilodaltons.

    Techniques Used: SDS Page, Mouse Assay

    Epitope mapping analysis of the RgpA27 adhesin protein of the RgpA-Kgp complex. Twenty-one overlapping 13mer peptides (overlay by six and offset by seven residues) were synthesized covering the N-terminal 148 residues of the RgpA27 by Chiron Technologies using the multipin peptide synthesis system. Epitope mapping of the pin-bound peptides was carried out by ELISA according to Chiron Technologies instructions. Pin-bound peptides were probed with sera from mice immunized with the RgpA-Kgp proteinase complex (■, n = 10) and normal mouse sera (□, n = 10).
    Figure Legend Snippet: Epitope mapping analysis of the RgpA27 adhesin protein of the RgpA-Kgp complex. Twenty-one overlapping 13mer peptides (overlay by six and offset by seven residues) were synthesized covering the N-terminal 148 residues of the RgpA27 by Chiron Technologies using the multipin peptide synthesis system. Epitope mapping of the pin-bound peptides was carried out by ELISA according to Chiron Technologies instructions. Pin-bound peptides were probed with sera from mice immunized with the RgpA-Kgp proteinase complex (■, n = 10) and normal mouse sera (□, n = 10).

    Techniques Used: Synthesized, Enzyme-linked Immunosorbent Assay, Mouse Assay

    6) Product Images from "Identification of a Diguanylate Cyclase and Its Role in Porphyromonas gingivalis Virulence"

    Article Title: Identification of a Diguanylate Cyclase and Its Role in Porphyromonas gingivalis Virulence

    Journal: Infection and Immunity

    doi: 10.1128/IAI.00084-14

    Comparison of the growth curves and gene expression of P. gingivalis ATCC 33277 and its mutant 1932E. (A) P. gingivalis cells were grown in standard TSB for 64 h. Bacterial growth is indicated by the means of the optical densities (OD 600 s) of the bacterial cultures. Error bars represent SDs ( n = 4 experiments). (B) The expression levels of the gene ( pgn_1931 ) located immediately downstream of pgn_1932 were measured by real-time RT-PCR. Each bar represents relative gene expression in the pgn_1932 mutant compared to that in P. gingivalis ATCC 33277 (which was given a value of 1). Error bars represent standard deviations ( n = 3).
    Figure Legend Snippet: Comparison of the growth curves and gene expression of P. gingivalis ATCC 33277 and its mutant 1932E. (A) P. gingivalis cells were grown in standard TSB for 64 h. Bacterial growth is indicated by the means of the optical densities (OD 600 s) of the bacterial cultures. Error bars represent SDs ( n = 4 experiments). (B) The expression levels of the gene ( pgn_1931 ) located immediately downstream of pgn_1932 were measured by real-time RT-PCR. Each bar represents relative gene expression in the pgn_1932 mutant compared to that in P. gingivalis ATCC 33277 (which was given a value of 1). Error bars represent standard deviations ( n = 3).

    Techniques Used: Expressing, Mutagenesis, Quantitative RT-PCR

    7) Product Images from "Oral Community Interactions of Filifactor alocis In Vitro"

    Article Title: Oral Community Interactions of Filifactor alocis In Vitro

    Journal: PLoS ONE

    doi: 10.1371/journal.pone.0076271

    Dual-species community formation between F. alocis and A. actinomycetemcomitans analyzed by CLSM. A. A. actinomycetemcomitans 652 (red, stained with hexidium iodide) was cultured on glass coverslips. F. alocis strains ATCC 35896 (upper left panel) and D-62D (upper right panel) were stained with FITC (green) and reacted with A. actinomycetemcomitans for 24 h, 48 h and 72 h. B. Time-resolved changes in the biovolume of A. actinomycetemcomitans 652, F. alocis ATCC 35896 and D-62D in dual species communities. Data are representative of four independent replicates. P-value compared with control single species communities was calculated by t-test, and significant differences are at p
    Figure Legend Snippet: Dual-species community formation between F. alocis and A. actinomycetemcomitans analyzed by CLSM. A. A. actinomycetemcomitans 652 (red, stained with hexidium iodide) was cultured on glass coverslips. F. alocis strains ATCC 35896 (upper left panel) and D-62D (upper right panel) were stained with FITC (green) and reacted with A. actinomycetemcomitans for 24 h, 48 h and 72 h. B. Time-resolved changes in the biovolume of A. actinomycetemcomitans 652, F. alocis ATCC 35896 and D-62D in dual species communities. Data are representative of four independent replicates. P-value compared with control single species communities was calculated by t-test, and significant differences are at p

    Techniques Used: Confocal Laser Scanning Microscopy, Staining, Cell Culture

    CLSM projections of monospecies communities of F.alocis strains ATCC 35896 and D-62D (green, stained with FITC), S. gordonii DL-1, F. nucleatum ATCC25586, A. actinomycetemcomitans 652, or P. gingivalis ATCC33277 (red, stained with hexidium iodide) after 24 h, 48 h, and 72 h.
    Figure Legend Snippet: CLSM projections of monospecies communities of F.alocis strains ATCC 35896 and D-62D (green, stained with FITC), S. gordonii DL-1, F. nucleatum ATCC25586, A. actinomycetemcomitans 652, or P. gingivalis ATCC33277 (red, stained with hexidium iodide) after 24 h, 48 h, and 72 h.

    Techniques Used: Confocal Laser Scanning Microscopy, Staining

    8) Product Images from "Oral Community Interactions of Filifactor alocis In Vitro"

    Article Title: Oral Community Interactions of Filifactor alocis In Vitro

    Journal: PLoS ONE

    doi: 10.1371/journal.pone.0076271

    Role of P. gingivalis LuxS in dual-species community formation with F. alocis . A. P. gingivalis ATCC 33277 (WT), and Δ luxS (1 × 10 8 , blue, stained with DAPI) were cultured on glass coverslips. F. alocis strains ATCC 35896 and D-62D were stained with FITC (green) and reacted with the P. gingivalis strains for 72 h. B. Biovolume of P. gingivalis or F. alocis in dual species communities at 72 h. Data are representative of four independent replicates. P-value compared with control single species communities was calculated by t-test, and significant differences are p
    Figure Legend Snippet: Role of P. gingivalis LuxS in dual-species community formation with F. alocis . A. P. gingivalis ATCC 33277 (WT), and Δ luxS (1 × 10 8 , blue, stained with DAPI) were cultured on glass coverslips. F. alocis strains ATCC 35896 and D-62D were stained with FITC (green) and reacted with the P. gingivalis strains for 72 h. B. Biovolume of P. gingivalis or F. alocis in dual species communities at 72 h. Data are representative of four independent replicates. P-value compared with control single species communities was calculated by t-test, and significant differences are p

    Techniques Used: Staining, Cell Culture

    9) Product Images from "Identification of an O-antigen chain length regulator, WzzP, in Porphyromonas gingivalis"

    Article Title: Identification of an O-antigen chain length regulator, WzzP, in Porphyromonas gingivalis

    Journal: MicrobiologyOpen

    doi: 10.1002/mbo3.84

    Analysis of LPS to assess Wzz activity by heterologous complementation. Silver-stained polyacrylamide gel displaying the O-antigen LPS profiles of Escherichia coli W3110 (lane 1), EVV16 ( wzzB ) containing pMF19 (lane 2), EVV16/pMF19 containing pBAD vector control (lane 3), EVV16/pMF19 containing PGN_2005-expressing plasmid from Porphyromonas gingivalis (lane 4), EVV16/pMF19 containing WzzB-expressing plasmid (pWzzB-SF) from Shigella flexneri (lane 5), and EVV16/pMF19 containing WzzB-expressing plasmid (pWzzB-ST) from Salmonella typhimurium (lane 6) (A). Immunoblot analysis of the cell lysates was performed with anti-PGN_2005 mouse polyclonal antiserum to confirm the expression of the PGN_2005 protein in the E. coli EVV16 (pMF19) strain (B).
    Figure Legend Snippet: Analysis of LPS to assess Wzz activity by heterologous complementation. Silver-stained polyacrylamide gel displaying the O-antigen LPS profiles of Escherichia coli W3110 (lane 1), EVV16 ( wzzB ) containing pMF19 (lane 2), EVV16/pMF19 containing pBAD vector control (lane 3), EVV16/pMF19 containing PGN_2005-expressing plasmid from Porphyromonas gingivalis (lane 4), EVV16/pMF19 containing WzzB-expressing plasmid (pWzzB-SF) from Shigella flexneri (lane 5), and EVV16/pMF19 containing WzzB-expressing plasmid (pWzzB-ST) from Salmonella typhimurium (lane 6) (A). Immunoblot analysis of the cell lysates was performed with anti-PGN_2005 mouse polyclonal antiserum to confirm the expression of the PGN_2005 protein in the E. coli EVV16 (pMF19) strain (B).

    Techniques Used: Activity Assay, Staining, Plasmid Preparation, Expressing

    Localization of the PGN_2005 protein in Porphyromonas gingivalis . The cell lysates of the wild type and the PGN_2005 mutant were subjected to immunodetection with anti-PGN_2005 (A). Cell fractionation analysis from the wild type. W, C/P, and TM indicate the whole cell lysate, cytoplasm/periplasm, and total membrane fraction, respectively (B). Five micrograms of protein from the inner membrane (IM) or outer membrane (OM) fractions that were separated by sucrose density gradient centrifugation from the membrane fraction of the wild type were subjected to immunodetection with anti-PGN_2005, anti-HBP35, anti-Rgp, mAb 1B5, and mAb TDC-5-2-1 (C).
    Figure Legend Snippet: Localization of the PGN_2005 protein in Porphyromonas gingivalis . The cell lysates of the wild type and the PGN_2005 mutant were subjected to immunodetection with anti-PGN_2005 (A). Cell fractionation analysis from the wild type. W, C/P, and TM indicate the whole cell lysate, cytoplasm/periplasm, and total membrane fraction, respectively (B). Five micrograms of protein from the inner membrane (IM) or outer membrane (OM) fractions that were separated by sucrose density gradient centrifugation from the membrane fraction of the wild type were subjected to immunodetection with anti-PGN_2005, anti-HBP35, anti-Rgp, mAb 1B5, and mAb TDC-5-2-1 (C).

    Techniques Used: Mutagenesis, Immunodetection, Cell Fractionation, Gradient Centrifugation

    Physical map of the area around the PGN_2005 gene and pigmentation of PGN_2005 mutant. Physical map of the PGN_2005 gene region (A). A triangle indicates the Tn 4400 ' insertion site of the PGN_2005 insertion mutant. Colony pigmentation (B). Porphyromonas gingivalis cells were anaerobically grown on blood agar plates at 35°C for 2 days.
    Figure Legend Snippet: Physical map of the area around the PGN_2005 gene and pigmentation of PGN_2005 mutant. Physical map of the PGN_2005 gene region (A). A triangle indicates the Tn 4400 ' insertion site of the PGN_2005 insertion mutant. Colony pigmentation (B). Porphyromonas gingivalis cells were anaerobically grown on blood agar plates at 35°C for 2 days.

    Techniques Used: Mutagenesis

    Purified LPS from the wild type or the PGN_2005 mutant. The LPS fraction was purified by the hot phenol method and subjected to SDS-PAGE followed by silver staining. Immunoblot analyses were also performed with mAb 1B5 and mAb TDC-5-2-1.
    Figure Legend Snippet: Purified LPS from the wild type or the PGN_2005 mutant. The LPS fraction was purified by the hot phenol method and subjected to SDS-PAGE followed by silver staining. Immunoblot analyses were also performed with mAb 1B5 and mAb TDC-5-2-1.

    Techniques Used: Purification, Mutagenesis, SDS Page, Silver Staining

    Immunoblot analyses of various Porphyromonas gingivalis strains. Immunoblot analyses of cell lysates of various P. gingivalis strains were performed with mAb 1B5, mAb TDC-5-2-1, anti-HBP35, or anti-Rgp. Three sets of PGN_2005/PGN_2005+ or PGN_2005 porT/PGN_2005+ strains were obtained from each single clone. The asterisks indicate nonspecific cross-reactive bands.
    Figure Legend Snippet: Immunoblot analyses of various Porphyromonas gingivalis strains. Immunoblot analyses of cell lysates of various P. gingivalis strains were performed with mAb 1B5, mAb TDC-5-2-1, anti-HBP35, or anti-Rgp. Three sets of PGN_2005/PGN_2005+ or PGN_2005 porT/PGN_2005+ strains were obtained from each single clone. The asterisks indicate nonspecific cross-reactive bands.

    Techniques Used:

    Gingipain and hemagglutination activities of Porphyromonas gingivalis . Porphyromonas gingivalis cells were anaerobically grown in enriched BHI medium at 35°C. The Kgp and Rgp activities of the cell lysates (cell) and vesicle-containing culture supernatants (sup) of ATCC 33277 (wild type), PGN_2005, or PGN_2005/PGN2005+ were measured (A). The hemagglutination activities of various P. gingivalis strains were measured (B). Twofold serial dilutions of various P. gingivalis cells were mixed with 1% sheep red blood cells and stored for 3 h at room temperature.
    Figure Legend Snippet: Gingipain and hemagglutination activities of Porphyromonas gingivalis . Porphyromonas gingivalis cells were anaerobically grown in enriched BHI medium at 35°C. The Kgp and Rgp activities of the cell lysates (cell) and vesicle-containing culture supernatants (sup) of ATCC 33277 (wild type), PGN_2005, or PGN_2005/PGN2005+ were measured (A). The hemagglutination activities of various P. gingivalis strains were measured (B). Twofold serial dilutions of various P. gingivalis cells were mixed with 1% sheep red blood cells and stored for 3 h at room temperature.

    Techniques Used:

    10) Product Images from "CRISPR Regulation of Intraspecies Diversification by Limiting IS Transposition and Intercellular Recombination"

    Article Title: CRISPR Regulation of Intraspecies Diversification by Limiting IS Transposition and Intercellular Recombination

    Journal: Genome Biology and Evolution

    doi: 10.1093/gbe/evt075

    Clustering by spacer content in CRISPR type 36.2 of Porphyromonas gingivalis . In type 36.2, the presence of each unique spacer is shown using a heatmap. The dendrogram was constructed from Euclidian distances. In the heatmap, the boxes indicate unique spacers and are arrayed horizontally. In the heatmap, 2 colors were used according to the bit score; red: ≥50, yellow:
    Figure Legend Snippet: Clustering by spacer content in CRISPR type 36.2 of Porphyromonas gingivalis . In type 36.2, the presence of each unique spacer is shown using a heatmap. The dendrogram was constructed from Euclidian distances. In the heatmap, the boxes indicate unique spacers and are arrayed horizontally. In the heatmap, 2 colors were used according to the bit score; red: ≥50, yellow:

    Techniques Used: CRISPR, Construct

    Characteristics of recombination breakpoints among three Porphyromonas gingivalis genomes. ( A ) Fragments are shown in the alignment of two genome sequences (TDC60, ATCC 33277). The positions of MGEs or rRNA operons in the breakpoint gaps are indicated by colored broken lines, connecting the gaps and the bars (indicating the positions of the features on the genome), which are arrayed along the outside of the plot area. The red boxes on the plot area are the regions shown in ( B ) in detail. ( B ) Breakpoint gaps of TDC60 are enlarged in light gray areas surrounded by broken lines. The regions of ATCC 33277, which correspond to the enlarged gap of TDC60, are enlarged similarly. The fragments in TDC60 and ATCC 33277 are colored by red and deep blue, respectively. The regions exhibiting high nucleotide similarity to each other are shown by a yellow belt between two fragments. The 3-kb regions of the breakpoints are indicated by dark gray rectangles on the upper or lower side of the fragments. (i) rRNA operons in the breakpoint gap. The black arrows indicate rRNA genes. The light blue-filled boxes with arrows inside indicate ISs. (ii) ISs in the breakpoint gap. ( C ) The number of each feature in the breakpoint gap is plotted. The regions without any characteristic features are included under “Others.” The mean and standard deviations are provided by the horizontal and vertical lines, respectively. Statistical significance is indicated by an asterisk ( P
    Figure Legend Snippet: Characteristics of recombination breakpoints among three Porphyromonas gingivalis genomes. ( A ) Fragments are shown in the alignment of two genome sequences (TDC60, ATCC 33277). The positions of MGEs or rRNA operons in the breakpoint gaps are indicated by colored broken lines, connecting the gaps and the bars (indicating the positions of the features on the genome), which are arrayed along the outside of the plot area. The red boxes on the plot area are the regions shown in ( B ) in detail. ( B ) Breakpoint gaps of TDC60 are enlarged in light gray areas surrounded by broken lines. The regions of ATCC 33277, which correspond to the enlarged gap of TDC60, are enlarged similarly. The fragments in TDC60 and ATCC 33277 are colored by red and deep blue, respectively. The regions exhibiting high nucleotide similarity to each other are shown by a yellow belt between two fragments. The 3-kb regions of the breakpoints are indicated by dark gray rectangles on the upper or lower side of the fragments. (i) rRNA operons in the breakpoint gap. The black arrows indicate rRNA genes. The light blue-filled boxes with arrows inside indicate ISs. (ii) ISs in the breakpoint gap. ( C ) The number of each feature in the breakpoint gap is plotted. The regions without any characteristic features are included under “Others.” The mean and standard deviations are provided by the horizontal and vertical lines, respectively. Statistical significance is indicated by an asterisk ( P

    Techniques Used:

    Split network of 60 Porphyromonas gingivalis isolates obtained from concatenated seven loci sequences. A split network tree based upon the MLST data is shown. Circles indicate external nodes (each isolate) and are colored according to geographic origin (black: Japan; outlined: overseas or unspecified). fimA types are shown by light gray shadows. The numbers outside the isolate’s name indicate the patient source. Eleven colors are used to emphasize the clusters.
    Figure Legend Snippet: Split network of 60 Porphyromonas gingivalis isolates obtained from concatenated seven loci sequences. A split network tree based upon the MLST data is shown. Circles indicate external nodes (each isolate) and are colored according to geographic origin (black: Japan; outlined: overseas or unspecified). fimA types are shown by light gray shadows. The numbers outside the isolate’s name indicate the patient source. Eleven colors are used to emphasize the clusters.

    Techniques Used:

    Regions exhibiting high nucleotide similarity to P. gingivalis CRISPR spacers. Two examples of the 19 spacers exhibiting high nucleotide similarity to the P. gingivalis genome are shown. The white and black arrows indicate CDSs and rRNA genes, respectively. The arrows within the light blue-filled boxes indicate ISs. The orange regions indicate the sequences exhibiting high nucleotide similarity to CRISPR spacers. (i) Region exhibiting high nucleotide similarity to spacer 37_259: the transposase gene in IS Pg2 , in the TDC60 genome. (ii) Region exhibiting high nucleotide similarity to spacer 37_90: close to the IS both 2-kb upstream and 2-kb downstream in the 3 genomes.
    Figure Legend Snippet: Regions exhibiting high nucleotide similarity to P. gingivalis CRISPR spacers. Two examples of the 19 spacers exhibiting high nucleotide similarity to the P. gingivalis genome are shown. The white and black arrows indicate CDSs and rRNA genes, respectively. The arrows within the light blue-filled boxes indicate ISs. The orange regions indicate the sequences exhibiting high nucleotide similarity to CRISPR spacers. (i) Region exhibiting high nucleotide similarity to spacer 37_259: the transposase gene in IS Pg2 , in the TDC60 genome. (ii) Region exhibiting high nucleotide similarity to spacer 37_90: close to the IS both 2-kb upstream and 2-kb downstream in the 3 genomes.

    Techniques Used: CRISPR

    Spacer contents of Porphyromonas gingivalis isolates from seven patients in four CRISPR loci. Spacer arrays of 26 isolates from 7 patients are shown at each CRISPR locus. Each box indicates one spacer. The spacers in the arrays exhibit high nucleotide similarity to each other among the isolates if they are aligned vertically and have the same color. Blank boxes indicate absent spacers in the particular isolates. In patient no. 2, two colors are used because the D5 isolate has a type 30 spacer array that is distinct from those of D8 and D9. The spacers in type 36.2, shared among seven isolates of three patients, are indicated by deep yellow boxes and emphasized by dark gray belts.
    Figure Legend Snippet: Spacer contents of Porphyromonas gingivalis isolates from seven patients in four CRISPR loci. Spacer arrays of 26 isolates from 7 patients are shown at each CRISPR locus. Each box indicates one spacer. The spacers in the arrays exhibit high nucleotide similarity to each other among the isolates if they are aligned vertically and have the same color. Blank boxes indicate absent spacers in the particular isolates. In patient no. 2, two colors are used because the D5 isolate has a type 30 spacer array that is distinct from those of D8 and D9. The spacers in type 36.2, shared among seven isolates of three patients, are indicated by deep yellow boxes and emphasized by dark gray belts.

    Techniques Used: CRISPR

    11) Product Images from "CD24 activates the NLRP3 inflammasome through c-Src kinase activity in a model of the lining epithelium of inflamed periodontal tissues"

    Article Title: CD24 activates the NLRP3 inflammasome through c-Src kinase activity in a model of the lining epithelium of inflamed periodontal tissues

    Journal: Immunity, Inflammation and Disease

    doi: 10.1002/iid3.40

    The bar graph shows significant downregulation of inflammasome genes encoding NLRP3, associated adaptor protein (ASC) and the pro-inflammatory cytokine IL-18 in H413 epithelial cells ligated with CD24 peptide antibody in response to Src kinase inhibitor for 3 h compared to H413 epithelial cells ligated with anti-CD24 peptide antibody only (* P
    Figure Legend Snippet: The bar graph shows significant downregulation of inflammasome genes encoding NLRP3, associated adaptor protein (ASC) and the pro-inflammatory cytokine IL-18 in H413 epithelial cells ligated with CD24 peptide antibody in response to Src kinase inhibitor for 3 h compared to H413 epithelial cells ligated with anti-CD24 peptide antibody only (* P

    Techniques Used:

    NLRP3 staining (a, b, c, and d) and Western blot (e). (a) and (e) lane-1: H413-1 cells treated with isotype control IgG1 negative antibody (5 µg/ml) for 3 h. (b) and (e) lane-2: cells treated with CD24 peptide antibody (5 µg/ml) for 3 h. An increased NLRP3 signal was observed by both confocal microscopy and Western blot (* P
    Figure Legend Snippet: NLRP3 staining (a, b, c, and d) and Western blot (e). (a) and (e) lane-1: H413-1 cells treated with isotype control IgG1 negative antibody (5 µg/ml) for 3 h. (b) and (e) lane-2: cells treated with CD24 peptide antibody (5 µg/ml) for 3 h. An increased NLRP3 signal was observed by both confocal microscopy and Western blot (* P

    Techniques Used: Staining, Western Blot, Confocal Microscopy

    Illustration of a model for protective/destructive feedback through CD24-NLRP3 and P. gingivalis -NLRP3 interactions in healthy and diseased periodontal tissues. The non-keratinised lining epithelium (junctional epithelium) provides initial responses to bacterial products by signalling through receptors of innate immunity to activate NLRP3 inflammasome pathways (right panel). These comprise an intracellular network of regulatory and effector molecules leading to synthesis and activation of pro-inflammatory cytokines [IL-1β (green dots) IL-18 (red dots)]. However, activated IL-18 potently suppresses NLRP3 and release of IL-18. Destructive feedback accelerates periodontal tissue damage. Conversely, CD24 is characteristically strongly expressed in the lining epithelium of inflamed tissues and functions as an important negative regulator for response to danger signals, protecting tissues from excessive leukocyte activity mediated by microbial activity (left panel).
    Figure Legend Snippet: Illustration of a model for protective/destructive feedback through CD24-NLRP3 and P. gingivalis -NLRP3 interactions in healthy and diseased periodontal tissues. The non-keratinised lining epithelium (junctional epithelium) provides initial responses to bacterial products by signalling through receptors of innate immunity to activate NLRP3 inflammasome pathways (right panel). These comprise an intracellular network of regulatory and effector molecules leading to synthesis and activation of pro-inflammatory cytokines [IL-1β (green dots) IL-18 (red dots)]. However, activated IL-18 potently suppresses NLRP3 and release of IL-18. Destructive feedback accelerates periodontal tissue damage. Conversely, CD24 is characteristically strongly expressed in the lining epithelium of inflamed tissues and functions as an important negative regulator for response to danger signals, protecting tissues from excessive leukocyte activity mediated by microbial activity (left panel).

