Fig 1.
Effects of MOI on adhesion and invasion of Ca9-22 cells by P. gulae D049.
Antibiotic protection invasion assays with P. gulae D049. Ca9-22 cells were infected with P. gulae at an MOI of 1, 10, 100, or 1000 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 <0.01 (Student’s t test), as compared with infected cells (MOI 1, 10, and/or 1000). A; Adhesion efficiency, B; Invasion efficiency.
Fig 2.
Time course for adhesion and invasion of Ca9-22 cells by P. gulae D049.
Ca9-22 cells were infected with P. gulae at an MOI of 100 for the indicated times. 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. A; Adhesion efficiency, B; Invasion efficiency.
Fig 3.
Adhesion/invasion of Ca9-22 cells by P. gulae strains with distinct types of fimbriae (types A, B and C). Antibiotic protection invasion assays of P. gulae strains. Ca9-22 cells were infected with the 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. A; Adhesion efficiency, B; Invasion efficiency. *P <0.01 (Student’s t test), as compared with infected cells (51700, D066 and/or D044 strains).
Fig 4.
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 <0.05, and **P <0.01 (Student’s t test) as compared with P. gingivalis 33277 and/or OMZ314. A; Adhesion efficiency, B; Invasion efficiency.
Table 1.
Effects of metabolic inhibitors on P. gulae D049 adhesion and invasion of Ca9-22 cells.
Table 2.
Effects of human serum on P. gulae adhesion and invasion of Ca9-22 cells.
Table 3.
Effects of human plasma on P. gulae adhesion and invasion of Ca9-22 cells.
Fig 5.
Inhibition of P. gulae D049 adhesion and invasion by anti-integrin α5β1 antibody.
A: Expressions of integrins in Ca9-22 cells. Cells were lysed and immunoblotted with anti-integrin α5, αV, β1, β3, or β-actin antibodies. SAS cells were used as a positive control. B: Ca9-22 cells were infected with P. gulae D049 at an MOI of 100 for 90 min with/without anti-integrin antibodies. The numbers of adherent 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 <0.01 (Student’s t test) as compared with 0 (no anti-integrin antibodies or IgG). IgG, immunoglobulin G. C: Ca9-22 cells were infected with P. gulae D049 at an MOI of 100 for 90 min with/without anti-integrin antibodies. The numbers of 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. *and **, P < 0.05 and P <0.01 (Student’s t test) as compared with 0 (no anti-integrin antibodies or IgG). IgG, immunoglobulin G.
Fig 6.
Scanning electron microscopy of Ca9-22 cells infected with P. gulae D049.
SEM examinations of Ca9-22 cells infected with/without P. gulae D049 at an MOI of 100 for 90 min. Bar markers represent 1 μm. A, Uninfected Ca9-22 cells (lower magnification). B: Uninfected Ca9-22 cells (higher magnification). C: Lower magnification showing engulfment of P. gulae by cellular protrusions during localized adhesion step of infection (arrows). D: Higher magnification showing cellular protrusions engulfing P. gulae (arrows).
Fig 7.
Transmission electron microscopy of Ca9-22 cells infected with P. gulae D049.
TEM examinations of Ca9-22 infected with/without P. gulae D049 at an MOI of 100 for 90 min. Bar markers represent 5 μm (A, C and E) or 1 μm (B, D and F). A: Uninfected Ca9-22 cells (lower magnification). B: Uninfected Ca9-22 cells (higher magnification). C: Lower magnification showing adherent P. gulae and subsequent engulfment by cellular protrusions (arrows). D: Higher magnification showing adherent P. gulae and subsequent engulfment by cellular protrusions (arrows). E: Lower magnification showing intracellular P. gulae in vacuole (arrows). F: Higher magnification showing intracellular P. gulae in vacuole (arrows).