Porphyromonas gingivalis minor fimbriae are required for cell-cell interactions

Abstract

Two distinctive types of fimbriae have been identified in Porphyromonas gingivalis. In this report, we demonstrate that minor fimbriae are involved in P. gingivalis autoaggregation and colonization. A mutant with a deficiency in minor fimbriae can bind to a saliva-coated surface but does not form microcolonies as the wild-type strain does.

Figures

FIG. 1.

Comparison of the growth curves of P. gingivalis strains. Cells were grown in TSB medium. Shown in the curves are means of four samples, with error bars representing the standard error of the mean. One-milliliter aliquots were taken, and the OD600 was measured over a period of 72 h.

FIG. 2.

Aggregation of P. gingivalis strains in TSB. (A) Immunodetection of fimbrial production with rabbit polyclonal anti-FimA and anti-Mfa1 antibodies. Lane 1, wild-type strain 33277 (fimA+ mfa1+); lane 2, strain DFAET (ΔfimA Δmfa1); lane 3, strain SMFC (fimA+ mfa1+); lane 4, strain FAT (ΔfimA mfa1+); lane 5, strain MFAE (fimA+ Δmfa1). (B) P. gingivalis cells were grown in TSB for 24 h. Aggregated cells collected at the bottoms of the test tubes. (C) The number of P. gingivalis cells was estimated in a spectrophotometer as described by Soukos et al. (17). OD600 was determined. Readings of supernatant were taken from 9 ml of top culture, and readings of aggregation were from 1 ml of bottom culture. An OD600 of 0.1 equals approximately 108 P. gingivalis cells per ml.

FIG. 3.

Quantitation of biofilm formation by P. gingivalis on saliva-coated polystyrene wells. P. gingivalis biofilms were allowed to form in saliva-coated six-well polystyrene microtiter dishes. The cells bound to the well surfaces were suspended in 1 ml PBS for 4, 16, 24, 40, 48, 64, and 72 h of incubation. The ability of P. gingivalis strains to attach and form microcolonies on the surface was scored by measuring cell density. Biofilm formation by wild-type strain 33277, strain DFAET (ΔfimA Δmfa1), strain FAT (ΔfimA), and strain MFAE (Δmfa1) was measured by determining OD600. Each data point represents the mean ± the standard deviation from at least three independent experiments.

FIG. 4.

Biofilm formation by P. gingivalis on saliva-coated MatTek glass bottom culture dishes. P. gingivalis cells were stained with FITC and incubated in saliva-coated glass bottom culture dishes for 1 (A, B, C, and D) or 24 (E, F, G, and H) h. P. gingivalis biofilms were visualized with a confocal laser scanning microscope.