    Techniques Used: Activation Assay, Activity Assay

    Quantitative real-time reverse transcription (RT)-PCR for gene expression of inflammasome and tight junction components in H413 epithelial cells in response to CD24 peptide antibody (a), active recombinant IL-18 (b and c), P. gingivalis (d) (* P
    Figure Legend Snippet: Quantitative real-time reverse transcription (RT)-PCR for gene expression of inflammasome and tight junction components in H413 epithelial cells in response to CD24 peptide antibody (a), active recombinant IL-18 (b and c), P. gingivalis (d) (* P

    Techniques Used: Reverse Transcription Polymerase Chain Reaction, Expressing, Recombinant

    12) Product Images from "Recognition of Porphyromonas gingivalis Gingipain Epitopes by Natural IgM Binding to Malondialdehyde Modified Low-Density Lipoprotein"

    Article Title: Recognition of Porphyromonas gingivalis Gingipain Epitopes by Natural IgM Binding to Malondialdehyde Modified Low-Density Lipoprotein

    Journal: PLoS ONE

    doi: 10.1371/journal.pone.0034910

    Dilution curves of mouse plasma IgM binding to antigens. A, B) C57BL/6 mice were immunized with heat-killed P. gingivalis ATCC33277 (Pg) and controls (Co) received saline. The plasma was diluted 1∶100–1∶6400 and IgM binding to MDA-LDL, CuOx-LDL, native LDL, PC-BSA (A), P. gingivalis and recombinant gingipain domains Rgp44, Rgp15–27 and RgpCAT (B) were determined before (pre) and after (post) immunization with chemiluminescence immunoassay. Mean ± SD for two samples is shown. C, D) LDLR −/− mice were immunized with killed P. gingivalis (3 strains mixed) (Pg) and controls (Co) received PBS. Pooled plasma from two mice was used for each dilution curve, and mean ± SD for two dilution curves is shown. IgM binding to MDA-LDL, CuOx-LDL, native LDL, PC-BSA (C), P. gingivalis and recombinant gingipain domain Rgp44, Rgp15–27 and RgpCAT (D) were determined before (pre) and after the second booster immunization (imm) as described.
    Figure Legend Snippet: Dilution curves of mouse plasma IgM binding to antigens. A, B) C57BL/6 mice were immunized with heat-killed P. gingivalis ATCC33277 (Pg) and controls (Co) received saline. The plasma was diluted 1∶100–1∶6400 and IgM binding to MDA-LDL, CuOx-LDL, native LDL, PC-BSA (A), P. gingivalis and recombinant gingipain domains Rgp44, Rgp15–27 and RgpCAT (B) were determined before (pre) and after (post) immunization with chemiluminescence immunoassay. Mean ± SD for two samples is shown. C, D) LDLR −/− mice were immunized with killed P. gingivalis (3 strains mixed) (Pg) and controls (Co) received PBS. Pooled plasma from two mice was used for each dilution curve, and mean ± SD for two dilution curves is shown. IgM binding to MDA-LDL, CuOx-LDL, native LDL, PC-BSA (C), P. gingivalis and recombinant gingipain domain Rgp44, Rgp15–27 and RgpCAT (D) were determined before (pre) and after the second booster immunization (imm) as described.

    Techniques Used: Binding Assay, Mouse Assay, Multiple Displacement Amplification, Recombinant, Chemiluminescence Immunoassay

    Identification of P. gingivalis epitopes for anti-MDA-LDL-IgM. A) Schematic presentation of arg-gingipain (RgpA) functional domains cloned and produced in a recombinant system [41] . B) Proteins of P. gingivalis were separated on SDS-PAGE (Pg, gel). Fragments recognized by MDmAb (45, 40 and 32 kDa, black arrows) were identified by mass spectrometry as arginine-specific gingipain or hemagglutinin A of P. gingivalis . Figures S1 , S2 , S3 contain the Mascot results of the database searches and the MSMS spectrum showing the matching amino acids in the peptide sequence. Three domains of the recombinant arg-specific gingipain, RgpCAT, Rgp44 and Rgp15–27 were produced in E. coli and analyzed for recognition by MDmAb and anti-PC-IgM control antibody (α-PC-mAb). C) Specific binding of MDmAb to recombinant gingipain domains Rgp15–27, Rgp44 and RgpCAT was tested with a competitive immunoassay. D) Reciprocally, soluble MDA-LDL, nLDL and PC-BSA were used as competitors for MDmAb binding to Rgp44. B/B 0 indicates the ratio of IgM binding with and without a competitor. MW, molecular weight. PC-BSA, phosphocholine-conjugated bovine serum albumin.
    Figure Legend Snippet: Identification of P. gingivalis epitopes for anti-MDA-LDL-IgM. A) Schematic presentation of arg-gingipain (RgpA) functional domains cloned and produced in a recombinant system [41] . B) Proteins of P. gingivalis were separated on SDS-PAGE (Pg, gel). Fragments recognized by MDmAb (45, 40 and 32 kDa, black arrows) were identified by mass spectrometry as arginine-specific gingipain or hemagglutinin A of P. gingivalis . Figures S1 , S2 , S3 contain the Mascot results of the database searches and the MSMS spectrum showing the matching amino acids in the peptide sequence. Three domains of the recombinant arg-specific gingipain, RgpCAT, Rgp44 and Rgp15–27 were produced in E. coli and analyzed for recognition by MDmAb and anti-PC-IgM control antibody (α-PC-mAb). C) Specific binding of MDmAb to recombinant gingipain domains Rgp15–27, Rgp44 and RgpCAT was tested with a competitive immunoassay. D) Reciprocally, soluble MDA-LDL, nLDL and PC-BSA were used as competitors for MDmAb binding to Rgp44. B/B 0 indicates the ratio of IgM binding with and without a competitor. MW, molecular weight. PC-BSA, phosphocholine-conjugated bovine serum albumin.

    Techniques Used: Multiple Displacement Amplification, Functional Assay, Clone Assay, Produced, Recombinant, SDS Page, Mass Spectrometry, Sequencing, Binding Assay, Molecular Weight

    Cross-reactive epitopes on MDA-LDL and bacteria. Binding of anti-MDA-LDL-IgM (MDmAb) and anti-PC-IgM control antibody (α-PC-mAb) to P. gingivalis (Pg) and PC-conjugated bovine serum albumin (PC-BSA) on Western blot (A). Binding of MDmAb to MDA- and MAA-modified and native LDL (nLDL) and PC-BSA using direct binding (B) and competitive (C) chemiluminescence immunoassays. Specific binding of MDmAb to P. gingivalis (Pg) and E. coli was tested with competitive chemiluminescence immunoassay (D). Bacterial suspensions were adjusted to an absorbance of 0.15 at 580 nm with PBS and further diluted as indicated. B/B 0 indicates the ratio of IgM binding with and without a competitor. RLU, relative light unit. Binding of MDmAb and isotype control (cntrl_IgM) to P. gingivalis (E) and E. coli (F) in native conditions was tested with flow cytometry. Fluorescence (FL-1) of the cells with the secondary antibody, +2°Ab (red), MDmAb (blue) and isotype control (green).
    Figure Legend Snippet: Cross-reactive epitopes on MDA-LDL and bacteria. Binding of anti-MDA-LDL-IgM (MDmAb) and anti-PC-IgM control antibody (α-PC-mAb) to P. gingivalis (Pg) and PC-conjugated bovine serum albumin (PC-BSA) on Western blot (A). Binding of MDmAb to MDA- and MAA-modified and native LDL (nLDL) and PC-BSA using direct binding (B) and competitive (C) chemiluminescence immunoassays. Specific binding of MDmAb to P. gingivalis (Pg) and E. coli was tested with competitive chemiluminescence immunoassay (D). Bacterial suspensions were adjusted to an absorbance of 0.15 at 580 nm with PBS and further diluted as indicated. B/B 0 indicates the ratio of IgM binding with and without a competitor. RLU, relative light unit. Binding of MDmAb and isotype control (cntrl_IgM) to P. gingivalis (E) and E. coli (F) in native conditions was tested with flow cytometry. Fluorescence (FL-1) of the cells with the secondary antibody, +2°Ab (red), MDmAb (blue) and isotype control (green).

    Techniques Used: Multiple Displacement Amplification, Binding Assay, Western Blot, Modification, Chemiluminescence Immunoassay, Flow Cytometry, Cytometry, Fluorescence

    Mouse plasma IgM binding to apoptotic T lymphocytes after P. gingivalis immunization. C57BL/6 mice were immunized with heat-killed Pg and controls received sterile saline (n = 8 per group). Mouse plasma (1∶70) IgM binding to UV-irradiated Jurkat T cells was measured with flow cytometry. A, B) Apoptotic T cell population (R1) was verified with propidium iodide (PI) staining. C) Plasma IgM binding in gate R2 of preimmune (black) and postimmune (blue) plasma samples, and competition of IgM binding with 250 µg/ml MDA-LDL (green) or native LDL (red). Inset plots (in Fig. 6C ) represent the secondary antibody control (2°Ab control) and plasma IgM binding to apoptotic cells in a Pg-immunized mouse (post). D) IgM binding to Jurkat cells was determined for each mouse in Pg-immunized and control group as geometric mean value in R2 subtracted by the 2°Ab control. Box-plot graphs represent the distribution of sample means calculated for two repeated assays. **P
    Figure Legend Snippet: Mouse plasma IgM binding to apoptotic T lymphocytes after P. gingivalis immunization. C57BL/6 mice were immunized with heat-killed Pg and controls received sterile saline (n = 8 per group). Mouse plasma (1∶70) IgM binding to UV-irradiated Jurkat T cells was measured with flow cytometry. A, B) Apoptotic T cell population (R1) was verified with propidium iodide (PI) staining. C) Plasma IgM binding in gate R2 of preimmune (black) and postimmune (blue) plasma samples, and competition of IgM binding with 250 µg/ml MDA-LDL (green) or native LDL (red). Inset plots (in Fig. 6C ) represent the secondary antibody control (2°Ab control) and plasma IgM binding to apoptotic cells in a Pg-immunized mouse (post). D) IgM binding to Jurkat cells was determined for each mouse in Pg-immunized and control group as geometric mean value in R2 subtracted by the 2°Ab control. Box-plot graphs represent the distribution of sample means calculated for two repeated assays. **P

    Techniques Used: Binding Assay, Mouse Assay, Irradiation, Flow Cytometry, Cytometry, Staining, Multiple Displacement Amplification

    Mouse plasma IgM and IgG binding to MDA-LDL after immunization with P. gingivalis . C57BL/6 mice were immunized with heat-killed P. gingivalis ATCC33277 (Pg; n = 8) and controls received saline (Co; n = 8). Plasma IgM (A) and IgG (B) to MDA-LDL before (pre) and after immunization (post) were determined with chemiluminescence immunoassay. Each C57BL/6 plasma sample (1∶500) was measured in duplicate and an average for each individual was calculated. LDLR −/− mice were immunized with killed P. gingivalis (3 strains mixed) (Pg; n = 7) and controls received PBS (Co; n = 8). Plasma IgM (C) and IgG (D) to MDA-LDL after the second booster immunization (imm) and after the HFD (end) were determined. Each LDLR −/− plasma sample (1∶1000) was measured in duplicate and an average for each individual in two repeated assays was calculated. Additionally, mouse plasma IgM binding to CuOx-LDL (E, G) and PC-BSA (F, H) was determined. For C57BL/6 mice (E, F) this was done similarly as described for panel A. Plasma samples of LDLR −/− mice (G, H) were pooled between three or four mice (1∶1000) for a single assay, in which the mean ± SD within a group is shown.
    Figure Legend Snippet: Mouse plasma IgM and IgG binding to MDA-LDL after immunization with P. gingivalis . C57BL/6 mice were immunized with heat-killed P. gingivalis ATCC33277 (Pg; n = 8) and controls received saline (Co; n = 8). Plasma IgM (A) and IgG (B) to MDA-LDL before (pre) and after immunization (post) were determined with chemiluminescence immunoassay. Each C57BL/6 plasma sample (1∶500) was measured in duplicate and an average for each individual was calculated. LDLR −/− mice were immunized with killed P. gingivalis (3 strains mixed) (Pg; n = 7) and controls received PBS (Co; n = 8). Plasma IgM (C) and IgG (D) to MDA-LDL after the second booster immunization (imm) and after the HFD (end) were determined. Each LDLR −/− plasma sample (1∶1000) was measured in duplicate and an average for each individual in two repeated assays was calculated. Additionally, mouse plasma IgM binding to CuOx-LDL (E, G) and PC-BSA (F, H) was determined. For C57BL/6 mice (E, F) this was done similarly as described for panel A. Plasma samples of LDLR −/− mice (G, H) were pooled between three or four mice (1∶1000) for a single assay, in which the mean ± SD within a group is shown.

    Techniques Used: Binding Assay, Multiple Displacement Amplification, Mouse Assay, Chemiluminescence Immunoassay

    Association between human serum IgM to P. gingivalis and MDA-LDL, and competitive binding with recombinant gingipain domains. Sera from 29 healthy adults were analyzed for IgM (A) and IgG (B) binding to Pg and MDA-LDL by chemiluminescence immunoassay. Associations between antibody levels were analyzed with Spearman rank correlation test. Human sera were pre-incubated with recombinant gingipain domains Rgp44, Rgp15–27, RgpCAT (C, D) in a competitive immunoassay detecting IgM binding to immobilized MDA-LDL. The ratio of serum IgM binding (B/B 0 ) to MDA-LDL with and without competitor (175 µg/ml) in 29 human serum samples (C) and dose-dependent competition assays of one sample (D). Reciprocal competition assay was performed to analyze human serum IgM binding to Pg antigen competed with MDA-LDL, nLDL and PC-BSA in a representative sample (E). RU, relative units.
    Figure Legend Snippet: Association between human serum IgM to P. gingivalis and MDA-LDL, and competitive binding with recombinant gingipain domains. Sera from 29 healthy adults were analyzed for IgM (A) and IgG (B) binding to Pg and MDA-LDL by chemiluminescence immunoassay. Associations between antibody levels were analyzed with Spearman rank correlation test. Human sera were pre-incubated with recombinant gingipain domains Rgp44, Rgp15–27, RgpCAT (C, D) in a competitive immunoassay detecting IgM binding to immobilized MDA-LDL. The ratio of serum IgM binding (B/B 0 ) to MDA-LDL with and without competitor (175 µg/ml) in 29 human serum samples (C) and dose-dependent competition assays of one sample (D). Reciprocal competition assay was performed to analyze human serum IgM binding to Pg antigen competed with MDA-LDL, nLDL and PC-BSA in a representative sample (E). RU, relative units.

    Techniques Used: Multiple Displacement Amplification, Binding Assay, Recombinant, Chemiluminescence Immunoassay, Incubation, Competitive Binding Assay

    13) Product Images from "Identification of genes required for the survival of B. fragilis using massive parallel sequencing of a saturated transposon mutant library"

    Article Title: Identification of genes required for the survival of B. fragilis using massive parallel sequencing of a saturated transposon mutant library

    Journal: BMC Genomics

    doi: 10.1186/1471-2164-15-429

    Comparison of B. fragilis 638R essential genes (blue circle) with B. fragilis 9343 (red circle), B. fragilis YCH46 (green circle), B. thetaiotaomicron VPI-5482 (pink circle), P. gingivalis ATCC 33277 (light red). Not all genes are labeled on the figure.
    Figure Legend Snippet: Comparison of B. fragilis 638R essential genes (blue circle) with B. fragilis 9343 (red circle), B. fragilis YCH46 (green circle), B. thetaiotaomicron VPI-5482 (pink circle), P. gingivalis ATCC 33277 (light red). Not all genes are labeled on the figure.

    Techniques Used: Labeling

    Distribution of essential genes on B. fragilis 638R genome. Distribution of essential genes on positive (outside circle) or negative (inside circle) strands of B. fragilis 638R chromosome.
    Figure Legend Snippet: Distribution of essential genes on B. fragilis 638R genome. Distribution of essential genes on positive (outside circle) or negative (inside circle) strands of B. fragilis 638R chromosome.

    Techniques Used:

    COG Classification of B. fragilis 638R essential genes. Legend: A- RNA processing and modification, C-Energy production and conversion, D- Cell cycle control, cell division, chromosome partitioning, E- Amino acid transport and metabolism, F- Nucleotide transport and metabolism, G- Carbohydrate transport and metabolism, H- Coenzyme transport and metabolism, I- Lipid transport and metabolism, J- Translation, ribosomal structure and biogenesis, K- Transcription, L- Replication, recombination and repair, M- Cell wall/membrane/envelope biogenesis, N- Cell motility, O- Posttranslational modification, protein turnover, chaperones, P- Inorganic ion transport and metabolism, Q- Secondary metabolites biosynthesis, transport and catabolism, R- General function prediction only, T- Signal transduction mechanisms, U- Intracellular trafficking, secretion, and vesicular transport, V- Defense mechanisms, S- Function unknown, X- Essential genes not in COG.
    Figure Legend Snippet: COG Classification of B. fragilis 638R essential genes. Legend: A- RNA processing and modification, C-Energy production and conversion, D- Cell cycle control, cell division, chromosome partitioning, E- Amino acid transport and metabolism, F- Nucleotide transport and metabolism, G- Carbohydrate transport and metabolism, H- Coenzyme transport and metabolism, I- Lipid transport and metabolism, J- Translation, ribosomal structure and biogenesis, K- Transcription, L- Replication, recombination and repair, M- Cell wall/membrane/envelope biogenesis, N- Cell motility, O- Posttranslational modification, protein turnover, chaperones, P- Inorganic ion transport and metabolism, Q- Secondary metabolites biosynthesis, transport and catabolism, R- General function prediction only, T- Signal transduction mechanisms, U- Intracellular trafficking, secretion, and vesicular transport, V- Defense mechanisms, S- Function unknown, X- Essential genes not in COG.

    Techniques Used: Modification, Transduction

    Venn diagram of classifications of B. fragilis 638R essential genes. BF638R- B. fragilis 638R; BF9343- B. fragilis 9343; BFYCH46- B. fragilis YCH46; DEG-database of essential genes. There are 346 essential genes which are common to B. fragilis strains and have homologs in the DEG. Two hundred essential genes of B. fragilis 638R have no homologs in the DEG and 31 genes are specific to B. fragilis 638R.
    Figure Legend Snippet: Venn diagram of classifications of B. fragilis 638R essential genes. BF638R- B. fragilis 638R; BF9343- B. fragilis 9343; BFYCH46- B. fragilis YCH46; DEG-database of essential genes. There are 346 essential genes which are common to B. fragilis strains and have homologs in the DEG. Two hundred essential genes of B. fragilis 638R have no homologs in the DEG and 31 genes are specific to B. fragilis 638R.

    Techniques Used:

    Distribution of GC content in B. fragilis 638R genes. No. of genes with GC percentage indicated on Y-axis. Blue line: B. fragilis 638R genes. Green line: B. fragilis 638R essential genes. Red line: B. fragilis 638R essential genes that are common to other B. fragilis species . Red dots: B. fragilis 638R essential genes unique to B. fragilis 638R. Blue Dots: B. fragilis 638R essential genes with matches in the DEG but not to P. gingivalis or B. thetaiotaomicron . Note: The lines are mapped to the primary Y-axis and the dots to the secondary (right side) Y-axis.
    Figure Legend Snippet: Distribution of GC content in B. fragilis 638R genes. No. of genes with GC percentage indicated on Y-axis. Blue line: B. fragilis 638R genes. Green line: B. fragilis 638R essential genes. Red line: B. fragilis 638R essential genes that are common to other B. fragilis species . Red dots: B. fragilis 638R essential genes unique to B. fragilis 638R. Blue Dots: B. fragilis 638R essential genes with matches in the DEG but not to P. gingivalis or B. thetaiotaomicron . Note: The lines are mapped to the primary Y-axis and the dots to the secondary (right side) Y-axis.

    Techniques Used:

    14) Product Images from "The Porphyromonas gingivalis Ferric Uptake Regulator Orthologue Binds Hemin and Regulates Hemin-Responsive Biofilm Development"

    Article Title: The Porphyromonas gingivalis Ferric Uptake Regulator Orthologue Binds Hemin and Regulates Hemin-Responsive Biofilm Development

    Journal: PLoS ONE

    doi: 10.1371/journal.pone.0111168

    P. gingivalis biofilm development. Orthogonal projections of CLSM images showing a representative region of the x-y plane over the depth of the biofilm in both xz and yz dimensions of the ATCC 33277 wild-type, har mutant ECR455 and har complement ECR 475 strains grown in excess hemin ( A ) or hemin-limitation ( B ). Comparison of the Biovolume ( C ), Average Thickness ( D ) and SA:Biovolume ( E ) calculated for each strain's biofilm growth in either excess hemin (dark bars) or limited hemin (light bars) over three independent experiments. All biometric parameters analysed for the biofilms formed by ATCC 33277 and ECR475 were significantly (p
    Figure Legend Snippet: P. gingivalis biofilm development. Orthogonal projections of CLSM images showing a representative region of the x-y plane over the depth of the biofilm in both xz and yz dimensions of the ATCC 33277 wild-type, har mutant ECR455 and har complement ECR 475 strains grown in excess hemin ( A ) or hemin-limitation ( B ). Comparison of the Biovolume ( C ), Average Thickness ( D ) and SA:Biovolume ( E ) calculated for each strain's biofilm growth in either excess hemin (dark bars) or limited hemin (light bars) over three independent experiments. All biometric parameters analysed for the biofilms formed by ATCC 33277 and ECR475 were significantly (p

    Techniques Used: Confocal Laser Scanning Microscopy, Mutagenesis

    Genomic arrangement of P. gingivalis ATCC 33277 in (A) the wild-type strain, (B) har mutant strain ECR455 and (C) har complemented strain ECR475. ‘P’ denotes promoter positions, the arrows above ‘P’ denote the direction of transcription whilst the stem loop following ermF indicates a Rho-independent transcriptional terminator. Not drawn to scale. (D) RT-PCR analysis of PGN_1504 and PGN_1503 ( har ). Reverse transcription of ECR455 and ECR475 RNA was performed using random hexamers. PCR was then performed using oligonucleotide primers specific for PGN_1504 (lanes 1–4) or PGN_1503 ( har ) (lanes 5–8 and 9–12). The templates used for PCR were: reverse transcribed ECR455 RNA (lanes 1 and 5), reverse transcribed ECR475 RNA (lane 9), RNA that was not reverse transcribed (lanes 2, 6 and 10), no template (lanes 3, 7 and 11) and P. gingivalis ATCC 33277 genomic DNA (lanes 4, 8 and 12). PGN_1504 transcript was detected in the har mutant ECR455 (lane 1), whilst PGN_1503 ( har ) transcript was not detected in the har mutant strain ECR455 (lane 5), but was detected in the har complemented strain ECR475 (lane 9). (E) Western blot detection of Har expression in P. gingivalis 33277, ECR455 and ECR475. Cytoplasmic protein extracts (25 µg) from P. gingivalis strains 33277 (B), ECR455 (C), ECR475 (D) and 5 ng purified Har (A) were separated on a 4–12% Bis-Tris polyacrylamide gel (Invitrogen) before Western transfer and blotting with anti-rHar sera. Har protein was detected in the 33277 wild-type and ECR475 complement, but not the ECR455 mutant strain.
    Figure Legend Snippet: Genomic arrangement of P. gingivalis ATCC 33277 in (A) the wild-type strain, (B) har mutant strain ECR455 and (C) har complemented strain ECR475. ‘P’ denotes promoter positions, the arrows above ‘P’ denote the direction of transcription whilst the stem loop following ermF indicates a Rho-independent transcriptional terminator. Not drawn to scale. (D) RT-PCR analysis of PGN_1504 and PGN_1503 ( har ). Reverse transcription of ECR455 and ECR475 RNA was performed using random hexamers. PCR was then performed using oligonucleotide primers specific for PGN_1504 (lanes 1–4) or PGN_1503 ( har ) (lanes 5–8 and 9–12). The templates used for PCR were: reverse transcribed ECR455 RNA (lanes 1 and 5), reverse transcribed ECR475 RNA (lane 9), RNA that was not reverse transcribed (lanes 2, 6 and 10), no template (lanes 3, 7 and 11) and P. gingivalis ATCC 33277 genomic DNA (lanes 4, 8 and 12). PGN_1504 transcript was detected in the har mutant ECR455 (lane 1), whilst PGN_1503 ( har ) transcript was not detected in the har mutant strain ECR455 (lane 5), but was detected in the har complemented strain ECR475 (lane 9). (E) Western blot detection of Har expression in P. gingivalis 33277, ECR455 and ECR475. Cytoplasmic protein extracts (25 µg) from P. gingivalis strains 33277 (B), ECR455 (C), ECR475 (D) and 5 ng purified Har (A) were separated on a 4–12% Bis-Tris polyacrylamide gel (Invitrogen) before Western transfer and blotting with anti-rHar sera. Har protein was detected in the 33277 wild-type and ECR475 complement, but not the ECR455 mutant strain.

    Techniques Used: Mutagenesis, Reverse Transcription Polymerase Chain Reaction, Polymerase Chain Reaction, Western Blot, Expressing, Purification

    15) Product Images from "The impact of virulence factors of Porphyromonas gingivalis on wound healing in vitro"

    Article Title: The impact of virulence factors of Porphyromonas gingivalis on wound healing in vitro

    Journal: Journal of Oral Microbiology

    doi: 10.3402/jom.v7.27543

    Mean relative closure (±SEM) from all biological replicates of scratches in oral epithelial cells challenged with LPS from P. gingivalis and LPS inactivated P. gingivalis by polymyxin B (a) and heat-inactivated P. gingivalis strains W83 and EpsC mutant (b). * p
    Figure Legend Snippet: Mean relative closure (±SEM) from all biological replicates of scratches in oral epithelial cells challenged with LPS from P. gingivalis and LPS inactivated P. gingivalis by polymyxin B (a) and heat-inactivated P. gingivalis strains W83 and EpsC mutant (b). * p

    Techniques Used: Mutagenesis

    Mean relative closure (±SEM) from all biological replicates of scratches in oral epithelial cells challenged with heat-inactivated and viable P. gingivalis strains ATCC 33277, W83, and W50. a=significantly different from control ( p
    Figure Legend Snippet: Mean relative closure (±SEM) from all biological replicates of scratches in oral epithelial cells challenged with heat-inactivated and viable P. gingivalis strains ATCC 33277, W83, and W50. a=significantly different from control ( p

    Techniques Used:

    16) Product Images from "Intraspecies Variability Affects Heterotypic Biofilms of Porphyromonas gingivalis and Prevotella intermedia: Evidences of Strain-Dependence Biofilm Modulation by Physical Contact and by Released Soluble Factors"

    Article Title: Intraspecies Variability Affects Heterotypic Biofilms of Porphyromonas gingivalis and Prevotella intermedia: Evidences of Strain-Dependence Biofilm Modulation by Physical Contact and by Released Soluble Factors

    Journal: PLoS ONE

    doi: 10.1371/journal.pone.0138687

    Quantification of biomass (OD 490 ) in two-species biofilms of Porphyromonas gingivalis , Prevotella intermedia and Prevotella nigrescens . a means significant difference between values of heterotypic biofilm and values of monotypic biofilm of the first strain in the association (Kruskal-Wallis test followed by nonparametric post-tests, p
    Figure Legend Snippet: Quantification of biomass (OD 490 ) in two-species biofilms of Porphyromonas gingivalis , Prevotella intermedia and Prevotella nigrescens . a means significant difference between values of heterotypic biofilm and values of monotypic biofilm of the first strain in the association (Kruskal-Wallis test followed by nonparametric post-tests, p

    Techniques Used:

    Effect of co-culture of Porphyromonas gingivalis , Prevotella intermedia and Prevotella nigrescens on biofilm formation in a two-compartment co-culture system. Panel A : P . gingivalis ATCC33277 and co-culture partners P . intermedia strains 17 and ATCC25611, and P . nigrescens ATCC33563. Panel B : P . gingivalis W83 and co-culture partners P . intermedia strains 17 and ATCC25611, and P . nigrescens ATCC33563. Panel C : P . intermedia strains 17 and ATCC25611 and co-culture partner P . nigrescens ATCC33563 Values represent average of 3 experiments (Kruskal-Wallis test followed by nonparametric post-tests, * p
    Figure Legend Snippet: Effect of co-culture of Porphyromonas gingivalis , Prevotella intermedia and Prevotella nigrescens on biofilm formation in a two-compartment co-culture system. Panel A : P . gingivalis ATCC33277 and co-culture partners P . intermedia strains 17 and ATCC25611, and P . nigrescens ATCC33563. Panel B : P . gingivalis W83 and co-culture partners P . intermedia strains 17 and ATCC25611, and P . nigrescens ATCC33563. Panel C : P . intermedia strains 17 and ATCC25611 and co-culture partner P . nigrescens ATCC33563 Values represent average of 3 experiments (Kruskal-Wallis test followed by nonparametric post-tests, * p

    Techniques Used: Co-Culture Assay

    Values inside the bars represent the percentage (%) of each strain in two-species biofilm measured by q-PCR after 24 hrs of incubation. Panel A : two-species biofilms of P . gingivalis W83 mixed with either P . intermedia 17, P . intermedia ATCC25611 or P . nigrescens ATCC33563; Panel B : two-species biofilms of P . gingivalis ATCC33277 mixed with either P . intermedia 17, P . intermedia ATCC25611 or P . nigrescens ATCC 33563; Panel C : two-species biofilms of P . nigrescens ATCC33563 mixed with either P . intermedia 17 or P . intermedia ATCC25611.
    Figure Legend Snippet: Values inside the bars represent the percentage (%) of each strain in two-species biofilm measured by q-PCR after 24 hrs of incubation. Panel A : two-species biofilms of P . gingivalis W83 mixed with either P . intermedia 17, P . intermedia ATCC25611 or P . nigrescens ATCC33563; Panel B : two-species biofilms of P . gingivalis ATCC33277 mixed with either P . intermedia 17, P . intermedia ATCC25611 or P . nigrescens ATCC 33563; Panel C : two-species biofilms of P . nigrescens ATCC33563 mixed with either P . intermedia 17 or P . intermedia ATCC25611.

    Techniques Used: Polymerase Chain Reaction, Incubation

    Representative confocal micrographs of 24 hrs biofilms of P . gingivalis and P . intermedia in chamber cells and thickness measurements. Panel A : P . gingivalis W83 (100x); Panel B : P . gingivalis ATCC33277 (100x); Panel C : P . intermedia 17 (40x); Panel D : P . intermedia ATCC25611 (40x); Panel E : P . gingivalis W83 + P . intermedia 17 (100x); Panel F : P . gingivalis ATCC33277 + P . intermedia 17 (100x); Panel G : P . gingivalis W83 + P . intermedia ATCC25611 (40x); Panel H : P . gingivalis ATCC33277 + P . intermedia ATCC25611 (40x). Bacterial cells were stained with species-specific fluorophore-conjugated probes.
    Figure Legend Snippet: Representative confocal micrographs of 24 hrs biofilms of P . gingivalis and P . intermedia in chamber cells and thickness measurements. Panel A : P . gingivalis W83 (100x); Panel B : P . gingivalis ATCC33277 (100x); Panel C : P . intermedia 17 (40x); Panel D : P . intermedia ATCC25611 (40x); Panel E : P . gingivalis W83 + P . intermedia 17 (100x); Panel F : P . gingivalis ATCC33277 + P . intermedia 17 (100x); Panel G : P . gingivalis W83 + P . intermedia ATCC25611 (40x); Panel H : P . gingivalis ATCC33277 + P . intermedia ATCC25611 (40x). Bacterial cells were stained with species-specific fluorophore-conjugated probes.

    Techniques Used: Staining

    17) Product Images from "Three CoA Transferases Involved in the Production of Short Chain Fatty Acids in Porphyromonas gingivalis"

    Article Title: Three CoA Transferases Involved in the Production of Short Chain Fatty Acids in Porphyromonas gingivalis

    Journal: Frontiers in Microbiology

    doi: 10.3389/fmicb.2016.01146

    Steady-state kinetic analysis of butyryl-CoA:acetate CoA transferase activities of recombinant PGN_0725, PGN_1341, and PGN_1888 proteins . (A) Double reciprocal plots of the initial velocities of acetyl-CoA and butyrate formation from butyryl-CoA and sodium acetate catalyzed by purified recombinant enzymes. Different butyryl-CoA concentrations (0.5–5 mM) were assayed at fixed sodium acetate concentrations (25 mM, filled squares; 50 mM, open squares; 100 mM, filled triangles; 125 mM, open triangles; 200 mM, filled circles; or 250 mM, open circles). (B) Secondary plots of y intercepts (velocity reciprocals) vs. sodium acetate concentrations. (C) Secondary plots of the reciprocal slopes from panel (A) vs. sodium acetate concentration. Data represent the mean ± standard deviation ( n = 3).
    Figure Legend Snippet: Steady-state kinetic analysis of butyryl-CoA:acetate CoA transferase activities of recombinant PGN_0725, PGN_1341, and PGN_1888 proteins . (A) Double reciprocal plots of the initial velocities of acetyl-CoA and butyrate formation from butyryl-CoA and sodium acetate catalyzed by purified recombinant enzymes. Different butyryl-CoA concentrations (0.5–5 mM) were assayed at fixed sodium acetate concentrations (25 mM, filled squares; 50 mM, open squares; 100 mM, filled triangles; 125 mM, open triangles; 200 mM, filled circles; or 250 mM, open circles). (B) Secondary plots of y intercepts (velocity reciprocals) vs. sodium acetate concentrations. (C) Secondary plots of the reciprocal slopes from panel (A) vs. sodium acetate concentration. Data represent the mean ± standard deviation ( n = 3).

    Techniques Used: Recombinant, Purification, Concentration Assay, Standard Deviation

    GC-MS chromatograms of butyrate and propionate in reaction mixtures . Total ion chromatogram of butyrate (A) and propionate (B) is shown. The mass spectra of each sample agree well with those of the corresponding standard. The reaction mixtures, containing 200 mM sodium acetate and either 1 mM butyryl-CoA or 1 mM propionyl-CoA, with recombinant PGN_0725, PGN_1341, PGN_1888, or PGN_1171 proteins, were incubated for 1 h. After the proteins were removed by acetone precipitation treatment, aqueous phase aliquot (1 μl) was analyzed.
    Figure Legend Snippet: GC-MS chromatograms of butyrate and propionate in reaction mixtures . Total ion chromatogram of butyrate (A) and propionate (B) is shown. The mass spectra of each sample agree well with those of the corresponding standard. The reaction mixtures, containing 200 mM sodium acetate and either 1 mM butyryl-CoA or 1 mM propionyl-CoA, with recombinant PGN_0725, PGN_1341, PGN_1888, or PGN_1171 proteins, were incubated for 1 h. After the proteins were removed by acetone precipitation treatment, aqueous phase aliquot (1 μl) was analyzed.

    Techniques Used: Gas Chromatography-Mass Spectrometry, Recombinant, Incubation

    Steady-state kinetic analysis of propionyl-CoA:acetate CoA transferase activities of recombinant PGN_0725 and PGN_1888 proteins . (A) Double reciprocal plots of the initial velocities of acetyl-CoA and propionate formation from propionyl-CoA and sodium acetate catalyzed by purified recombinant enzymes. Different concentrations of propionyl-CoA (0.2–3 mM) were assayed at fixed sodium acetate concentrations (20 mM, filled circles; 25 mM, open circles; 40 mM, filled triangles; 50 mM, open triangles; 75 mM, filled squares; 100 mM, open squares; 125 mM, filled diamonds; 200 mM, open diamonds; and 250 mM, crosses). (B) Secondary plots of y intercepts (velocity reciprocals) vs. sodium acetate concentrations. (C) Secondary plots of the reciprocal slopes from panel (A) vs. sodium acetate concentrations. Data represent the mean ± standard deviation ( n = 3).
    Figure Legend Snippet: Steady-state kinetic analysis of propionyl-CoA:acetate CoA transferase activities of recombinant PGN_0725 and PGN_1888 proteins . (A) Double reciprocal plots of the initial velocities of acetyl-CoA and propionate formation from propionyl-CoA and sodium acetate catalyzed by purified recombinant enzymes. Different concentrations of propionyl-CoA (0.2–3 mM) were assayed at fixed sodium acetate concentrations (20 mM, filled circles; 25 mM, open circles; 40 mM, filled triangles; 50 mM, open triangles; 75 mM, filled squares; 100 mM, open squares; 125 mM, filled diamonds; 200 mM, open diamonds; and 250 mM, crosses). (B) Secondary plots of y intercepts (velocity reciprocals) vs. sodium acetate concentrations. (C) Secondary plots of the reciprocal slopes from panel (A) vs. sodium acetate concentrations. Data represent the mean ± standard deviation ( n = 3).

    Techniques Used: Recombinant, Purification, Standard Deviation

    18) Product Images from "Porphyromonas gingivalis Gingipain-Dependently Enhances IL-33 Production in Human Gingival Epithelial Cells"

    Article Title: Porphyromonas gingivalis Gingipain-Dependently Enhances IL-33 Production in Human Gingival Epithelial Cells

    Journal: PLoS ONE

    doi: 10.1371/journal.pone.0152794

    P . gingivalis -induced IL-33 mRNA expression requires activation of PAR-2 and PLC. (A) Expression of PAR-1 , -2 , -3 , and -4 mRNA by Ca9-22 cells analyzed by RT-PCR. Data are representative of two independent experiments. Fragments are 156 bp ( GAPDH ), 150 bp ( PAR-1 ), 153 bp ( PAR-2 ), 170 bp ( PAR-3 ), and 177 bp ( PAR-4 ), respectively. Ca9-22 cells were transfected with 100 pmol of PAR-2 siRNA or control siRNA for 14 h (C and D), and stimulated with 50 μg/ml of whole P . gingivalis W83 cells for 48 h (B and D). Ca9-22 cells were incubated with indicated concentrations of U-73122 (PLC inhibitor) (E) or GF109203X (PKC inhibitor) (F) for 30 min, and then stimulated with 50 μg/ml of whole P . gingivalis W83 cells for 48 h. Expression of PAR-2 mRNA (B and C) and IL-33 mRNA (D, E, and F) was analyzed using RT-qPCR. Data are representative of three independent experiments, and are shown as means ± SD of triplicate assays. Statistical significant differences are indicated (*, P
    Figure Legend Snippet: P . gingivalis -induced IL-33 mRNA expression requires activation of PAR-2 and PLC. (A) Expression of PAR-1 , -2 , -3 , and -4 mRNA by Ca9-22 cells analyzed by RT-PCR. Data are representative of two independent experiments. Fragments are 156 bp ( GAPDH ), 150 bp ( PAR-1 ), 153 bp ( PAR-2 ), 170 bp ( PAR-3 ), and 177 bp ( PAR-4 ), respectively. Ca9-22 cells were transfected with 100 pmol of PAR-2 siRNA or control siRNA for 14 h (C and D), and stimulated with 50 μg/ml of whole P . gingivalis W83 cells for 48 h (B and D). Ca9-22 cells were incubated with indicated concentrations of U-73122 (PLC inhibitor) (E) or GF109203X (PKC inhibitor) (F) for 30 min, and then stimulated with 50 μg/ml of whole P . gingivalis W83 cells for 48 h. Expression of PAR-2 mRNA (B and C) and IL-33 mRNA (D, E, and F) was analyzed using RT-qPCR. Data are representative of three independent experiments, and are shown as means ± SD of triplicate assays. Statistical significant differences are indicated (*, P

    Techniques Used: Expressing, Activation Assay, Planar Chromatography, Reverse Transcription Polymerase Chain Reaction, Transfection, Incubation, Quantitative RT-PCR

    Induction of IL-33 mRNA expression by P . gingivalis requires activation of the NF-κB pathway. (A) Ca9-22 cells stimulated with 50 μg/ml of whole P . gingivalis W83 cells for indicated periods. (B) Whole P . gingivalis W83 cells (50 μg/ml) were incubated with gingipain inhibitors (0.3 μM FPR-cmk plus 0.3 μM KYT-36) for 15 min at 37°C, and then used to stimulate Ca9-22 cells for 9 h. (C) Ca9-22 cells stimulated with 50 μg/ml of whole P . gingivalis ATCC 33277 wild-type or KDP136 cells for 9 h. (A-C) Cells were transiently transfected with pNFκB- Metridia luciferase reporter or control p Metridia luciferase reporter plasmids. Amount of secreted luciferase in culture supernatants were analyzed using a luminometer. (D) Ca9-22 cells incubated with indicated concentrations of PDTC (NF-κB inhibitor) for 1 h and then stimulated with 50 μg/ml of whole P . gingivalis W83 cells for 48 h. Total cellular RNA was extracted and transcripts were analyzed by RT-qPCR. Data are representative of three independent experiments and are shown as means ± SD of triplicate assays. Statistical significant differences are indicated (*, P
    Figure Legend Snippet: Induction of IL-33 mRNA expression by P . gingivalis requires activation of the NF-κB pathway. (A) Ca9-22 cells stimulated with 50 μg/ml of whole P . gingivalis W83 cells for indicated periods. (B) Whole P . gingivalis W83 cells (50 μg/ml) were incubated with gingipain inhibitors (0.3 μM FPR-cmk plus 0.3 μM KYT-36) for 15 min at 37°C, and then used to stimulate Ca9-22 cells for 9 h. (C) Ca9-22 cells stimulated with 50 μg/ml of whole P . gingivalis ATCC 33277 wild-type or KDP136 cells for 9 h. (A-C) Cells were transiently transfected with pNFκB- Metridia luciferase reporter or control p Metridia luciferase reporter plasmids. Amount of secreted luciferase in culture supernatants were analyzed using a luminometer. (D) Ca9-22 cells incubated with indicated concentrations of PDTC (NF-κB inhibitor) for 1 h and then stimulated with 50 μg/ml of whole P . gingivalis W83 cells for 48 h. Total cellular RNA was extracted and transcripts were analyzed by RT-qPCR. Data are representative of three independent experiments and are shown as means ± SD of triplicate assays. Statistical significant differences are indicated (*, P

    Techniques Used: Expressing, Activation Assay, Incubation, Transfection, Luciferase, Quantitative RT-PCR

    P . gingivalis increases IL-33 mRNA expression in gingival epithelial cells. Ca9-22 cells were infected with fresh P . gingivalis W83 cultures at MOI of 0.5 for the indicated periods (A) or indicated MOI for 48 h (B). Ca9-22 cells were stimulated with 50 μg/ml (MOI of 0.1) of whole P . gingivalis W83 cells for the indicated periods (C) and indicated amounts of whole P . gingivalis W83 cells, fimbriae, PGTP2-RL, and LPS derived from P . gingivalis for 48 h (D). Cells were incubated with 10 μg/ml cycloheximide for 45 min (E), 1 μg/ml nocodazole for 1 h (F) or 0.5 μM cytochalasin D for 30 min (G) and then stimulated for 48 h with 50 μg/ml of P . gingivalis W83 whole cells with the respective inhibitors. Total cellular RNA was then extracted at the indicated times, and IL-33 transcripts were analyzed by RT-qPCR. Data are representative of three independent experiments, and are shown as means ± SD of triplicate assays. Statistically significant differences are indicated. *, P
    Figure Legend Snippet: P . gingivalis increases IL-33 mRNA expression in gingival epithelial cells. Ca9-22 cells were infected with fresh P . gingivalis W83 cultures at MOI of 0.5 for the indicated periods (A) or indicated MOI for 48 h (B). Ca9-22 cells were stimulated with 50 μg/ml (MOI of 0.1) of whole P . gingivalis W83 cells for the indicated periods (C) and indicated amounts of whole P . gingivalis W83 cells, fimbriae, PGTP2-RL, and LPS derived from P . gingivalis for 48 h (D). Cells were incubated with 10 μg/ml cycloheximide for 45 min (E), 1 μg/ml nocodazole for 1 h (F) or 0.5 μM cytochalasin D for 30 min (G) and then stimulated for 48 h with 50 μg/ml of P . gingivalis W83 whole cells with the respective inhibitors. Total cellular RNA was then extracted at the indicated times, and IL-33 transcripts were analyzed by RT-qPCR. Data are representative of three independent experiments, and are shown as means ± SD of triplicate assays. Statistically significant differences are indicated. *, P

    Techniques Used: Expressing, Infection, Derivative Assay, Incubation, Quantitative RT-PCR

    P . gingivalis increases IL-33 protein expression in gingival epithelial cells. (A) Ca9-22 cells were stimulated with 50 μg/ml of whole P . gingivalis W83 cells for the indicated periods. Cell lysates were analyzed by Western blotting with an anti-human IL-33 mAb. Expression levels of IL-33 were quantified by densitometry using ImageJ software and normalized to medium alone. (B) Ca9-22 cells were stimulated with 50 μg/ml of whole P . gingivalis W83 cells for 4 d, and intracellular IL-33 protein was stained with PE-conjugated anti-human IL-33 mAb. Nuclei were stained with DAPI. Bar = 25 μm. Data are representative of three independent experiments.
    Figure Legend Snippet: P . gingivalis increases IL-33 protein expression in gingival epithelial cells. (A) Ca9-22 cells were stimulated with 50 μg/ml of whole P . gingivalis W83 cells for the indicated periods. Cell lysates were analyzed by Western blotting with an anti-human IL-33 mAb. Expression levels of IL-33 were quantified by densitometry using ImageJ software and normalized to medium alone. (B) Ca9-22 cells were stimulated with 50 μg/ml of whole P . gingivalis W83 cells for 4 d, and intracellular IL-33 protein was stained with PE-conjugated anti-human IL-33 mAb. Nuclei were stained with DAPI. Bar = 25 μm. Data are representative of three independent experiments.

    Techniques Used: Expressing, Western Blot, Software, Staining

    Participation of gingipains in P . gingivalis -induced IL-33 mRNA expression in gingival/oral epithelial cells. Whole P . gingivalis W83 cells (50 μg/ml) were incubated with 0.3 μM FPR-cmk (Rgp inhibitor) or 0.3 μM KYT-36 (Kgp inhibitor) for 15 min at 37°C and then used to stimulate Ca9-22 (A) or primary oral epithelial (B) cells for 48 h. (C) Ca9-22 cells stimulated with 50 μg/ml of whole cells of P . gingivalis ATCC 33277 wild-type, P . gingivalis KDP131 (Δ rgpA ), KDP132 (Δ rgpB ), KDP129 (Δ kgp ), KDP133 (Δ rgpA Δ rgpB ), or KDP136 (Δ kgp Δ rgpA Δ rgpB ) gingipain-null mutant for 48 h. (D) Primary oral epithelial cells stimulated with 50 μg/ml of P . gingivalis ATCC 33277 wild-type or KDP136 for 48 h. (E) Ca9-22 cells cultured for 48 h with 50 μg/ml of whole P . gingivalis W83 cells after incubation with or without at 70°C for 1 h. Total cellular RNA was extracted and transcripts were analyzed by RT-qPCR. Data are representative of three independent experiments, and are shown as means ± SD of triplicate assays. Statistical significant differences are indicated (*, P
    Figure Legend Snippet: Participation of gingipains in P . gingivalis -induced IL-33 mRNA expression in gingival/oral epithelial cells. Whole P . gingivalis W83 cells (50 μg/ml) were incubated with 0.3 μM FPR-cmk (Rgp inhibitor) or 0.3 μM KYT-36 (Kgp inhibitor) for 15 min at 37°C and then used to stimulate Ca9-22 (A) or primary oral epithelial (B) cells for 48 h. (C) Ca9-22 cells stimulated with 50 μg/ml of whole cells of P . gingivalis ATCC 33277 wild-type, P . gingivalis KDP131 (Δ rgpA ), KDP132 (Δ rgpB ), KDP129 (Δ kgp ), KDP133 (Δ rgpA Δ rgpB ), or KDP136 (Δ kgp Δ rgpA Δ rgpB ) gingipain-null mutant for 48 h. (D) Primary oral epithelial cells stimulated with 50 μg/ml of P . gingivalis ATCC 33277 wild-type or KDP136 for 48 h. (E) Ca9-22 cells cultured for 48 h with 50 μg/ml of whole P . gingivalis W83 cells after incubation with or without at 70°C for 1 h. Total cellular RNA was extracted and transcripts were analyzed by RT-qPCR. Data are representative of three independent experiments, and are shown as means ± SD of triplicate assays. Statistical significant differences are indicated (*, P

    Techniques Used: Expressing, Incubation, Mutagenesis, Cell Culture, Quantitative RT-PCR

    Induction of IL-33 mRNA expression by P . gingivalis requires activation of the p38 pathway. (A) Ca9-22 cells stimulated for indicated periods with 50 μg/ml of whole P . gingivalis W83 cells in medium containing 1% FBS. Phosphorylation of p38 was detected in cell lysates by Western blotting against anti-phospho-p38 antibody (p-p38). Controls comprised antibody against total p38. Data are representative of three independent experiments. Relative expression of phosphorylated p38 was quantified using densitometry. Relative expression of phosphorylated p38 was normalized to that of p38. (B) Ca9-22 cells incubated with 10 μM PD98059, SP600125, or SB203580 for 30 min and then stimulated with 50 μg/ml of whole P . gingivalis W83 cells for 48 h in medium containing 5% FBS. Total cellular RNA was extracted and transcripts were analyzed by RT-qPCR. Data are representative of three independent experiments, and are shown as means ± SD of triplicate assays. Statistical significant differences are indicated (*, P
    Figure Legend Snippet: Induction of IL-33 mRNA expression by P . gingivalis requires activation of the p38 pathway. (A) Ca9-22 cells stimulated for indicated periods with 50 μg/ml of whole P . gingivalis W83 cells in medium containing 1% FBS. Phosphorylation of p38 was detected in cell lysates by Western blotting against anti-phospho-p38 antibody (p-p38). Controls comprised antibody against total p38. Data are representative of three independent experiments. Relative expression of phosphorylated p38 was quantified using densitometry. Relative expression of phosphorylated p38 was normalized to that of p38. (B) Ca9-22 cells incubated with 10 μM PD98059, SP600125, or SB203580 for 30 min and then stimulated with 50 μg/ml of whole P . gingivalis W83 cells for 48 h in medium containing 5% FBS. Total cellular RNA was extracted and transcripts were analyzed by RT-qPCR. Data are representative of three independent experiments, and are shown as means ± SD of triplicate assays. Statistical significant differences are indicated (*, P

    Techniques Used: Expressing, Activation Assay, Western Blot, Incubation, Quantitative RT-PCR

    19) Product Images from "The Porphyromonas gingivalis Ferric Uptake Regulator Orthologue Binds Hemin and Regulates Hemin-Responsive Biofilm Development"

    Article Title: The Porphyromonas gingivalis Ferric Uptake Regulator Orthologue Binds Hemin and Regulates Hemin-Responsive Biofilm Development

    Journal: PLoS ONE

    doi: 10.1371/journal.pone.0111168

    P. gingivalis biofilm development. Orthogonal projections of CLSM images showing a representative region of the x-y plane over the depth of the biofilm in both xz and yz dimensions of the ATCC 33277 wild-type, har mutant ECR455 and har complement ECR 475 strains grown in excess hemin ( A ) or hemin-limitation ( B ). Comparison of the Biovolume ( C ), Average Thickness ( D ) and SA:Biovolume ( E ) calculated for each strain's biofilm growth in either excess hemin (dark bars) or limited hemin (light bars) over three independent experiments. All biometric parameters analysed for the biofilms formed by ATCC 33277 and ECR475 were significantly (p
    Figure Legend Snippet: P. gingivalis biofilm development. Orthogonal projections of CLSM images showing a representative region of the x-y plane over the depth of the biofilm in both xz and yz dimensions of the ATCC 33277 wild-type, har mutant ECR455 and har complement ECR 475 strains grown in excess hemin ( A ) or hemin-limitation ( B ). Comparison of the Biovolume ( C ), Average Thickness ( D ) and SA:Biovolume ( E ) calculated for each strain's biofilm growth in either excess hemin (dark bars) or limited hemin (light bars) over three independent experiments. All biometric parameters analysed for the biofilms formed by ATCC 33277 and ECR475 were significantly (p

    Techniques Used: Confocal Laser Scanning Microscopy, Mutagenesis

    Genomic arrangement of P. gingivalis ATCC 33277 in (A) the wild-type strain, (B) har mutant strain ECR455 and (C) har complemented strain ECR475. ‘P’ denotes promoter positions, the arrows above ‘P’ denote the direction of transcription whilst the stem loop following ermF indicates a Rho-independent transcriptional terminator. Not drawn to scale. (D) RT-PCR analysis of PGN_1504 and PGN_1503 ( har ). Reverse transcription of ECR455 and ECR475 RNA was performed using random hexamers. PCR was then performed using oligonucleotide primers specific for PGN_1504 (lanes 1–4) or PGN_1503 ( har ) (lanes 5–8 and 9–12). The templates used for PCR were: reverse transcribed ECR455 RNA (lanes 1 and 5), reverse transcribed ECR475 RNA (lane 9), RNA that was not reverse transcribed (lanes 2, 6 and 10), no template (lanes 3, 7 and 11) and P. gingivalis ATCC 33277 genomic DNA (lanes 4, 8 and 12). PGN_1504 transcript was detected in the har mutant ECR455 (lane 1), whilst PGN_1503 ( har ) transcript was not detected in the har mutant strain ECR455 (lane 5), but was detected in the har complemented strain ECR475 (lane 9). (E) Western blot detection of Har expression in P. gingivalis 33277, ECR455 and ECR475. Cytoplasmic protein extracts (25 µg) from P. gingivalis strains 33277 (B), ECR455 (C), ECR475 (D) and 5 ng purified Har (A) were separated on a 4–12% Bis-Tris polyacrylamide gel (Invitrogen) before Western transfer and blotting with anti-rHar sera. Har protein was detected in the 33277 wild-type and ECR475 complement, but not the ECR455 mutant strain.
    Figure Legend Snippet: Genomic arrangement of P. gingivalis ATCC 33277 in (A) the wild-type strain, (B) har mutant strain ECR455 and (C) har complemented strain ECR475. ‘P’ denotes promoter positions, the arrows above ‘P’ denote the direction of transcription whilst the stem loop following ermF indicates a Rho-independent transcriptional terminator. Not drawn to scale. (D) RT-PCR analysis of PGN_1504 and PGN_1503 ( har ). Reverse transcription of ECR455 and ECR475 RNA was performed using random hexamers. PCR was then performed using oligonucleotide primers specific for PGN_1504 (lanes 1–4) or PGN_1503 ( har ) (lanes 5–8 and 9–12). The templates used for PCR were: reverse transcribed ECR455 RNA (lanes 1 and 5), reverse transcribed ECR475 RNA (lane 9), RNA that was not reverse transcribed (lanes 2, 6 and 10), no template (lanes 3, 7 and 11) and P. gingivalis ATCC 33277 genomic DNA (lanes 4, 8 and 12). PGN_1504 transcript was detected in the har mutant ECR455 (lane 1), whilst PGN_1503 ( har ) transcript was not detected in the har mutant strain ECR455 (lane 5), but was detected in the har complemented strain ECR475 (lane 9). (E) Western blot detection of Har expression in P. gingivalis 33277, ECR455 and ECR475. Cytoplasmic protein extracts (25 µg) from P. gingivalis strains 33277 (B), ECR455 (C), ECR475 (D) and 5 ng purified Har (A) were separated on a 4–12% Bis-Tris polyacrylamide gel (Invitrogen) before Western transfer and blotting with anti-rHar sera. Har protein was detected in the 33277 wild-type and ECR475 complement, but not the ECR455 mutant strain.

    Techniques Used: Mutagenesis, Reverse Transcription Polymerase Chain Reaction, Polymerase Chain Reaction, Western Blot, Expressing, Purification

    20) Product Images from "The Porphyromonas gingivalis Ferric Uptake Regulator Orthologue Binds Hemin and Regulates Hemin-Responsive Biofilm Development"

    Article Title: The Porphyromonas gingivalis Ferric Uptake Regulator Orthologue Binds Hemin and Regulates Hemin-Responsive Biofilm Development

    Journal: PLoS ONE

    doi: 10.1371/journal.pone.0111168

    Genomic arrangement of P. gingivalis ATCC 33277 in (A) the wild-type strain, (B) har mutant strain ECR455 and (C) har complemented strain ECR475. ‘P’ denotes promoter positions, the arrows above ‘P’ denote the direction of transcription whilst the stem loop following ermF indicates a Rho-independent transcriptional terminator. Not drawn to scale. (D) RT-PCR analysis of PGN_1504 and PGN_1503 ( har ). Reverse transcription of ECR455 and ECR475 RNA was performed using random hexamers. PCR was then performed using oligonucleotide primers specific for PGN_1504 (lanes 1–4) or PGN_1503 ( har ) (lanes 5–8 and 9–12). The templates used for PCR were: reverse transcribed ECR455 RNA (lanes 1 and 5), reverse transcribed ECR475 RNA (lane 9), RNA that was not reverse transcribed (lanes 2, 6 and 10), no template (lanes 3, 7 and 11) and P. gingivalis ATCC 33277 genomic DNA (lanes 4, 8 and 12). PGN_1504 transcript was detected in the har mutant ECR455 (lane 1), whilst PGN_1503 ( har ) transcript was not detected in the har mutant strain ECR455 (lane 5), but was detected in the har complemented strain ECR475 (lane 9). (E) Western blot detection of Har expression in P. gingivalis 33277, ECR455 and ECR475. Cytoplasmic protein extracts (25 µg) from P. gingivalis strains 33277 (B), ECR455 (C), ECR475 (D) and 5 ng purified Har (A) were separated on a 4–12% Bis-Tris polyacrylamide gel (Invitrogen) before Western transfer and blotting with anti-rHar sera. Har protein was detected in the 33277 wild-type and ECR475 complement, but not the ECR455 mutant strain.
    Figure Legend Snippet: Genomic arrangement of P. gingivalis ATCC 33277 in (A) the wild-type strain, (B) har mutant strain ECR455 and (C) har complemented strain ECR475. ‘P’ denotes promoter positions, the arrows above ‘P’ denote the direction of transcription whilst the stem loop following ermF indicates a Rho-independent transcriptional terminator. Not drawn to scale. (D) RT-PCR analysis of PGN_1504 and PGN_1503 ( har ). Reverse transcription of ECR455 and ECR475 RNA was performed using random hexamers. PCR was then performed using oligonucleotide primers specific for PGN_1504 (lanes 1–4) or PGN_1503 ( har ) (lanes 5–8 and 9–12). The templates used for PCR were: reverse transcribed ECR455 RNA (lanes 1 and 5), reverse transcribed ECR475 RNA (lane 9), RNA that was not reverse transcribed (lanes 2, 6 and 10), no template (lanes 3, 7 and 11) and P. gingivalis ATCC 33277 genomic DNA (lanes 4, 8 and 12). PGN_1504 transcript was detected in the har mutant ECR455 (lane 1), whilst PGN_1503 ( har ) transcript was not detected in the har mutant strain ECR455 (lane 5), but was detected in the har complemented strain ECR475 (lane 9). (E) Western blot detection of Har expression in P. gingivalis 33277, ECR455 and ECR475. Cytoplasmic protein extracts (25 µg) from P. gingivalis strains 33277 (B), ECR455 (C), ECR475 (D) and 5 ng purified Har (A) were separated on a 4–12% Bis-Tris polyacrylamide gel (Invitrogen) before Western transfer and blotting with anti-rHar sera. Har protein was detected in the 33277 wild-type and ECR475 complement, but not the ECR455 mutant strain.

    Techniques Used: Mutagenesis, Reverse Transcription Polymerase Chain Reaction, Polymerase Chain Reaction, Western Blot, Expressing, Purification

    21) Product Images from "The Porphyromonas gingivalis Ferric Uptake Regulator Orthologue Binds Hemin and Regulates Hemin-Responsive Biofilm Development"

    Article Title: The Porphyromonas gingivalis Ferric Uptake Regulator Orthologue Binds Hemin and Regulates Hemin-Responsive Biofilm Development

    Journal: PLoS ONE

    doi: 10.1371/journal.pone.0111168

    P. gingivalis biofilm development. Orthogonal projections of CLSM images showing a representative region of the x-y plane over the depth of the biofilm in both xz and yz dimensions of the ATCC 33277 wild-type, har mutant ECR455 and har complement ECR 475 strains grown in excess hemin ( A ) or hemin-limitation ( B ). Comparison of the Biovolume ( C ), Average Thickness ( D ) and SA:Biovolume ( E ) calculated for each strain's biofilm growth in either excess hemin (dark bars) or limited hemin (light bars) over three independent experiments. All biometric parameters analysed for the biofilms formed by ATCC 33277 and ECR475 were significantly (p
    Figure Legend Snippet: P. gingivalis biofilm development. Orthogonal projections of CLSM images showing a representative region of the x-y plane over the depth of the biofilm in both xz and yz dimensions of the ATCC 33277 wild-type, har mutant ECR455 and har complement ECR 475 strains grown in excess hemin ( A ) or hemin-limitation ( B ). Comparison of the Biovolume ( C ), Average Thickness ( D ) and SA:Biovolume ( E ) calculated for each strain's biofilm growth in either excess hemin (dark bars) or limited hemin (light bars) over three independent experiments. All biometric parameters analysed for the biofilms formed by ATCC 33277 and ECR475 were significantly (p

    Techniques Used: Confocal Laser Scanning Microscopy, Mutagenesis

    Genomic arrangement of P. gingivalis ATCC 33277 in (A) the wild-type strain, (B) har mutant strain ECR455 and (C) har complemented strain ECR475. ‘P’ denotes promoter positions, the arrows above ‘P’ denote the direction of transcription whilst the stem loop following ermF indicates a Rho-independent transcriptional terminator. Not drawn to scale. (D) RT-PCR analysis of PGN_1504 and PGN_1503 ( har ). Reverse transcription of ECR455 and ECR475 RNA was performed using random hexamers. PCR was then performed using oligonucleotide primers specific for PGN_1504 (lanes 1–4) or PGN_1503 ( har ) (lanes 5–8 and 9–12). The templates used for PCR were: reverse transcribed ECR455 RNA (lanes 1 and 5), reverse transcribed ECR475 RNA (lane 9), RNA that was not reverse transcribed (lanes 2, 6 and 10), no template (lanes 3, 7 and 11) and P. gingivalis ATCC 33277 genomic DNA (lanes 4, 8 and 12). PGN_1504 transcript was detected in the har mutant ECR455 (lane 1), whilst PGN_1503 ( har ) transcript was not detected in the har mutant strain ECR455 (lane 5), but was detected in the har complemented strain ECR475 (lane 9). (E) Western blot detection of Har expression in P. gingivalis 33277, ECR455 and ECR475. Cytoplasmic protein extracts (25 µg) from P. gingivalis strains 33277 (B), ECR455 (C), ECR475 (D) and 5 ng purified Har (A) were separated on a 4–12% Bis-Tris polyacrylamide gel (Invitrogen) before Western transfer and blotting with anti-rHar sera. Har protein was detected in the 33277 wild-type and ECR475 complement, but not the ECR455 mutant strain.
    Figure Legend Snippet: Genomic arrangement of P. gingivalis ATCC 33277 in (A) the wild-type strain, (B) har mutant strain ECR455 and (C) har complemented strain ECR475. ‘P’ denotes promoter positions, the arrows above ‘P’ denote the direction of transcription whilst the stem loop following ermF indicates a Rho-independent transcriptional terminator. Not drawn to scale. (D) RT-PCR analysis of PGN_1504 and PGN_1503 ( har ). Reverse transcription of ECR455 and ECR475 RNA was performed using random hexamers. PCR was then performed using oligonucleotide primers specific for PGN_1504 (lanes 1–4) or PGN_1503 ( har ) (lanes 5–8 and 9–12). The templates used for PCR were: reverse transcribed ECR455 RNA (lanes 1 and 5), reverse transcribed ECR475 RNA (lane 9), RNA that was not reverse transcribed (lanes 2, 6 and 10), no template (lanes 3, 7 and 11) and P. gingivalis ATCC 33277 genomic DNA (lanes 4, 8 and 12). PGN_1504 transcript was detected in the har mutant ECR455 (lane 1), whilst PGN_1503 ( har ) transcript was not detected in the har mutant strain ECR455 (lane 5), but was detected in the har complemented strain ECR475 (lane 9). (E) Western blot detection of Har expression in P. gingivalis 33277, ECR455 and ECR475. Cytoplasmic protein extracts (25 µg) from P. gingivalis strains 33277 (B), ECR455 (C), ECR475 (D) and 5 ng purified Har (A) were separated on a 4–12% Bis-Tris polyacrylamide gel (Invitrogen) before Western transfer and blotting with anti-rHar sera. Har protein was detected in the 33277 wild-type and ECR475 complement, but not the ECR455 mutant strain.

    Techniques Used: Mutagenesis, Reverse Transcription Polymerase Chain Reaction, Polymerase Chain Reaction, Western Blot, Expressing, Purification

    22) Product Images from "Porphyromonas gingivalis can invade periodontal ligament stem cells"

    Article Title: Porphyromonas gingivalis can invade periodontal ligament stem cells

    Journal: BMC Microbiology

    doi: 10.1186/s12866-017-0950-5

    The efficiency of P. gingivalis ATCC 33277 infection in PDLSCs. The infection rate of P. gingivalis in PDLSCs at MOIs of 50, 100, 200, and 500 were 5.83%, 8.12%, 7.77% and 7.53%, respectively, according to the agar plate culture method. The efficiencies of P. gingivalis infection of PDLSCs at MOIs of 50, 100, 200, and 500 were 6.74%, 10.56%, 10.36% and 9.78%, respectively, according to the q-PCR method. The data are presented as the mean ± SD of triplicate experiments. *Significant difference ( P
    Figure Legend Snippet: The efficiency of P. gingivalis ATCC 33277 infection in PDLSCs. The infection rate of P. gingivalis in PDLSCs at MOIs of 50, 100, 200, and 500 were 5.83%, 8.12%, 7.77% and 7.53%, respectively, according to the agar plate culture method. The efficiencies of P. gingivalis infection of PDLSCs at MOIs of 50, 100, 200, and 500 were 6.74%, 10.56%, 10.36% and 9.78%, respectively, according to the q-PCR method. The data are presented as the mean ± SD of triplicate experiments. *Significant difference ( P

    Techniques Used: Infection, Polymerase Chain Reaction

    PDLSCs infected with P. gingivalis ATCC 33277 under transmission electron microscopy. The nucleus was large and round, and organelles were abundant in PDLSCs ( a ). P. gingivalis ATCC 33277 could invade PDLSCs after 2 h of incubation ( b , c - A ). Endocytic vacuoles were not found surrounding internalized P. gingivalis . The bumps observed were stretched membrane where the PDLSCs packaged P. gingivalis ATCC 33277 ( b , c - B )
    Figure Legend Snippet: PDLSCs infected with P. gingivalis ATCC 33277 under transmission electron microscopy. The nucleus was large and round, and organelles were abundant in PDLSCs ( a ). P. gingivalis ATCC 33277 could invade PDLSCs after 2 h of incubation ( b , c - A ). Endocytic vacuoles were not found surrounding internalized P. gingivalis . The bumps observed were stretched membrane where the PDLSCs packaged P. gingivalis ATCC 33277 ( b , c - B )

    Techniques Used: Infection, Transmission Assay, Electron Microscopy, Incubation

    23) Product Images from "Porphyromonas gingivalis can invade periodontal ligament stem cells"

    Article Title: Porphyromonas gingivalis can invade periodontal ligament stem cells

    Journal: BMC Microbiology

    doi: 10.1186/s12866-017-0950-5

    The efficiency of P. gingivalis ATCC 33277 infection in PDLSCs. The infection rate of P. gingivalis in PDLSCs at MOIs of 50, 100, 200, and 500 were 5.83%, 8.12%, 7.77% and 7.53%, respectively, according to the agar plate culture method. The efficiencies of P. gingivalis infection of PDLSCs at MOIs of 50, 100, 200, and 500 were 6.74%, 10.56%, 10.36% and 9.78%, respectively, according to the q-PCR method. The data are presented as the mean ± SD of triplicate experiments. *Significant difference ( P
    Figure Legend Snippet: The efficiency of P. gingivalis ATCC 33277 infection in PDLSCs. The infection rate of P. gingivalis in PDLSCs at MOIs of 50, 100, 200, and 500 were 5.83%, 8.12%, 7.77% and 7.53%, respectively, according to the agar plate culture method. The efficiencies of P. gingivalis infection of PDLSCs at MOIs of 50, 100, 200, and 500 were 6.74%, 10.56%, 10.36% and 9.78%, respectively, according to the q-PCR method. The data are presented as the mean ± SD of triplicate experiments. *Significant difference ( P

    Techniques Used: Infection, Polymerase Chain Reaction

    PDLSCs infected with P. gingivalis ATCC 33277 under transmission electron microscopy. The nucleus was large and round, and organelles were abundant in PDLSCs ( a ). P. gingivalis ATCC 33277 could invade PDLSCs after 2 h of incubation ( b , c - A ). Endocytic vacuoles were not found surrounding internalized P. gingivalis . The bumps observed were stretched membrane where the PDLSCs packaged P. gingivalis ATCC 33277 ( b , c - B )
    Figure Legend Snippet: PDLSCs infected with P. gingivalis ATCC 33277 under transmission electron microscopy. The nucleus was large and round, and organelles were abundant in PDLSCs ( a ). P. gingivalis ATCC 33277 could invade PDLSCs after 2 h of incubation ( b , c - A ). Endocytic vacuoles were not found surrounding internalized P. gingivalis . The bumps observed were stretched membrane where the PDLSCs packaged P. gingivalis ATCC 33277 ( b , c - B )

    Techniques Used: Infection, Transmission Assay, Electron Microscopy, Incubation

    24) Product Images from "Structural and functional characterization of shaft, anchor, and tip proteins of the Mfa1 fimbria from the periodontal pathogen Porphyromonas gingivalis"

    Article Title: Structural and functional characterization of shaft, anchor, and tip proteins of the Mfa1 fimbria from the periodontal pathogen Porphyromonas gingivalis

    Journal: Scientific Reports

    doi: 10.1038/s41598-018-20067-z

    Overall structures of Mfa1, Mfa2 and Mfa3. ( A ) The mature form of Mfa1 with the full-length C-terminus located in the first β-sheet (mMfa1). ( B ) The precursor form of Mfa1 with the final nine amino acid residues removed (pMfa1 Δ9 ). ( C ) The structure of Mfa2. ( D ) The structure of the truncated Mfa3 with the missing C-terminal domain depicted as an orange sphere (pMfa3). The RgpA/B cleavage sites are marked with arrows in (B) and ( D) . The cleavable N-terminal extensions (β1-strands) are depicted in magenta in pMfa1 Δ9 and Mfa3, and the uncleavable N-terminal extension in Mfa2 in green. N mature is shown in green (pMfa1Δ9, mMfa1 and pMfa3) and the C-terminal strand in mMfa1 is shown in orange. The structurally conserved tryptophan residue located between the N- and C-terminal domains are depicted as orange stick models in all structures. In Mfa2 the two disulphide bonds are shown as stick models. ( E ) Overall topology of the Mfa proteins: an N-terminal domain and a C-terminal domain consisting of two sheets each (sheets 1 and 2 and sheets 3 and 4, respectively). Upon maturation the β1β2-loop is cleaved and the β1-strand is removed. The C-terminal β-strand, depicted in red, is present in several of the Bacteroidetes fimbrial shaft proteins and can adopt different conformations.
    Figure Legend Snippet: Overall structures of Mfa1, Mfa2 and Mfa3. ( A ) The mature form of Mfa1 with the full-length C-terminus located in the first β-sheet (mMfa1). ( B ) The precursor form of Mfa1 with the final nine amino acid residues removed (pMfa1 Δ9 ). ( C ) The structure of Mfa2. ( D ) The structure of the truncated Mfa3 with the missing C-terminal domain depicted as an orange sphere (pMfa3). The RgpA/B cleavage sites are marked with arrows in (B) and ( D) . The cleavable N-terminal extensions (β1-strands) are depicted in magenta in pMfa1 Δ9 and Mfa3, and the uncleavable N-terminal extension in Mfa2 in green. N mature is shown in green (pMfa1Δ9, mMfa1 and pMfa3) and the C-terminal strand in mMfa1 is shown in orange. The structurally conserved tryptophan residue located between the N- and C-terminal domains are depicted as orange stick models in all structures. In Mfa2 the two disulphide bonds are shown as stick models. ( E ) Overall topology of the Mfa proteins: an N-terminal domain and a C-terminal domain consisting of two sheets each (sheets 1 and 2 and sheets 3 and 4, respectively). Upon maturation the β1β2-loop is cleaved and the β1-strand is removed. The C-terminal β-strand, depicted in red, is present in several of the Bacteroidetes fimbrial shaft proteins and can adopt different conformations.

    Techniques Used:

    Deletion of the nine final Mfa3 residues does not affect the polymerization or downstream proteins. Expression of ( A ) Mfa3 or ( B ) Mfa1 or ( C ) polymerization of Mfa1. Whole cell lysates were solubilized in SDS buffer, heated to 100 °C for 5 min ( A and B ) or 60 °C for 10 min ( C ). The samples were separated on SDS-PAGE, blotted to a PVDF membrane and probed with a polyclonal Mfa3 antibody ( A ) or an Mfa1 fimbriae antibody ( B and C ). Lanes: 1, JI-1 (positive control); 2, Δmfa3ΔfimA ; 3, + mfa3 ; 4, + mfa3ΔC ( A – C ). Effect of mfa3ΔC mutation on incorporation of accessory proteins ( D – F ). ( D ) SDS-PAGE of pure Mfa1 fimbriae. ( E ) Immunoblot analysis of pure Mfa1 fimbriae using an anti-Mfa3 antibody. ( F ), Immunoblot analysis of pure Mfa1 fimbriae using an anti-Mfa4 antibody Lanes: 1, JI-1 (positive control); 2, Δmfa3ΔfimA ; 3, + mfa3ΔC ( D – F ).
    Figure Legend Snippet: Deletion of the nine final Mfa3 residues does not affect the polymerization or downstream proteins. Expression of ( A ) Mfa3 or ( B ) Mfa1 or ( C ) polymerization of Mfa1. Whole cell lysates were solubilized in SDS buffer, heated to 100 °C for 5 min ( A and B ) or 60 °C for 10 min ( C ). The samples were separated on SDS-PAGE, blotted to a PVDF membrane and probed with a polyclonal Mfa3 antibody ( A ) or an Mfa1 fimbriae antibody ( B and C ). Lanes: 1, JI-1 (positive control); 2, Δmfa3ΔfimA ; 3, + mfa3 ; 4, + mfa3ΔC ( A – C ). Effect of mfa3ΔC mutation on incorporation of accessory proteins ( D – F ). ( D ) SDS-PAGE of pure Mfa1 fimbriae. ( E ) Immunoblot analysis of pure Mfa1 fimbriae using an anti-Mfa3 antibody. ( F ), Immunoblot analysis of pure Mfa1 fimbriae using an anti-Mfa4 antibody Lanes: 1, JI-1 (positive control); 2, Δmfa3ΔfimA ; 3, + mfa3ΔC ( D – F ).

    Techniques Used: Expressing, SDS Page, Positive Control, Mutagenesis

    Mfa2 C54 and C285 are involved in Mfa1-Mfa2 interaction. Whole cell lysates were solubilized in SDS buffer (+2BME) or (−2BME), heated to 80 °C for 5 min, separated on SDS-PAGE, and blotted to a PVDF membrane. The samples were probed with an Mfa2 polyclonal antibody. Lanes: 1, JI-1 (positive control); 2, Δmfa1Δfim ; 3, + mfa1 ; 4, + mfa1ΔC ; 5, Δmfa2ΔfimA (negative control); 6, + mfa2 ; 7, + mfa2C54A ; 8, + mfa2C285A .
    Figure Legend Snippet: Mfa2 C54 and C285 are involved in Mfa1-Mfa2 interaction. Whole cell lysates were solubilized in SDS buffer (+2BME) or (−2BME), heated to 80 °C for 5 min, separated on SDS-PAGE, and blotted to a PVDF membrane. The samples were probed with an Mfa2 polyclonal antibody. Lanes: 1, JI-1 (positive control); 2, Δmfa1Δfim ; 3, + mfa1 ; 4, + mfa1ΔC ; 5, Δmfa2ΔfimA (negative control); 6, + mfa2 ; 7, + mfa2C54A ; 8, + mfa2C285A .

    Techniques Used: SDS Page, Positive Control, Negative Control

    Surface representation of Mfa1. Mfa1 in surface representation in light green. The proline-rich region is illustrated in dark green. The left insert shows the proline-rich region coordinating the metal ion. The right insert shows Trp554 and the hydrogen bonds between Asp71(OD1 and OD2) and Arg236 (NE and NH2) and between Val556(O) and Arg236(NH1 and NH2). Prolines, metal coordinating residues and residues in the tryptophan pockets are labelled.
    Figure Legend Snippet: Surface representation of Mfa1. Mfa1 in surface representation in light green. The proline-rich region is illustrated in dark green. The left insert shows the proline-rich region coordinating the metal ion. The right insert shows Trp554 and the hydrogen bonds between Asp71(OD1 and OD2) and Arg236 (NE and NH2) and between Val556(O) and Arg236(NH1 and NH2). Prolines, metal coordinating residues and residues in the tryptophan pockets are labelled.

    Techniques Used:

    The last strand and Trp binding pocket of Mfa1 are involved in polymerization. Whole cell lysates were solubilized in SDS buffer and heated to: ( A ) 100 °C for 5 min, ( B ) 80 °C for 5 min, ( C ) 70 °C for 10 min, or ( D ) 60 °C for 10 min. The samples were separated on SDS-PAGE, blotted to a membrane and probed with a polyclonal Mfa1 fimbriae antibody. Lanes: 1, JI-1(positive control); 2, Δmfa1Δfim ; 3, + mfa1 ; 4, + mfa1ΔC ; 5, + mfa1R236A ; 6, + mfa1 W554A .
    Figure Legend Snippet: The last strand and Trp binding pocket of Mfa1 are involved in polymerization. Whole cell lysates were solubilized in SDS buffer and heated to: ( A ) 100 °C for 5 min, ( B ) 80 °C for 5 min, ( C ) 70 °C for 10 min, or ( D ) 60 °C for 10 min. The samples were separated on SDS-PAGE, blotted to a membrane and probed with a polyclonal Mfa1 fimbriae antibody. Lanes: 1, JI-1(positive control); 2, Δmfa1Δfim ; 3, + mfa1 ; 4, + mfa1ΔC ; 5, + mfa1R236A ; 6, + mfa1 W554A .

    Techniques Used: Binding Assay, SDS Page

    Fimbrial gene clusters, organization of proteins, and constructs. ( A ) Five genes encode each fimbria: Mfa1 constitutes the shaft, Mfa2 the anchor, and Mfa3-5 the tip proteins of Mfa1 fimbria (upper) while FimA, FimB and FimC-E constitute shaft, anchor and tip proteins respectively of the FimA fimbria (lower). ( B ) Mfa1, 3, and 4 start with a signal peptide, followed by a lipidated cysteine. The mature forms are obtained when RgpA/B cleaves the precursors at an exposed arginine (Mfa1, 3 and 4). Mfa2, the anchor protein, is not cleaved. (C) Schematic description of constructs used for crystallographic studies. All constructs used for crystallization screening are depicted as blue bars. The parts of the protein that were observed and modelled in the crystal structures are marked with vertical stripes.
    Figure Legend Snippet: Fimbrial gene clusters, organization of proteins, and constructs. ( A ) Five genes encode each fimbria: Mfa1 constitutes the shaft, Mfa2 the anchor, and Mfa3-5 the tip proteins of Mfa1 fimbria (upper) while FimA, FimB and FimC-E constitute shaft, anchor and tip proteins respectively of the FimA fimbria (lower). ( B ) Mfa1, 3, and 4 start with a signal peptide, followed by a lipidated cysteine. The mature forms are obtained when RgpA/B cleaves the precursors at an exposed arginine (Mfa1, 3 and 4). Mfa2, the anchor protein, is not cleaved. (C) Schematic description of constructs used for crystallographic studies. All constructs used for crystallization screening are depicted as blue bars. The parts of the protein that were observed and modelled in the crystal structures are marked with vertical stripes.

    Techniques Used: Construct, Crystallization Assay

    Effect of mfa1ΔC mutation on expression of tip proteins. Whole cell lysates and culture supernatant were solubilized in SDS buffer, heated at 100 °C for 5 min, separated on SDS-PAGE, and blotted to a membrane. Next, the samples were probed with polyclonal antibodies against ( A ) Mfa3, ( B ) Mfa4 or ( C ) Mfa5. Lanes: 1, JI-1 (positive control); 2, Δmfa1Δfim ; 3, + mfa1 ; 4, + mfa1ΔC . Asterisks indicate possible degradation products.
    Figure Legend Snippet: Effect of mfa1ΔC mutation on expression of tip proteins. Whole cell lysates and culture supernatant were solubilized in SDS buffer, heated at 100 °C for 5 min, separated on SDS-PAGE, and blotted to a membrane. Next, the samples were probed with polyclonal antibodies against ( A ) Mfa3, ( B ) Mfa4 or ( C ) Mfa5. Lanes: 1, JI-1 (positive control); 2, Δmfa1Δfim ; 3, + mfa1 ; 4, + mfa1ΔC . Asterisks indicate possible degradation products.

    Techniques Used: Mutagenesis, Expressing, SDS Page, Positive Control

    25) Product Images from "Structural and functional characterization of shaft, anchor, and tip proteins of the Mfa1 fimbria from the periodontal pathogen Porphyromonas gingivalis"

    Article Title: Structural and functional characterization of shaft, anchor, and tip proteins of the Mfa1 fimbria from the periodontal pathogen Porphyromonas gingivalis

    Journal: Scientific Reports

    doi: 10.1038/s41598-018-20067-z

    Overall structures of Mfa1, Mfa2 and Mfa3. ( A ) The mature form of Mfa1 with the full-length C-terminus located in the first β-sheet (mMfa1). ( B ) The precursor form of Mfa1 with the final nine amino acid residues removed (pMfa1 Δ9 ). ( C ) The structure of Mfa2. ( D ) The structure of the truncated Mfa3 with the missing C-terminal domain depicted as an orange sphere (pMfa3). The RgpA/B cleavage sites are marked with arrows in (B) and ( D) . The cleavable N-terminal extensions (β1-strands) are depicted in magenta in pMfa1 Δ9 and Mfa3, and the uncleavable N-terminal extension in Mfa2 in green. N mature is shown in green (pMfa1Δ9, mMfa1 and pMfa3) and the C-terminal strand in mMfa1 is shown in orange. The structurally conserved tryptophan residue located between the N- and C-terminal domains are depicted as orange stick models in all structures. In Mfa2 the two disulphide bonds are shown as stick models. ( E ) Overall topology of the Mfa proteins: an N-terminal domain and a C-terminal domain consisting of two sheets each (sheets 1 and 2 and sheets 3 and 4, respectively). Upon maturation the β1β2-loop is cleaved and the β1-strand is removed. The C-terminal β-strand, depicted in red, is present in several of the Bacteroidetes fimbrial shaft proteins and can adopt different conformations.
    Figure Legend Snippet: Overall structures of Mfa1, Mfa2 and Mfa3. ( A ) The mature form of Mfa1 with the full-length C-terminus located in the first β-sheet (mMfa1). ( B ) The precursor form of Mfa1 with the final nine amino acid residues removed (pMfa1 Δ9 ). ( C ) The structure of Mfa2. ( D ) The structure of the truncated Mfa3 with the missing C-terminal domain depicted as an orange sphere (pMfa3). The RgpA/B cleavage sites are marked with arrows in (B) and ( D) . The cleavable N-terminal extensions (β1-strands) are depicted in magenta in pMfa1 Δ9 and Mfa3, and the uncleavable N-terminal extension in Mfa2 in green. N mature is shown in green (pMfa1Δ9, mMfa1 and pMfa3) and the C-terminal strand in mMfa1 is shown in orange. The structurally conserved tryptophan residue located between the N- and C-terminal domains are depicted as orange stick models in all structures. In Mfa2 the two disulphide bonds are shown as stick models. ( E ) Overall topology of the Mfa proteins: an N-terminal domain and a C-terminal domain consisting of two sheets each (sheets 1 and 2 and sheets 3 and 4, respectively). Upon maturation the β1β2-loop is cleaved and the β1-strand is removed. The C-terminal β-strand, depicted in red, is present in several of the Bacteroidetes fimbrial shaft proteins and can adopt different conformations.

    Techniques Used:

    Deletion of the nine final Mfa3 residues does not affect the polymerization or downstream proteins. Expression of ( A ) Mfa3 or ( B ) Mfa1 or ( C ) polymerization of Mfa1. Whole cell lysates were solubilized in SDS buffer, heated to 100 °C for 5 min ( A and B ) or 60 °C for 10 min ( C ). The samples were separated on SDS-PAGE, blotted to a PVDF membrane and probed with a polyclonal Mfa3 antibody ( A ) or an Mfa1 fimbriae antibody ( B and C ). Lanes: 1, JI-1 (positive control); 2, Δmfa3ΔfimA ; 3, + mfa3 ; 4, + mfa3ΔC ( A – C ). Effect of mfa3ΔC mutation on incorporation of accessory proteins ( D – F ). ( D ) SDS-PAGE of pure Mfa1 fimbriae. ( E ) Immunoblot analysis of pure Mfa1 fimbriae using an anti-Mfa3 antibody. ( F ), Immunoblot analysis of pure Mfa1 fimbriae using an anti-Mfa4 antibody Lanes: 1, JI-1 (positive control); 2, Δmfa3ΔfimA ; 3, + mfa3ΔC ( D – F ).
    Figure Legend Snippet: Deletion of the nine final Mfa3 residues does not affect the polymerization or downstream proteins. Expression of ( A ) Mfa3 or ( B ) Mfa1 or ( C ) polymerization of Mfa1. Whole cell lysates were solubilized in SDS buffer, heated to 100 °C for 5 min ( A and B ) or 60 °C for 10 min ( C ). The samples were separated on SDS-PAGE, blotted to a PVDF membrane and probed with a polyclonal Mfa3 antibody ( A ) or an Mfa1 fimbriae antibody ( B and C ). Lanes: 1, JI-1 (positive control); 2, Δmfa3ΔfimA ; 3, + mfa3 ; 4, + mfa3ΔC ( A – C ). Effect of mfa3ΔC mutation on incorporation of accessory proteins ( D – F ). ( D ) SDS-PAGE of pure Mfa1 fimbriae. ( E ) Immunoblot analysis of pure Mfa1 fimbriae using an anti-Mfa3 antibody. ( F ), Immunoblot analysis of pure Mfa1 fimbriae using an anti-Mfa4 antibody Lanes: 1, JI-1 (positive control); 2, Δmfa3ΔfimA ; 3, + mfa3ΔC ( D – F ).

    Techniques Used: Expressing, SDS Page, Positive Control, Mutagenesis

    Mfa2 C54 and C285 are involved in Mfa1-Mfa2 interaction. Whole cell lysates were solubilized in SDS buffer (+2BME) or (−2BME), heated to 80 °C for 5 min, separated on SDS-PAGE, and blotted to a PVDF membrane. The samples were probed with an Mfa2 polyclonal antibody. Lanes: 1, JI-1 (positive control); 2, Δmfa1Δfim ; 3, + mfa1 ; 4, + mfa1ΔC ; 5, Δmfa2ΔfimA (negative control); 6, + mfa2 ; 7, + mfa2C54A ; 8, + mfa2C285A .
    Figure Legend Snippet: Mfa2 C54 and C285 are involved in Mfa1-Mfa2 interaction. Whole cell lysates were solubilized in SDS buffer (+2BME) or (−2BME), heated to 80 °C for 5 min, separated on SDS-PAGE, and blotted to a PVDF membrane. The samples were probed with an Mfa2 polyclonal antibody. Lanes: 1, JI-1 (positive control); 2, Δmfa1Δfim ; 3, + mfa1 ; 4, + mfa1ΔC ; 5, Δmfa2ΔfimA (negative control); 6, + mfa2 ; 7, + mfa2C54A ; 8, + mfa2C285A .

    Techniques Used: SDS Page, Positive Control, Negative Control

    Surface representation of Mfa1. Mfa1 in surface representation in light green. The proline-rich region is illustrated in dark green. The left insert shows the proline-rich region coordinating the metal ion. The right insert shows Trp554 and the hydrogen bonds between Asp71(OD1 and OD2) and Arg236 (NE and NH2) and between Val556(O) and Arg236(NH1 and NH2). Prolines, metal coordinating residues and residues in the tryptophan pockets are labelled.
    Figure Legend Snippet: Surface representation of Mfa1. Mfa1 in surface representation in light green. The proline-rich region is illustrated in dark green. The left insert shows the proline-rich region coordinating the metal ion. The right insert shows Trp554 and the hydrogen bonds between Asp71(OD1 and OD2) and Arg236 (NE and NH2) and between Val556(O) and Arg236(NH1 and NH2). Prolines, metal coordinating residues and residues in the tryptophan pockets are labelled.

    Techniques Used:

    The last strand and Trp binding pocket of Mfa1 are involved in polymerization. Whole cell lysates were solubilized in SDS buffer and heated to: ( A ) 100 °C for 5 min, ( B ) 80 °C for 5 min, ( C ) 70 °C for 10 min, or ( D ) 60 °C for 10 min. The samples were separated on SDS-PAGE, blotted to a membrane and probed with a polyclonal Mfa1 fimbriae antibody. Lanes: 1, JI-1(positive control); 2, Δmfa1Δfim ; 3, + mfa1 ; 4, + mfa1ΔC ; 5, + mfa1R236A ; 6, + mfa1 W554A .
    Figure Legend Snippet: The last strand and Trp binding pocket of Mfa1 are involved in polymerization. Whole cell lysates were solubilized in SDS buffer and heated to: ( A ) 100 °C for 5 min, ( B ) 80 °C for 5 min, ( C ) 70 °C for 10 min, or ( D ) 60 °C for 10 min. The samples were separated on SDS-PAGE, blotted to a membrane and probed with a polyclonal Mfa1 fimbriae antibody. Lanes: 1, JI-1(positive control); 2, Δmfa1Δfim ; 3, + mfa1 ; 4, + mfa1ΔC ; 5, + mfa1R236A ; 6, + mfa1 W554A .

    Techniques Used: Binding Assay, SDS Page

    Fimbrial gene clusters, organization of proteins, and constructs. ( A ) Five genes encode each fimbria: Mfa1 constitutes the shaft, Mfa2 the anchor, and Mfa3-5 the tip proteins of Mfa1 fimbria (upper) while FimA, FimB and FimC-E constitute shaft, anchor and tip proteins respectively of the FimA fimbria (lower). ( B ) Mfa1, 3, and 4 start with a signal peptide, followed by a lipidated cysteine. The mature forms are obtained when RgpA/B cleaves the precursors at an exposed arginine (Mfa1, 3 and 4). Mfa2, the anchor protein, is not cleaved. (C) Schematic description of constructs used for crystallographic studies. All constructs used for crystallization screening are depicted as blue bars. The parts of the protein that were observed and modelled in the crystal structures are marked with vertical stripes.
    Figure Legend Snippet: Fimbrial gene clusters, organization of proteins, and constructs. ( A ) Five genes encode each fimbria: Mfa1 constitutes the shaft, Mfa2 the anchor, and Mfa3-5 the tip proteins of Mfa1 fimbria (upper) while FimA, FimB and FimC-E constitute shaft, anchor and tip proteins respectively of the FimA fimbria (lower). ( B ) Mfa1, 3, and 4 start with a signal peptide, followed by a lipidated cysteine. The mature forms are obtained when RgpA/B cleaves the precursors at an exposed arginine (Mfa1, 3 and 4). Mfa2, the anchor protein, is not cleaved. (C) Schematic description of constructs used for crystallographic studies. All constructs used for crystallization screening are depicted as blue bars. The parts of the protein that were observed and modelled in the crystal structures are marked with vertical stripes.

    Techniques Used: Construct, Crystallization Assay

    Effect of mfa1ΔC mutation on expression of tip proteins. Whole cell lysates and culture supernatant were solubilized in SDS buffer, heated at 100 °C for 5 min, separated on SDS-PAGE, and blotted to a membrane. Next, the samples were probed with polyclonal antibodies against ( A ) Mfa3, ( B ) Mfa4 or ( C ) Mfa5. Lanes: 1, JI-1 (positive control); 2, Δmfa1Δfim ; 3, + mfa1 ; 4, + mfa1ΔC . Asterisks indicate possible degradation products.
    Figure Legend Snippet: Effect of mfa1ΔC mutation on expression of tip proteins. Whole cell lysates and culture supernatant were solubilized in SDS buffer, heated at 100 °C for 5 min, separated on SDS-PAGE, and blotted to a membrane. Next, the samples were probed with polyclonal antibodies against ( A ) Mfa3, ( B ) Mfa4 or ( C ) Mfa5. Lanes: 1, JI-1 (positive control); 2, Δmfa1Δfim ; 3, + mfa1 ; 4, + mfa1ΔC . Asterisks indicate possible degradation products.

    Techniques Used: Mutagenesis, Expressing, SDS Page, Positive Control

    26) Product Images from "Modulation of the Substitution Pattern of 5-Aryl-2-Aminoimidazoles Allows Fine-Tuning of Their Antibiofilm Activity Spectrum and Toxicity"

    Article Title: Modulation of the Substitution Pattern of 5-Aryl-2-Aminoimidazoles Allows Fine-Tuning of Their Antibiofilm Activity Spectrum and Toxicity

    Journal: Antimicrobial Agents and Chemotherapy

    doi: 10.1128/AAC.00035-16

    Structures of 5-Ar-2-AI-based compounds whose activities against a broad panel of monospecies and mixed-species biofilm models were tested in this study.
    Figure Legend Snippet: Structures of 5-Ar-2-AI-based compounds whose activities against a broad panel of monospecies and mixed-species biofilm models were tested in this study.

    Techniques Used:

    Synthesis and structures of eight novel N 1-,2 N -disubstituted 5-Ar-2-AIs tested against monospecies and mixed-species biofilms. MeOH, methanol; rt, room temperature. Percentages indicate compound yield.
    Figure Legend Snippet: Synthesis and structures of eight novel N 1-,2 N -disubstituted 5-Ar-2-AIs tested against monospecies and mixed-species biofilms. MeOH, methanol; rt, room temperature. Percentages indicate compound yield.

    Techniques Used:

    27) Product Images from "LptO (PG0027) Is Required for Lipid A 1-Phosphatase Activity in Porphyromonas gingivalis W50"

    Article Title: LptO (PG0027) Is Required for Lipid A 1-Phosphatase Activity in Porphyromonas gingivalis W50

    Journal: Journal of Bacteriology

    doi: 10.1128/JB.00751-16

    SEM of P. gingivalis W50 and the Δ PG0027 mutant strain. Samples were prepared for SEM as described in Materials and Methods. The scale bar represents 1 μm. P. gingivalis W50 shows the characteristic membrane blebbing forming OMVs, which is not present in the Δ PG0027 mutant strain.
    Figure Legend Snippet: SEM of P. gingivalis W50 and the Δ PG0027 mutant strain. Samples were prepared for SEM as described in Materials and Methods. The scale bar represents 1 μm. P. gingivalis W50 shows the characteristic membrane blebbing forming OMVs, which is not present in the Δ PG0027 mutant strain.

    Techniques Used: Mutagenesis

    TEM of P. gingivalis W50, the Δ PG0027 and Δ PG1051 ( waaL ) mutant strains, and the C Δ PG0027 strain. Samples were prepared for TEM as described in Materials and Methods. (A) The scale bar represents 200 nm. OMV formation in P. gingivalis W50 and the C Δ PG0027 strain is clearly visible as defined structures. (B) Close-up view of the outer surface layers of P. gingivalis W50, the Δ PG0027 and Δ PG1051 ( waaL ) mutant strains, and the C Δ PG0027 strain.
    Figure Legend Snippet: TEM of P. gingivalis W50, the Δ PG0027 and Δ PG1051 ( waaL ) mutant strains, and the C Δ PG0027 strain. Samples were prepared for TEM as described in Materials and Methods. (A) The scale bar represents 200 nm. OMV formation in P. gingivalis W50 and the C Δ PG0027 strain is clearly visible as defined structures. (B) Close-up view of the outer surface layers of P. gingivalis W50, the Δ PG0027 and Δ PG1051 ( waaL ) mutant strains, and the C Δ PG0027 strain.

    Techniques Used: Transmission Electron Microscopy, Mutagenesis

    MALDI TOF/TOF tandem mass spectrum of m / z 1,448 of lipid A from P. gingivalis W50 (A) and the Δ PG0027 mutant strain (B). Inset structures show the proposed phosphate positioning, and dashed lines and arrows indicate possible cleavage sites.
    Figure Legend Snippet: MALDI TOF/TOF tandem mass spectrum of m / z 1,448 of lipid A from P. gingivalis W50 (A) and the Δ PG0027 mutant strain (B). Inset structures show the proposed phosphate positioning, and dashed lines and arrows indicate possible cleavage sites.

    Techniques Used: Mutagenesis

    MALDI-TOF MS analysis of lipid A isolated from OMVs from P. gingivalis W50, the Δ PG0027 mutant strain, and the C Δ PG0027 strain. MALDI-TOF MS was performed in negative-ion mode as described in Materials and Methods. Boxes represent mono-P-triacyl, mono-P-tetraacyl, and phosphorylated-pentaacyl lipid A species.
    Figure Legend Snippet: MALDI-TOF MS analysis of lipid A isolated from OMVs from P. gingivalis W50, the Δ PG0027 mutant strain, and the C Δ PG0027 strain. MALDI-TOF MS was performed in negative-ion mode as described in Materials and Methods. Boxes represent mono-P-triacyl, mono-P-tetraacyl, and phosphorylated-pentaacyl lipid A species.

    Techniques Used: Mass Spectrometry, Isolation, Mutagenesis

    SDS-PAGE and Western blotting of proteins from E. coli DH5α cells containing pEXT20, pMFH15, and pWEL1 ( S . Typhimurium PagL cloned into pEXT20) versus anti-His and anti-PG0027 antibodies. (A) Proteins from EXT20 and MFH15 (NI, noninduced; I, induced) for 5 h or overnight were subjected to SDS-PAGE and Western blotting and probed with an anti-His antibody. (B) SDS-PAGE and Western blotting of membranes from EXT20 (M) and WEL1 (M) and samples from MFH15, namely, whole cells (WC), supernatants (SN), and membranes (M) probed with an anti-His antibody. Membranes from EXT20 (M), WEL1 (M), and MFH15 (M) were also probed with an anti-PG0027 antibody.
    Figure Legend Snippet: SDS-PAGE and Western blotting of proteins from E. coli DH5α cells containing pEXT20, pMFH15, and pWEL1 ( S . Typhimurium PagL cloned into pEXT20) versus anti-His and anti-PG0027 antibodies. (A) Proteins from EXT20 and MFH15 (NI, noninduced; I, induced) for 5 h or overnight were subjected to SDS-PAGE and Western blotting and probed with an anti-His antibody. (B) SDS-PAGE and Western blotting of membranes from EXT20 (M) and WEL1 (M) and samples from MFH15, namely, whole cells (WC), supernatants (SN), and membranes (M) probed with an anti-His antibody. Membranes from EXT20 (M), WEL1 (M), and MFH15 (M) were also probed with an anti-PG0027 antibody.

    Techniques Used: SDS Page, Western Blot, Clone Assay

    MALDI-TOF MS analysis of lipid A from P. gingivalis W50, the Δ PG0027 mutant strain, and the C Δ PG0027 and C Δ PG0027R strains. Negative-ion MALDI-TOF MS was performed on lipid A samples with norharmane as the matrix as described in Materials and Methods. Boxes with solid lines represent the mono-P-tetraacyl, mono-P-pentaacyl, and bis-P-pentaacyl lipid A clusters, whereas boxes with dashed lines represent the non-P-tetraacyl and non-P-pentaacyl lipid A clusters.
    Figure Legend Snippet: MALDI-TOF MS analysis of lipid A from P. gingivalis W50, the Δ PG0027 mutant strain, and the C Δ PG0027 and C Δ PG0027R strains. Negative-ion MALDI-TOF MS was performed on lipid A samples with norharmane as the matrix as described in Materials and Methods. Boxes with solid lines represent the mono-P-tetraacyl, mono-P-pentaacyl, and bis-P-pentaacyl lipid A clusters, whereas boxes with dashed lines represent the non-P-tetraacyl and non-P-pentaacyl lipid A clusters.

    Techniques Used: Mass Spectrometry, Mutagenesis

    Lipid A-modifying activity of S . Typhimurium PagL and P. gingivalis PG0027 expressed in E. coli with Salmonella LPS as the substrate. Salmonella LPS was incubated with PagL-expressing membranes (in E. coli ) and PG0027-expressing membranes (in E. coli ), and lipid A was isolated from the reaction mixture and analyzed by MALDI-TOF MS in linear negative-ion mode.
    Figure Legend Snippet: Lipid A-modifying activity of S . Typhimurium PagL and P. gingivalis PG0027 expressed in E. coli with Salmonella LPS as the substrate. Salmonella LPS was incubated with PagL-expressing membranes (in E. coli ) and PG0027-expressing membranes (in E. coli ), and lipid A was isolated from the reaction mixture and analyzed by MALDI-TOF MS in linear negative-ion mode.

    Techniques Used: Activity Assay, Incubation, Expressing, Isolation, Mass Spectrometry

    Properties of P. gingivalis W50, the Δ PG0027 mutant strain, and the C Δ PG0027 strain. (A) Growth in BHI broth. Samples were withdrawn at different time points, and the OD 540 was measured for 8 days. Curves: 1, W50; 2, Δ PG0027 mutant strain; 3, C Δ PG0027 strain. (B) Strains were grown in BHI broth as for panel A (solid line) or with the addition of 0.02% Tween 20 (dashed line). Curves: 1, W50; 2, Δ PG0027 mutant strain; 3, C Δ PG0027 strain. (C) Histogram showing OMV yields from P. gingivalis W50 and the Δ PG0027 mutant and C Δ PG0027 strains. Student's t test yielded a P value of
    Figure Legend Snippet: Properties of P. gingivalis W50, the Δ PG0027 mutant strain, and the C Δ PG0027 strain. (A) Growth in BHI broth. Samples were withdrawn at different time points, and the OD 540 was measured for 8 days. Curves: 1, W50; 2, Δ PG0027 mutant strain; 3, C Δ PG0027 strain. (B) Strains were grown in BHI broth as for panel A (solid line) or with the addition of 0.02% Tween 20 (dashed line). Curves: 1, W50; 2, Δ PG0027 mutant strain; 3, C Δ PG0027 strain. (C) Histogram showing OMV yields from P. gingivalis W50 and the Δ PG0027 mutant and C Δ PG0027 strains. Student's t test yielded a P value of

    Techniques Used: Mutagenesis

    Lipid A modification assays of PG0027-expressing membranes (in E. coli ) with P. gingivalis W50 LPS as the substrate. P. gingivalis W50 LPS was incubated with E. coli membranes containing EXT20 (top panel) or PG0027-expressing membranes (MFH15) (middle panel). Lipid A was isolated from the reaction mixture as described in Materials and Methods and analyzed by MALDI-TOF MS in linear negative-ion mode. The bottom panel shows lipid A isolated from E. coli DH5α cells. The arrows show E. coli lipid A. Intens. [a.u.], intensity in arbitrary units.
    Figure Legend Snippet: Lipid A modification assays of PG0027-expressing membranes (in E. coli ) with P. gingivalis W50 LPS as the substrate. P. gingivalis W50 LPS was incubated with E. coli membranes containing EXT20 (top panel) or PG0027-expressing membranes (MFH15) (middle panel). Lipid A was isolated from the reaction mixture as described in Materials and Methods and analyzed by MALDI-TOF MS in linear negative-ion mode. The bottom panel shows lipid A isolated from E. coli DH5α cells. The arrows show E. coli lipid A. Intens. [a.u.], intensity in arbitrary units.

    Techniques Used: Modification, Expressing, Incubation, Isolation, Mass Spectrometry

    MALDI TOF/TOF tandem mass spectrum of m / z 1,688 of lipid A from the P. gingivalis Δ PG0027 mutant strain. Inset structures show the proposed phosphate positioning, and dashed lines and arrows indicate possible cleavage sites.
    Figure Legend Snippet: MALDI TOF/TOF tandem mass spectrum of m / z 1,688 of lipid A from the P. gingivalis Δ PG0027 mutant strain. Inset structures show the proposed phosphate positioning, and dashed lines and arrows indicate possible cleavage sites.

    Techniques Used: Mutagenesis

    MALDI-TOF MS analysis of lipid A from the P. gingivalis Δ PG902 , Δ PG902 / PG0027 , Δ PG1711 , and Δ PG1711 / PG0027 mutant strains. Boxes with solid and dashed lines represent phosphorylated and nonphosphorylated species, respectively, as described in the legend to Fig. 4 .
    Figure Legend Snippet: MALDI-TOF MS analysis of lipid A from the P. gingivalis Δ PG902 , Δ PG902 / PG0027 , Δ PG1711 , and Δ PG1711 / PG0027 mutant strains. Boxes with solid and dashed lines represent phosphorylated and nonphosphorylated species, respectively, as described in the legend to Fig. 4 .

    Techniques Used: Mass Spectrometry, Mutagenesis

    Phosphatase activities of intact cells and sonicated supernatants of P. gingivalis strains. Phosphatase activities were measured with 4-nitro-phenylphosphate as the substrate as described in Materials and Methods and expressed as units based on the change in the A 405 /OD 600 ratio of intact cells and as units based on the change in the A 405 per milligram of protein in sonicated supernatants. Activities were measured at pHs 7.4, 7.8, 8.0, and 8.3 with either continuous or discontinuous assays. The activities obtained at pHs 7.8 and 8.0 are shown as histograms. Black bars, P. gingivalis W50; light gray bars, Δ PG0027 ; dark gray bars, C Δ PG0027R. P values (Student's t test) are indicated below the pairs. The activities obtained at pHs 7.4 and 8.3 and the P values determined from the data are shown at the bottom.
    Figure Legend Snippet: Phosphatase activities of intact cells and sonicated supernatants of P. gingivalis strains. Phosphatase activities were measured with 4-nitro-phenylphosphate as the substrate as described in Materials and Methods and expressed as units based on the change in the A 405 /OD 600 ratio of intact cells and as units based on the change in the A 405 per milligram of protein in sonicated supernatants. Activities were measured at pHs 7.4, 7.8, 8.0, and 8.3 with either continuous or discontinuous assays. The activities obtained at pHs 7.8 and 8.0 are shown as histograms. Black bars, P. gingivalis W50; light gray bars, Δ PG0027 ; dark gray bars, C Δ PG0027R. P values (Student's t test) are indicated below the pairs. The activities obtained at pHs 7.4 and 8.3 and the P values determined from the data are shown at the bottom.

    Techniques Used: Sonication

    28) Product Images from "Tolerance induced by Porphyromonas gingivalis may occur independently of TLR2 and TLR4"

    Article Title: Tolerance induced by Porphyromonas gingivalis may occur independently of TLR2 and TLR4

    Journal: PLoS ONE

    doi: 10.1371/journal.pone.0200946

    Interstrain variability of cytokine production in macrophages stimulated with P . gingivalis clinical isolates. Macrophages were incubated with 1x10 8 CFU/ml heat-inactiveted P . gingivalis clinical isolates (a total of 21 strains) for 24 h, washed and then restimulated with the same clinical isolates (1x10 8 CFU/ml) for another 24 h. TNF-α (A), IL-1β (B) and IL-10 (C) levels in the culture supernatants were determined by ELISA.
    Figure Legend Snippet: Interstrain variability of cytokine production in macrophages stimulated with P . gingivalis clinical isolates. Macrophages were incubated with 1x10 8 CFU/ml heat-inactiveted P . gingivalis clinical isolates (a total of 21 strains) for 24 h, washed and then restimulated with the same clinical isolates (1x10 8 CFU/ml) for another 24 h. TNF-α (A), IL-1β (B) and IL-10 (C) levels in the culture supernatants were determined by ELISA.

    Techniques Used: Incubation, Enzyme-linked Immunosorbent Assay

    Cytokine expression in macrophages stimulated with P . gingivalis . Macrophages were stimulated with medium, 1x10 8 CFU/ml heat-inactivated P . gingivalis clinical isolates, P . gingivalis ATCC 33277 or E . coli ATCC 25922 (positive control) for 24 h, washed and then challenged with medium, 1x10 8 CFU/ml P . gingivalis clinical isolates, P . gingivalis ATCC 33277 or E . coli ATCC 25922 for another 24 h. Expression levels of TNF-α (A), IL-1β (B) and IL-10 (C) in the cell-free supernatants were detected by ELISA. Data are presented as the mean±SD (n = 5 per group). *p
    Figure Legend Snippet: Cytokine expression in macrophages stimulated with P . gingivalis . Macrophages were stimulated with medium, 1x10 8 CFU/ml heat-inactivated P . gingivalis clinical isolates, P . gingivalis ATCC 33277 or E . coli ATCC 25922 (positive control) for 24 h, washed and then challenged with medium, 1x10 8 CFU/ml P . gingivalis clinical isolates, P . gingivalis ATCC 33277 or E . coli ATCC 25922 for another 24 h. Expression levels of TNF-α (A), IL-1β (B) and IL-10 (C) in the cell-free supernatants were detected by ELISA. Data are presented as the mean±SD (n = 5 per group). *p

    Techniques Used: Expressing, Positive Control, Enzyme-linked Immunosorbent Assay

    29) Product Images from "Tolerance induced by Porphyromonas gingivalis may occur independently of TLR2 and TLR4"

    Article Title: Tolerance induced by Porphyromonas gingivalis may occur independently of TLR2 and TLR4

    Journal: PLoS ONE

    doi: 10.1371/journal.pone.0200946

    Interstrain variability of cytokine production in macrophages stimulated with P . gingivalis clinical isolates. Macrophages were incubated with 1x10 8 CFU/ml heat-inactiveted P . gingivalis clinical isolates (a total of 21 strains) for 24 h, washed and then restimulated with the same clinical isolates (1x10 8 CFU/ml) for another 24 h. TNF-α (A), IL-1β (B) and IL-10 (C) levels in the culture supernatants were determined by ELISA.
    Figure Legend Snippet: Interstrain variability of cytokine production in macrophages stimulated with P . gingivalis clinical isolates. Macrophages were incubated with 1x10 8 CFU/ml heat-inactiveted P . gingivalis clinical isolates (a total of 21 strains) for 24 h, washed and then restimulated with the same clinical isolates (1x10 8 CFU/ml) for another 24 h. TNF-α (A), IL-1β (B) and IL-10 (C) levels in the culture supernatants were determined by ELISA.

    Techniques Used: Incubation, Enzyme-linked Immunosorbent Assay

    Cytokine expression in macrophages stimulated with P . gingivalis . Macrophages were stimulated with medium, 1x10 8 CFU/ml heat-inactivated P . gingivalis clinical isolates, P . gingivalis ATCC 33277 or E . coli ATCC 25922 (positive control) for 24 h, washed and then challenged with medium, 1x10 8 CFU/ml P . gingivalis clinical isolates, P . gingivalis ATCC 33277 or E . coli ATCC 25922 for another 24 h. Expression levels of TNF-α (A), IL-1β (B) and IL-10 (C) in the cell-free supernatants were detected by ELISA. Data are presented as the mean±SD (n = 5 per group). *p
    Figure Legend Snippet: Cytokine expression in macrophages stimulated with P . gingivalis . Macrophages were stimulated with medium, 1x10 8 CFU/ml heat-inactivated P . gingivalis clinical isolates, P . gingivalis ATCC 33277 or E . coli ATCC 25922 (positive control) for 24 h, washed and then challenged with medium, 1x10 8 CFU/ml P . gingivalis clinical isolates, P . gingivalis ATCC 33277 or E . coli ATCC 25922 for another 24 h. Expression levels of TNF-α (A), IL-1β (B) and IL-10 (C) in the cell-free supernatants were detected by ELISA. Data are presented as the mean±SD (n = 5 per group). *p

    Techniques Used: Expressing, Positive Control, Enzyme-linked Immunosorbent Assay

    30) Product Images from "Antimicrobial Activity of Piper marginatum Jacq and Ilex guayusa Loes on Microorganisms Associated with Periodontal Disease"

    Article Title: Antimicrobial Activity of Piper marginatum Jacq and Ilex guayusa Loes on Microorganisms Associated with Periodontal Disease

    Journal: International Journal of Microbiology

    doi: 10.1155/2018/4147383

    I. guayusa Loes and P. marginatum Jacq fraction antimicrobial activity against P. gingivalis ATCC 33277 at three concentrations. (a) Antimicrobial activities (inhibitory halos mm) from I. guayusa Loes and P. marginatum Jacq fractions against P. gingivalis. (b) 1: P. marginatum Jacq 4 mg/mL ethanol : water fraction, 19.0 mm inhibitory halo; 2: I. guayusa Loes 4 mg/mL acetone fraction, 12.3 mm inhibitory halo; 3: P. marginatum Jacq 4 mg/mL acetone fraction, 11.7 mm inhibitory halo; 4: I. guayusa Loes 4 mg/mL hexane fraction, 13.7 mm inhibitory halo; 5: 100 µ g/mL ampicillin positive control, 38.0 mm inhibitory halo.
    Figure Legend Snippet: I. guayusa Loes and P. marginatum Jacq fraction antimicrobial activity against P. gingivalis ATCC 33277 at three concentrations. (a) Antimicrobial activities (inhibitory halos mm) from I. guayusa Loes and P. marginatum Jacq fractions against P. gingivalis. (b) 1: P. marginatum Jacq 4 mg/mL ethanol : water fraction, 19.0 mm inhibitory halo; 2: I. guayusa Loes 4 mg/mL acetone fraction, 12.3 mm inhibitory halo; 3: P. marginatum Jacq 4 mg/mL acetone fraction, 11.7 mm inhibitory halo; 4: I. guayusa Loes 4 mg/mL hexane fraction, 13.7 mm inhibitory halo; 5: 100 µ g/mL ampicillin positive control, 38.0 mm inhibitory halo.

    Techniques Used: Activity Assay, Positive Control

    I. guayusa Loes and P. marginatum Jacq fraction antimicrobial activity against P. intermedia ATCC 25611 at three concentrations. (a) Antimicrobial activities (inhibitory halos mm) from I. guayusa Loes and P. marginatum Jacq fractions against P. intermedia. (b) 1: P. marginatum Jacq 4 mg/mL ethanol : water fraction, 12.3 mm inhibitory halo; 2: I. guayusa Loes 4 mg/mL acetone fraction, 12.0 mm inhibitory halo; 3: P. marginatum Jacq; 4 mg/mL acetone fraction, 11.3 mm inhibitory halo; 4: I. guayusa Loes 4 mg/mL hexane fraction, 13.3 mm inhibitory halo; 5: 50 IU/mL erythromycin positive control, 40 mm inhibitory halo.
    Figure Legend Snippet: I. guayusa Loes and P. marginatum Jacq fraction antimicrobial activity against P. intermedia ATCC 25611 at three concentrations. (a) Antimicrobial activities (inhibitory halos mm) from I. guayusa Loes and P. marginatum Jacq fractions against P. intermedia. (b) 1: P. marginatum Jacq 4 mg/mL ethanol : water fraction, 12.3 mm inhibitory halo; 2: I. guayusa Loes 4 mg/mL acetone fraction, 12.0 mm inhibitory halo; 3: P. marginatum Jacq; 4 mg/mL acetone fraction, 11.3 mm inhibitory halo; 4: I. guayusa Loes 4 mg/mL hexane fraction, 13.3 mm inhibitory halo; 5: 50 IU/mL erythromycin positive control, 40 mm inhibitory halo.

    Techniques Used: Activity Assay, Positive Control

    31) Product Images from "Antimicrobial Activity of Piper marginatum Jacq and Ilex guayusa Loes on Microorganisms Associated with Periodontal Disease"

    Article Title: Antimicrobial Activity of Piper marginatum Jacq and Ilex guayusa Loes on Microorganisms Associated with Periodontal Disease

    Journal: International Journal of Microbiology

    doi: 10.1155/2018/4147383

    I. guayusa Loes and P. marginatum Jacq fraction antimicrobial activity against P. gingivalis ATCC 33277 at three concentrations. (a) Antimicrobial activities (inhibitory halos mm) from I. guayusa Loes and P. marginatum Jacq fractions against P. gingivalis. (b) 1: P. marginatum Jacq 4 mg/mL ethanol : water fraction, 19.0 mm inhibitory halo; 2: I. guayusa Loes 4 mg/mL acetone fraction, 12.3 mm inhibitory halo; 3: P. marginatum Jacq 4 mg/mL acetone fraction, 11.7 mm inhibitory halo; 4: I. guayusa Loes 4 mg/mL hexane fraction, 13.7 mm inhibitory halo; 5: 100 µ g/mL ampicillin positive control, 38.0 mm inhibitory halo.
    Figure Legend Snippet: I. guayusa Loes and P. marginatum Jacq fraction antimicrobial activity against P. gingivalis ATCC 33277 at three concentrations. (a) Antimicrobial activities (inhibitory halos mm) from I. guayusa Loes and P. marginatum Jacq fractions against P. gingivalis. (b) 1: P. marginatum Jacq 4 mg/mL ethanol : water fraction, 19.0 mm inhibitory halo; 2: I. guayusa Loes 4 mg/mL acetone fraction, 12.3 mm inhibitory halo; 3: P. marginatum Jacq 4 mg/mL acetone fraction, 11.7 mm inhibitory halo; 4: I. guayusa Loes 4 mg/mL hexane fraction, 13.7 mm inhibitory halo; 5: 100 µ g/mL ampicillin positive control, 38.0 mm inhibitory halo.

    Techniques Used: Activity Assay, Positive Control

    I. guayusa Loes and P. marginatum Jacq fraction antimicrobial activity against P. intermedia ATCC 25611 at three concentrations. (a) Antimicrobial activities (inhibitory halos mm) from I. guayusa Loes and P. marginatum Jacq fractions against P. intermedia. (b) 1: P. marginatum Jacq 4 mg/mL ethanol : water fraction, 12.3 mm inhibitory halo; 2: I. guayusa Loes 4 mg/mL acetone fraction, 12.0 mm inhibitory halo; 3: P. marginatum Jacq; 4 mg/mL acetone fraction, 11.3 mm inhibitory halo; 4: I. guayusa Loes 4 mg/mL hexane fraction, 13.3 mm inhibitory halo; 5: 50 IU/mL erythromycin positive control, 40 mm inhibitory halo.
    Figure Legend Snippet: I. guayusa Loes and P. marginatum Jacq fraction antimicrobial activity against P. intermedia ATCC 25611 at three concentrations. (a) Antimicrobial activities (inhibitory halos mm) from I. guayusa Loes and P. marginatum Jacq fractions against P. intermedia. (b) 1: P. marginatum Jacq 4 mg/mL ethanol : water fraction, 12.3 mm inhibitory halo; 2: I. guayusa Loes 4 mg/mL acetone fraction, 12.0 mm inhibitory halo; 3: P. marginatum Jacq; 4 mg/mL acetone fraction, 11.3 mm inhibitory halo; 4: I. guayusa Loes 4 mg/mL hexane fraction, 13.3 mm inhibitory halo; 5: 50 IU/mL erythromycin positive control, 40 mm inhibitory halo.

    Techniques Used: Activity Assay, Positive Control

    32) Product Images from "Novel fimbrilin PGN_1808 in Porphyromonas gingivalis"

    Article Title: Novel fimbrilin PGN_1808 in Porphyromonas gingivalis

    Journal: PLoS ONE

    doi: 10.1371/journal.pone.0173541

    Transmission electron micrographs. Whole cells of the P . gingivalis mutant that was deficient in both FimA and Mfa1 fimbriae (A), whole cells of the PGN_1808-overexpressing mutant (B), and fractionated PGN_1808 (C) were negatively stained, then observed by TEM. Filamentous structures (2‒3 nm × 200‒400 nm) were observed on cell surfaces of the PGN_1808-overexpressing mutant (indicated by arrows) and in the PGN_1808 fraction. Scale bars indicate 200 nm.
    Figure Legend Snippet: Transmission electron micrographs. Whole cells of the P . gingivalis mutant that was deficient in both FimA and Mfa1 fimbriae (A), whole cells of the PGN_1808-overexpressing mutant (B), and fractionated PGN_1808 (C) were negatively stained, then observed by TEM. Filamentous structures (2‒3 nm × 200‒400 nm) were observed on cell surfaces of the PGN_1808-overexpressing mutant (indicated by arrows) and in the PGN_1808 fraction. Scale bars indicate 200 nm.

    Techniques Used: Transmission Assay, Mutagenesis, Staining, Transmission Electron Microscopy

    Fractionation of bacterial cell lysates. P . gingivalis TDC60 lysates were fractionated by anion-exchange chromatography after ammonium sulfate precipitation (A). Fractions 30 to 50 (10 μl each) were subjected to SDS-PAGE, followed by CBB staining (B) and western blotting using anti-FimA, anti-Mfa1, and anti-PGN_1808 antisera (C). We demonstrated reproducibility by repeating the fractionation experiment three times (data not shown). Numbers in left indicate molecular weights (kDa).
    Figure Legend Snippet: Fractionation of bacterial cell lysates. P . gingivalis TDC60 lysates were fractionated by anion-exchange chromatography after ammonium sulfate precipitation (A). Fractions 30 to 50 (10 μl each) were subjected to SDS-PAGE, followed by CBB staining (B) and western blotting using anti-FimA, anti-Mfa1, and anti-PGN_1808 antisera (C). We demonstrated reproducibility by repeating the fractionation experiment three times (data not shown). Numbers in left indicate molecular weights (kDa).

    Techniques Used: Fractionation, Chromatography, SDS Page, Staining, Western Blot

    33) Product Images from "Porphyromonas gingivalis Vesicles Enhance Attachment, and the Leucine-Rich Repeat BspA Protein Is Required for Invasion of Epithelial Cells by "Tannerella forsythia" "

    Article Title: Porphyromonas gingivalis Vesicles Enhance Attachment, and the Leucine-Rich Repeat BspA Protein Is Required for Invasion of Epithelial Cells by "Tannerella forsythia"

    Journal:

    doi: 10.1128/IAI.00062-06

    Attachment of T. forsythia 43037 (Tf) and BFM571 to KB cells in the absence and presence of P. gingivalis OMVs (Pgv). Attachment levels (means ± standard errors) were expressed as the percentage of attached bacteria relative to the total number
    Figure Legend Snippet: Attachment of T. forsythia 43037 (Tf) and BFM571 to KB cells in the absence and presence of P. gingivalis OMVs (Pgv). Attachment levels (means ± standard errors) were expressed as the percentage of attached bacteria relative to the total number

    Techniques Used:

    Coaggregation of T. forsythia in the presence of P. gingivalis and outer membrane vesicles purified from P. gingivalis . Data are representative of three independent experiments. Data points represent means ± standard errors of triplicate readings.
    Figure Legend Snippet: Coaggregation of T. forsythia in the presence of P. gingivalis and outer membrane vesicles purified from P. gingivalis . Data are representative of three independent experiments. Data points represent means ± standard errors of triplicate readings.

    Techniques Used: Purification

    Invasion by T. forsythia (Tf) of KB cells and dependence of invasion on the presence of P. gingivalis vesicles (Pgv). The invasion level was expressed as the percentage of bacteria retrieved following antibiotic killing of external bacteria and KB cell
    Figure Legend Snippet: Invasion by T. forsythia (Tf) of KB cells and dependence of invasion on the presence of P. gingivalis vesicles (Pgv). The invasion level was expressed as the percentage of bacteria retrieved following antibiotic killing of external bacteria and KB cell

    Techniques Used:

    34) Product Images from "Porphyromonas gingivalis Vesicles Enhance Attachment, and the Leucine-Rich Repeat BspA Protein Is Required for Invasion of Epithelial Cells by "Tannerella forsythia" "

    Article Title: Porphyromonas gingivalis Vesicles Enhance Attachment, and the Leucine-Rich Repeat BspA Protein Is Required for Invasion of Epithelial Cells by "Tannerella forsythia"

    Journal:

    doi: 10.1128/IAI.00062-06

    Attachment of T. forsythia 43037 (Tf) and BFM571 to KB cells in the absence and presence of P. gingivalis OMVs (Pgv). Attachment levels (means ± standard errors) were expressed as the percentage of attached bacteria relative to the total number
    Figure Legend Snippet: Attachment of T. forsythia 43037 (Tf) and BFM571 to KB cells in the absence and presence of P. gingivalis OMVs (Pgv). Attachment levels (means ± standard errors) were expressed as the percentage of attached bacteria relative to the total number

    Techniques Used:

    Coaggregation of T. forsythia in the presence of P. gingivalis and outer membrane vesicles purified from P. gingivalis . Data are representative of three independent experiments. Data points represent means ± standard errors of triplicate readings.
    Figure Legend Snippet: Coaggregation of T. forsythia in the presence of P. gingivalis and outer membrane vesicles purified from P. gingivalis . Data are representative of three independent experiments. Data points represent means ± standard errors of triplicate readings.

    Techniques Used: Purification

    35) Product Images from "Role for Gingipains in Porphyromonas gingivalis Traffic to Phagolysosomes and Survival in Human Aortic Endothelial Cells ▿"

    Article Title: Role for Gingipains in Porphyromonas gingivalis Traffic to Phagolysosomes and Survival in Human Aortic Endothelial Cells ▿

    Journal: Infection and Immunity

    doi: 10.1128/IAI.01013-06

    Effects of wortmannin and bafilomycin A 1 on viability of intracellular P. gingivalis ATCC 33277 in HAECs. (A) SDS-PAGE and immunoblot analysis of the intracellular protein levels of cathepsins B and D in uninfected and infected HAECs with ATCC 33277 or
    Figure Legend Snippet: Effects of wortmannin and bafilomycin A 1 on viability of intracellular P. gingivalis ATCC 33277 in HAECs. (A) SDS-PAGE and immunoblot analysis of the intracellular protein levels of cathepsins B and D in uninfected and infected HAECs with ATCC 33277 or

    Techniques Used: SDS Page, Infection

    Ultrastructural observations of P. gingivalis ATCC 33277- and KDP136-infected HAECs by thin-section electron microscopy. (A) At 2 h postinfection, most of the internalized ATCC 33277 organisms were enclosed by or fused with single or double membranous
    Figure Legend Snippet: Ultrastructural observations of P. gingivalis ATCC 33277- and KDP136-infected HAECs by thin-section electron microscopy. (A) At 2 h postinfection, most of the internalized ATCC 33277 organisms were enclosed by or fused with single or double membranous

    Techniques Used: Infection, Electron Microscopy

    Confocal microscopic images of intracellular P. gingivalis WT strains and KDP136 in infected cells. After infection with P. gingivalis (Pg) ATCC 33277 (A), 381 (B), W83 (C), and KDP136 (D) for 20 min, HAECs were washed and further cultured for the indicated
    Figure Legend Snippet: Confocal microscopic images of intracellular P. gingivalis WT strains and KDP136 in infected cells. After infection with P. gingivalis (Pg) ATCC 33277 (A), 381 (B), W83 (C), and KDP136 (D) for 20 min, HAECs were washed and further cultured for the indicated

    Techniques Used: Infection, Cell Culture

    Induction of autophagy in HAECs infected with P. gingivalis ATCC 33277 and KDP136. (A) Effect of starvation on LC3-staining patterns in HAECs. While the cells showed mainly diffuse staining for LC3, cells under starvation conditions for 2 h exhibited
    Figure Legend Snippet: Induction of autophagy in HAECs infected with P. gingivalis ATCC 33277 and KDP136. (A) Effect of starvation on LC3-staining patterns in HAECs. While the cells showed mainly diffuse staining for LC3, cells under starvation conditions for 2 h exhibited

    Techniques Used: Infection, Staining

    Immunoelectron microscopy of P. gingivalis -infected HAECs. After infection with ATCC 33277 and KDP136, HAECs were incubated in the absence of extracellular bacteria for the indicated times. The localization of cathepsin B, a lysosomal marker, was examined
    Figure Legend Snippet: Immunoelectron microscopy of P. gingivalis -infected HAECs. After infection with ATCC 33277 and KDP136, HAECs were incubated in the absence of extracellular bacteria for the indicated times. The localization of cathepsin B, a lysosomal marker, was examined

    Techniques Used: Immuno-Electron Microscopy, Infection, Incubation, Marker

    Viability of intracellular WT P. gingivalis (ATCC 33277 and 381) and KDP136 in HAECs. HAECs (1 × 10 5 cells per well) were infected with P. gingivalis ATCC 33277 (black bars), 381 (hatched bars), or KDP136 (white bars) for 20 min at an MOI of 10
    Figure Legend Snippet: Viability of intracellular WT P. gingivalis (ATCC 33277 and 381) and KDP136 in HAECs. HAECs (1 × 10 5 cells per well) were infected with P. gingivalis ATCC 33277 (black bars), 381 (hatched bars), or KDP136 (white bars) for 20 min at an MOI of 10

    Techniques Used: Infection

    36) Product Images from "Structure, function, and inhibition of a genomic/clinical variant of Porphyromonas gingivalis peptidylarginine deiminase"

    Article Title: Structure, function, and inhibition of a genomic/clinical variant of Porphyromonas gingivalis peptidylarginine deiminase

    Journal: Protein Science : A Publication of the Protein Society

    doi: 10.1002/pro.3571

    Citrullinating activity in cell cultures. Activity of P. gingivalis strains containing variants PPAD‐T1 (ATCC 33277), PPAD‐T2 (ATCC T2; strain ATCC 33277 after introducing mutations G 231 N, E 232 T, and N 235 D), and a control, in which the antibiotic cassette used in ATCC T2 was introduced in wild‐type ATCC 33277 (ATCC T1 Ctrl). The results are shown as relative activity determined in triplicates using three independent cultures of each strain and displayed as mean ± SD. The statistical difference between strains was analyzed by the analysis of variance; ns, not significant; *** P
    Figure Legend Snippet: Citrullinating activity in cell cultures. Activity of P. gingivalis strains containing variants PPAD‐T1 (ATCC 33277), PPAD‐T2 (ATCC T2; strain ATCC 33277 after introducing mutations G 231 N, E 232 T, and N 235 D), and a control, in which the antibiotic cassette used in ATCC T2 was introduced in wild‐type ATCC 33277 (ATCC T1 Ctrl). The results are shown as relative activity determined in triplicates using three independent cultures of each strain and displayed as mean ± SD. The statistical difference between strains was analyzed by the analysis of variance; ns, not significant; *** P

    Techniques Used: Activity Assay

    37) Product Images from "Strain-Specific Colonization Patterns and Serum Modulation of Multi-Species Oral Biofilm Development"

    Article Title: Strain-Specific Colonization Patterns and Serum Modulation of Multi-Species Oral Biofilm Development

    Journal: Anaerobe

    doi: 10.1016/j.anaerobe.2012.06.003

    Ability of avirulent P. gingivalis strains ATCC 33277 and 381 to integrate into 4h-old biofilm communities of health-associated species. Biofilms of A. oris and V. parvula or A. oris , V. parvula and F. nucleatum were allowed to develop for 4h in flow
    Figure Legend Snippet: Ability of avirulent P. gingivalis strains ATCC 33277 and 381 to integrate into 4h-old biofilm communities of health-associated species. Biofilms of A. oris and V. parvula or A. oris , V. parvula and F. nucleatum were allowed to develop for 4h in flow

    Techniques Used: Flow Cytometry

    Effect of serum on viability of A. oris (Ao), V. parvula (Vp), F. nucleatum ATCC 10953 (Fn) and P. gingivalis 381 (Pg). Planktonic growth of microorganisms was evaluated in their appropriate nutrient-rich medium supplemented with 10% native human serum
    Figure Legend Snippet: Effect of serum on viability of A. oris (Ao), V. parvula (Vp), F. nucleatum ATCC 10953 (Fn) and P. gingivalis 381 (Pg). Planktonic growth of microorganisms was evaluated in their appropriate nutrient-rich medium supplemented with 10% native human serum

    Techniques Used:

    Planktonic growth of F. nucleatum ATCC 10953 in salivary growth medium as a mono-culture or in co-culture with A. oris and V. parvula . Panel A shows planktonic growth curves. Data points represent measurements from three independent replicate experiments.
    Figure Legend Snippet: Planktonic growth of F. nucleatum ATCC 10953 in salivary growth medium as a mono-culture or in co-culture with A. oris and V. parvula . Panel A shows planktonic growth curves. Data points represent measurements from three independent replicate experiments.

    Techniques Used: Co-Culture Assay

    Ability of virulent P. gingivalis strains W50 and W83 to integrate into 4h-old biofilm communities of health-associated species. In panel A, biofilms of A. oris and V. parvula or A. oris , V. parvula and F. nucleatum were allowed to develop for 4h in flow
    Figure Legend Snippet: Ability of virulent P. gingivalis strains W50 and W83 to integrate into 4h-old biofilm communities of health-associated species. In panel A, biofilms of A. oris and V. parvula or A. oris , V. parvula and F. nucleatum were allowed to develop for 4h in flow

    Techniques Used: Flow Cytometry

    Ability of avirulent P. gingivalis strains ATCC 33277 and 381 to integrate into 4h-old biofilm communities of health-associated species. Biofilms of A. oris and V. parvula or A. oris , V. parvula and F. nucleatum were allowed to develop for 4h in flow
    Figure Legend Snippet: Ability of avirulent P. gingivalis strains ATCC 33277 and 381 to integrate into 4h-old biofilm communities of health-associated species. Biofilms of A. oris and V. parvula or A. oris , V. parvula and F. nucleatum were allowed to develop for 4h in flow

    Techniques Used: Flow Cytometry

    Effect of serum on viability of A. oris (Ao), V. parvula (Vp), F. nucleatum ATCC 10953 (Fn) and P. gingivalis 381 (Pg). Planktonic growth of microorganisms was evaluated in their appropriate nutrient-rich medium supplemented with 10% native human serum
    Figure Legend Snippet: Effect of serum on viability of A. oris (Ao), V. parvula (Vp), F. nucleatum ATCC 10953 (Fn) and P. gingivalis 381 (Pg). Planktonic growth of microorganisms was evaluated in their appropriate nutrient-rich medium supplemented with 10% native human serum

    Techniques Used:

    Ability of virulent P. gingivalis strains W50 and W83 to integrate into 4h-old biofilm communities of health-associated species. In panel A, biofilms of A. oris and V. parvula or A. oris , V. parvula and F. nucleatum were allowed to develop for 4h in flow
    Figure Legend Snippet: Ability of virulent P. gingivalis strains W50 and W83 to integrate into 4h-old biofilm communities of health-associated species. In panel A, biofilms of A. oris and V. parvula or A. oris , V. parvula and F. nucleatum were allowed to develop for 4h in flow

    Techniques Used: Flow Cytometry

    38) Product Images from "Strain-Specific Colonization Patterns and Serum Modulation of Multi-Species Oral Biofilm Development"

    Article Title: Strain-Specific Colonization Patterns and Serum Modulation of Multi-Species Oral Biofilm Development

    Journal: Anaerobe

    doi: 10.1016/j.anaerobe.2012.06.003

    Effect of serum on viability of A. oris (Ao), V. parvula (Vp), F. nucleatum ATCC 10953 (Fn) and P. gingivalis 381 (Pg). Planktonic growth of microorganisms was evaluated in their appropriate nutrient-rich medium supplemented with 10% native human serum
    Figure Legend Snippet: Effect of serum on viability of A. oris (Ao), V. parvula (Vp), F. nucleatum ATCC 10953 (Fn) and P. gingivalis 381 (Pg). Planktonic growth of microorganisms was evaluated in their appropriate nutrient-rich medium supplemented with 10% native human serum

    Techniques Used:

    39) Product Images from "Filifactor alocis Has Virulence Attributes That Can Enhance Its Persistence under Oxidative Stress Conditions and Mediate Invasion of Epithelial Cells by Porphyromonas gingivalis ▿ ▿ †"

    Article Title: Filifactor alocis Has Virulence Attributes That Can Enhance Its Persistence under Oxidative Stress Conditions and Mediate Invasion of Epithelial Cells by Porphyromonas gingivalis ▿ ▿ †

    Journal: Infection and Immunity

    doi: 10.1128/IAI.05631-11

    Biofilm formation by mono- and cocultures of Filifactor alocis and P. gingivalis ATCC 33277 (PG). Overnight cultures of F. alocis and/or P. gingivalis were diluted with fresh BHI medium to obtain a concentration of 5 × 10 7 CFU/ml. The cells were
    Figure Legend Snippet: Biofilm formation by mono- and cocultures of Filifactor alocis and P. gingivalis ATCC 33277 (PG). Overnight cultures of F. alocis and/or P. gingivalis were diluted with fresh BHI medium to obtain a concentration of 5 × 10 7 CFU/ml. The cells were

    Techniques Used: Concentration Assay

    40) Product Images from "Inhibition of Porphyromonas gingivalis-induced periodontal bone loss by CXCR4 antagonist treatment"

    Article Title: Inhibition of Porphyromonas gingivalis-induced periodontal bone loss by CXCR4 antagonist treatment

    Journal: Molecular oral microbiology

    doi: 10.1111/j.2041-1014.2012.00657.x

    Effect of AMD3100 on the numbers of P. gingivalis or total bacteria in the murine periodontal tissue
    Figure Legend Snippet: Effect of AMD3100 on the numbers of P. gingivalis or total bacteria in the murine periodontal tissue

    Techniques Used:

    Preventive treatment with AMD3100 abrogates P. gingivalis -induced periodontal bone loss
    Figure Legend Snippet: Preventive treatment with AMD3100 abrogates P. gingivalis -induced periodontal bone loss

    Techniques Used:

    Therapeutic treatment with AMD3100 inhibits P. gingivalis -induced periodontal bone loss
    Figure Legend Snippet: Therapeutic treatment with AMD3100 inhibits P. gingivalis -induced periodontal bone loss

    Techniques Used:

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    Article Snippet: .. Scanning Electron Microscopy In order to visualize the structural changes of E. faecalis biofilms after exposure to ClyR, E. faecalis ATCC 51299 biofilms grown on glass coverslips for 24 h were washed twice with PBS (to remove the planktonic cells), and treated with 50 μg/mL ClyR for 1 h. Biofilms were then washed twice with PBS, fixed with 2.5% glutaraldehyde overnight at 4 °C, dehydrated by granted ethanol (from 30% to 100%), and analyzed by a SU8010 scanning electron microscope (SEM, Hitachi, Tokyo, Japan). ..

    Positive Control:

    Article Title: Biofilm Formation by ica-Negative Ocular Isolates of Staphylococcus haemolyticus
    Article Snippet: .. One of the S. haemolyticus from healthy conjunctiva (SHN65) formed better biofilm and was comparable to that of the positive control strain of S. epidermidis ATCC 35984, but the composition of the matrix was different. ..

    Cell Culture:

    Article Title: Impact of Porphyromonas gingivalis Peptidylarginine Deiminase on Bacterial Biofilm Formation, Epithelial Cell Invasion, and Epithelial Cell Transcriptional Landscape
    Article Snippet: .. Bacterial strains and mammalian cell culture Wild-type (WT) P. gingivalis strain ATCC 33277, its isogenic mutants with a deleted PPAD gene (Δppad ) and expressing catalytically inactive PPAD (C351A, PPADC351A ), and T. forsythia ATCC 43037, F. nucleatum ATCC 25586, A. naeslundii ATCC 12104, and S. gordonii ATCC 10558 were used in this study. .. T. forsythia was grown on ATCC 1921-NAM agar, and the remaining strains were pre-cultivated on blood agar.

    Produced:

    Article Title: Inhibition of Candida albicans biofilm development by unencapsulated Enterococcus faecalis cps2
    Article Snippet: .. At this maturation stage, the number of viable cells detected within this biofilm was significantly reduced by approximately 50%, compared with the control (P < 0.005), because of the addition of spent medium produced by either unencapsulated E. faecalis cps2 or the reference strain ATCC 29212. .. Our data also found that 10 μg/mL protein concentration of the spent medium was sufficient to reduce biofilm mass, and a similar trend was observed when using a higher protein concentration (100 μg/mL; not shown).

    other:

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    Article Snippet: In addition, P . gulae D049 (type C) and P . gingivalis OMZ314 (type II) showed invasion at a higher level of efficiency than P . gingivalis ATCC 33277 (type I) ( ).

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    Microscopy:

    Article Title: Effects of a Chimeric Lysin against Planktonic and Sessile Enterococcus faecalis Hint at Potential Application in Endodontic Therapy
    Article Snippet: .. Scanning Electron Microscopy In order to visualize the structural changes of E. faecalis biofilms after exposure to ClyR, E. faecalis ATCC 51299 biofilms grown on glass coverslips for 24 h were washed twice with PBS (to remove the planktonic cells), and treated with 50 μg/mL ClyR for 1 h. Biofilms were then washed twice with PBS, fixed with 2.5% glutaraldehyde overnight at 4 °C, dehydrated by granted ethanol (from 30% to 100%), and analyzed by a SU8010 scanning electron microscope (SEM, Hitachi, Tokyo, Japan). ..

    Expressing:

    Article Title: Impact of Porphyromonas gingivalis Peptidylarginine Deiminase on Bacterial Biofilm Formation, Epithelial Cell Invasion, and Epithelial Cell Transcriptional Landscape
    Article Snippet: .. Bacterial strains and mammalian cell culture Wild-type (WT) P. gingivalis strain ATCC 33277, its isogenic mutants with a deleted PPAD gene (Δppad ) and expressing catalytically inactive PPAD (C351A, PPADC351A ), and T. forsythia ATCC 43037, F. nucleatum ATCC 25586, A. naeslundii ATCC 12104, and S. gordonii ATCC 10558 were used in this study. .. T. forsythia was grown on ATCC 1921-NAM agar, and the remaining strains were pre-cultivated on blood agar.

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    ATCC p gingivalis genomes
    Clustering by spacer content in CRISPR type 36.2 of Porphyromonas <t>gingivalis</t> . In type 36.2, the presence of each unique spacer is shown using a heatmap. The dendrogram was constructed from Euclidian distances. In the heatmap, the boxes indicate unique spacers and are arrayed horizontally. In the heatmap, 2 colors were used according to the bit score; red: ≥50, yellow:
    P Gingivalis Genomes, supplied by ATCC, used in various techniques. Bioz Stars score: 85/100, based on 1 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
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    92
    ATCC p gingivalis
    Attachment of T. forsythia 43037 (Tf) and BFM571 to KB cells in the absence and presence of P. <t>gingivalis</t> OMVs (Pgv). Attachment levels (means ± standard errors) were expressed as the percentage of attached bacteria relative to the total number
    P Gingivalis, supplied by ATCC, used in various techniques. Bioz Stars score: 92/100, based on 43 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
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    85
    ATCC p gingivalis omvs
    Attachment of T. forsythia 43037 (Tf) and BFM571 to KB cells in the absence and presence of P. <t>gingivalis</t> <t>OMVs</t> (Pgv). Attachment levels (means ± standard errors) were expressed as the percentage of attached bacteria relative to the total number
    P Gingivalis Omvs, supplied by ATCC, used in various techniques. Bioz Stars score: 85/100, based on 4 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
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    Clustering by spacer content in CRISPR type 36.2 of Porphyromonas gingivalis . In type 36.2, the presence of each unique spacer is shown using a heatmap. The dendrogram was constructed from Euclidian distances. In the heatmap, the boxes indicate unique spacers and are arrayed horizontally. In the heatmap, 2 colors were used according to the bit score; red: ≥50, yellow:

    Journal: Genome Biology and Evolution

    Article Title: CRISPR Regulation of Intraspecies Diversification by Limiting IS Transposition and Intercellular Recombination

    doi: 10.1093/gbe/evt075

    Figure Lengend Snippet: Clustering by spacer content in CRISPR type 36.2 of Porphyromonas gingivalis . In type 36.2, the presence of each unique spacer is shown using a heatmap. The dendrogram was constructed from Euclidian distances. In the heatmap, the boxes indicate unique spacers and are arrayed horizontally. In the heatmap, 2 colors were used according to the bit score; red: ≥50, yellow:

    Article Snippet: Our preliminary investigation of CRISPRs in three P. gingivalis strains demonstrated that CRISPR spacers exhibiting high nucleotide similarity to regions of P. gingivalis genomes were present and the number of spacers was diverse among the three genomes (TDC60: 89; W83: 44; and ATCC 33277: 137).

    Techniques: CRISPR, Construct

    Characteristics of recombination breakpoints among three Porphyromonas gingivalis genomes. ( A ) Fragments are shown in the alignment of two genome sequences (TDC60, ATCC 33277). The positions of MGEs or rRNA operons in the breakpoint gaps are indicated by colored broken lines, connecting the gaps and the bars (indicating the positions of the features on the genome), which are arrayed along the outside of the plot area. The red boxes on the plot area are the regions shown in ( B ) in detail. ( B ) Breakpoint gaps of TDC60 are enlarged in light gray areas surrounded by broken lines. The regions of ATCC 33277, which correspond to the enlarged gap of TDC60, are enlarged similarly. The fragments in TDC60 and ATCC 33277 are colored by red and deep blue, respectively. The regions exhibiting high nucleotide similarity to each other are shown by a yellow belt between two fragments. The 3-kb regions of the breakpoints are indicated by dark gray rectangles on the upper or lower side of the fragments. (i) rRNA operons in the breakpoint gap. The black arrows indicate rRNA genes. The light blue-filled boxes with arrows inside indicate ISs. (ii) ISs in the breakpoint gap. ( C ) The number of each feature in the breakpoint gap is plotted. The regions without any characteristic features are included under “Others.” The mean and standard deviations are provided by the horizontal and vertical lines, respectively. Statistical significance is indicated by an asterisk ( P

    Journal: Genome Biology and Evolution

    Article Title: CRISPR Regulation of Intraspecies Diversification by Limiting IS Transposition and Intercellular Recombination

    doi: 10.1093/gbe/evt075

    Figure Lengend Snippet: Characteristics of recombination breakpoints among three Porphyromonas gingivalis genomes. ( A ) Fragments are shown in the alignment of two genome sequences (TDC60, ATCC 33277). The positions of MGEs or rRNA operons in the breakpoint gaps are indicated by colored broken lines, connecting the gaps and the bars (indicating the positions of the features on the genome), which are arrayed along the outside of the plot area. The red boxes on the plot area are the regions shown in ( B ) in detail. ( B ) Breakpoint gaps of TDC60 are enlarged in light gray areas surrounded by broken lines. The regions of ATCC 33277, which correspond to the enlarged gap of TDC60, are enlarged similarly. The fragments in TDC60 and ATCC 33277 are colored by red and deep blue, respectively. The regions exhibiting high nucleotide similarity to each other are shown by a yellow belt between two fragments. The 3-kb regions of the breakpoints are indicated by dark gray rectangles on the upper or lower side of the fragments. (i) rRNA operons in the breakpoint gap. The black arrows indicate rRNA genes. The light blue-filled boxes with arrows inside indicate ISs. (ii) ISs in the breakpoint gap. ( C ) The number of each feature in the breakpoint gap is plotted. The regions without any characteristic features are included under “Others.” The mean and standard deviations are provided by the horizontal and vertical lines, respectively. Statistical significance is indicated by an asterisk ( P

    Article Snippet: Our preliminary investigation of CRISPRs in three P. gingivalis strains demonstrated that CRISPR spacers exhibiting high nucleotide similarity to regions of P. gingivalis genomes were present and the number of spacers was diverse among the three genomes (TDC60: 89; W83: 44; and ATCC 33277: 137).

    Techniques:

    Split network of 60 Porphyromonas gingivalis isolates obtained from concatenated seven loci sequences. A split network tree based upon the MLST data is shown. Circles indicate external nodes (each isolate) and are colored according to geographic origin (black: Japan; outlined: overseas or unspecified). fimA types are shown by light gray shadows. The numbers outside the isolate’s name indicate the patient source. Eleven colors are used to emphasize the clusters.

    Journal: Genome Biology and Evolution

    Article Title: CRISPR Regulation of Intraspecies Diversification by Limiting IS Transposition and Intercellular Recombination

    doi: 10.1093/gbe/evt075

    Figure Lengend Snippet: Split network of 60 Porphyromonas gingivalis isolates obtained from concatenated seven loci sequences. A split network tree based upon the MLST data is shown. Circles indicate external nodes (each isolate) and are colored according to geographic origin (black: Japan; outlined: overseas or unspecified). fimA types are shown by light gray shadows. The numbers outside the isolate’s name indicate the patient source. Eleven colors are used to emphasize the clusters.

    Article Snippet: Our preliminary investigation of CRISPRs in three P. gingivalis strains demonstrated that CRISPR spacers exhibiting high nucleotide similarity to regions of P. gingivalis genomes were present and the number of spacers was diverse among the three genomes (TDC60: 89; W83: 44; and ATCC 33277: 137).

    Techniques:

    Regions exhibiting high nucleotide similarity to P. gingivalis CRISPR spacers. Two examples of the 19 spacers exhibiting high nucleotide similarity to the P. gingivalis genome are shown. The white and black arrows indicate CDSs and rRNA genes, respectively. The arrows within the light blue-filled boxes indicate ISs. The orange regions indicate the sequences exhibiting high nucleotide similarity to CRISPR spacers. (i) Region exhibiting high nucleotide similarity to spacer 37_259: the transposase gene in IS Pg2 , in the TDC60 genome. (ii) Region exhibiting high nucleotide similarity to spacer 37_90: close to the IS both 2-kb upstream and 2-kb downstream in the 3 genomes.

    Journal: Genome Biology and Evolution

    Article Title: CRISPR Regulation of Intraspecies Diversification by Limiting IS Transposition and Intercellular Recombination

    doi: 10.1093/gbe/evt075

    Figure Lengend Snippet: Regions exhibiting high nucleotide similarity to P. gingivalis CRISPR spacers. Two examples of the 19 spacers exhibiting high nucleotide similarity to the P. gingivalis genome are shown. The white and black arrows indicate CDSs and rRNA genes, respectively. The arrows within the light blue-filled boxes indicate ISs. The orange regions indicate the sequences exhibiting high nucleotide similarity to CRISPR spacers. (i) Region exhibiting high nucleotide similarity to spacer 37_259: the transposase gene in IS Pg2 , in the TDC60 genome. (ii) Region exhibiting high nucleotide similarity to spacer 37_90: close to the IS both 2-kb upstream and 2-kb downstream in the 3 genomes.

    Article Snippet: Our preliminary investigation of CRISPRs in three P. gingivalis strains demonstrated that CRISPR spacers exhibiting high nucleotide similarity to regions of P. gingivalis genomes were present and the number of spacers was diverse among the three genomes (TDC60: 89; W83: 44; and ATCC 33277: 137).

    Techniques: CRISPR

    Spacer contents of Porphyromonas gingivalis isolates from seven patients in four CRISPR loci. Spacer arrays of 26 isolates from 7 patients are shown at each CRISPR locus. Each box indicates one spacer. The spacers in the arrays exhibit high nucleotide similarity to each other among the isolates if they are aligned vertically and have the same color. Blank boxes indicate absent spacers in the particular isolates. In patient no. 2, two colors are used because the D5 isolate has a type 30 spacer array that is distinct from those of D8 and D9. The spacers in type 36.2, shared among seven isolates of three patients, are indicated by deep yellow boxes and emphasized by dark gray belts.

    Journal: Genome Biology and Evolution

    Article Title: CRISPR Regulation of Intraspecies Diversification by Limiting IS Transposition and Intercellular Recombination

    doi: 10.1093/gbe/evt075

    Figure Lengend Snippet: Spacer contents of Porphyromonas gingivalis isolates from seven patients in four CRISPR loci. Spacer arrays of 26 isolates from 7 patients are shown at each CRISPR locus. Each box indicates one spacer. The spacers in the arrays exhibit high nucleotide similarity to each other among the isolates if they are aligned vertically and have the same color. Blank boxes indicate absent spacers in the particular isolates. In patient no. 2, two colors are used because the D5 isolate has a type 30 spacer array that is distinct from those of D8 and D9. The spacers in type 36.2, shared among seven isolates of three patients, are indicated by deep yellow boxes and emphasized by dark gray belts.

    Article Snippet: Our preliminary investigation of CRISPRs in three P. gingivalis strains demonstrated that CRISPR spacers exhibiting high nucleotide similarity to regions of P. gingivalis genomes were present and the number of spacers was diverse among the three genomes (TDC60: 89; W83: 44; and ATCC 33277: 137).

    Techniques: CRISPR

    Attachment of T. forsythia 43037 (Tf) and BFM571 to KB cells in the absence and presence of P. gingivalis OMVs (Pgv). Attachment levels (means ± standard errors) were expressed as the percentage of attached bacteria relative to the total number

    Journal:

    Article Title: Porphyromonas gingivalis Vesicles Enhance Attachment, and the Leucine-Rich Repeat BspA Protein Is Required for Invasion of Epithelial Cells by "Tannerella forsythia"

    doi: 10.1128/IAI.00062-06

    Figure Lengend Snippet: Attachment of T. forsythia 43037 (Tf) and BFM571 to KB cells in the absence and presence of P. gingivalis OMVs (Pgv). Attachment levels (means ± standard errors) were expressed as the percentage of attached bacteria relative to the total number

    Article Snippet: There were no significant differences between P. gingivalis - or P. gingivalis OMV-induced aggregations of T. forsythia ATCC 43037 and those of the BspA-defective mutant BFM571.

    Techniques:

    Coaggregation of T. forsythia in the presence of P. gingivalis and outer membrane vesicles purified from P. gingivalis . Data are representative of three independent experiments. Data points represent means ± standard errors of triplicate readings.

    Journal:

    Article Title: Porphyromonas gingivalis Vesicles Enhance Attachment, and the Leucine-Rich Repeat BspA Protein Is Required for Invasion of Epithelial Cells by "Tannerella forsythia"

    doi: 10.1128/IAI.00062-06

    Figure Lengend Snippet: Coaggregation of T. forsythia in the presence of P. gingivalis and outer membrane vesicles purified from P. gingivalis . Data are representative of three independent experiments. Data points represent means ± standard errors of triplicate readings.

    Article Snippet: There were no significant differences between P. gingivalis - or P. gingivalis OMV-induced aggregations of T. forsythia ATCC 43037 and those of the BspA-defective mutant BFM571.

    Techniques: Purification

    Invasion by T. forsythia (Tf) of KB cells and dependence of invasion on the presence of P. gingivalis vesicles (Pgv). The invasion level was expressed as the percentage of bacteria retrieved following antibiotic killing of external bacteria and KB cell

    Journal:

    Article Title: Porphyromonas gingivalis Vesicles Enhance Attachment, and the Leucine-Rich Repeat BspA Protein Is Required for Invasion of Epithelial Cells by "Tannerella forsythia"

    doi: 10.1128/IAI.00062-06

    Figure Lengend Snippet: Invasion by T. forsythia (Tf) of KB cells and dependence of invasion on the presence of P. gingivalis vesicles (Pgv). The invasion level was expressed as the percentage of bacteria retrieved following antibiotic killing of external bacteria and KB cell

    Article Snippet: There were no significant differences between P. gingivalis - or P. gingivalis OMV-induced aggregations of T. forsythia ATCC 43037 and those of the BspA-defective mutant BFM571.

    Techniques:

    Effect of serum on viability of A. oris (Ao), V. parvula (Vp), F. nucleatum ATCC 10953 (Fn) and P. gingivalis 381 (Pg). Planktonic growth of microorganisms was evaluated in their appropriate nutrient-rich medium supplemented with 10% native human serum

    Journal: Anaerobe

    Article Title: Strain-Specific Colonization Patterns and Serum Modulation of Multi-Species Oral Biofilm Development

    doi: 10.1016/j.anaerobe.2012.06.003

    Figure Lengend Snippet: Effect of serum on viability of A. oris (Ao), V. parvula (Vp), F. nucleatum ATCC 10953 (Fn) and P. gingivalis 381 (Pg). Planktonic growth of microorganisms was evaluated in their appropriate nutrient-rich medium supplemented with 10% native human serum

    Article Snippet: For these experiments, we inoculated flow cells simultaneously with A. oris , V. parvula , F. nucleatum ATCC 10953 and P. gingivalis 381. shows the effect of serum on multi-species biofilms and also on mono-species biofilms of F. nucleatum ATCC 10953 and P. gingivalis 381.

    Techniques:

    Attachment of T. forsythia 43037 (Tf) and BFM571 to KB cells in the absence and presence of P. gingivalis OMVs (Pgv). Attachment levels (means ± standard errors) were expressed as the percentage of attached bacteria relative to the total number

    Journal:

    Article Title: Porphyromonas gingivalis Vesicles Enhance Attachment, and the Leucine-Rich Repeat BspA Protein Is Required for Invasion of Epithelial Cells by "Tannerella forsythia"

    doi: 10.1128/IAI.00062-06

    Figure Lengend Snippet: Attachment of T. forsythia 43037 (Tf) and BFM571 to KB cells in the absence and presence of P. gingivalis OMVs (Pgv). Attachment levels (means ± standard errors) were expressed as the percentage of attached bacteria relative to the total number

    Article Snippet: In the presence of P. gingivalis OMVs, attachment of T. forsythia ATCC 43037 was increased fourfold.

    Techniques:

    Coaggregation of T. forsythia in the presence of P. gingivalis and outer membrane vesicles purified from P. gingivalis . Data are representative of three independent experiments. Data points represent means ± standard errors of triplicate readings.

    Journal:

    Article Title: Porphyromonas gingivalis Vesicles Enhance Attachment, and the Leucine-Rich Repeat BspA Protein Is Required for Invasion of Epithelial Cells by "Tannerella forsythia"

    doi: 10.1128/IAI.00062-06

    Figure Lengend Snippet: Coaggregation of T. forsythia in the presence of P. gingivalis and outer membrane vesicles purified from P. gingivalis . Data are representative of three independent experiments. Data points represent means ± standard errors of triplicate readings.

    Article Snippet: In the presence of P. gingivalis OMVs, attachment of T. forsythia ATCC 43037 was increased fourfold.

    Techniques: Purification