Yuichiro Noiri

Participation of bacterial biofilms in refractory and chronic periapical periodontitis.

The aim of this study was to examine morphologically the participation of extraradicular biofilm in refractory periapical periodontitis. Six teeth and five extruded root filling gutta-percha points associated with refractory periapical periodontitis were investigated by scanning electron microscope. In nine of 11 samples examined, bacterial biofilms were seen at the extraradicular area. The gutta-percha surface was covered with glycocalyx-like structures, and filaments, long rods, and spirochete-shaped bacteria were predominant in the extraradicular sites. Planktonic cells, which were filaments and spirochete-shaped bacteria, emigrated from the glycocalyx structures in some spots. In the extracted teeth, biofilm consisting of both bacteria and glycocalyx-like structures were observed on the periapical root surfaces. Next to the residual periodontal ligament, a few filaments, rods, and fusiforms were attached on the healthy cementum surface. The present findings suggested that bacterial biofilms formed in the extraradicular areas were related to refractory periapical periodontitis.

Inhibitory effects of resin composite containing bactericide-immobilized filler on plaque accumulation

OBJECTIVE Previously, we have reported that incorporation of the antibacterial monomer 12-methacryloyloxydodecylpyridinium bromide (MDPB) was effective in immobilizing bactericide in the resin matrix, and an antibacterial composite without release of the agent could be achieved. In this study, an attempt was made to increase the density of bactericide immobilized in composite, and the inhibitory effects of this modified antibacterial composite on plaque accumulation were determined, focusing on the reliability of the effects and the mechanisms to affect the plaque formation. METHODS An experimental composite containing immobilized bactericide at 2.83% was prepared by the incorporation of MDPB into a prepolymerized resin filler of control composite, and elution of antibacterial components and inhibition of in vitro plaque accumulation by Streptococcus mutans were determined. The inhibitory effects of the experimental composite on the attachment, glucan synthesis and growth of S. mutans on the surface were also examined in addition to the comparison of surface roughness and hydrophobicity with controls. The results were analyzed using the Students t-test. RESULTS The experimental composite had reproducible inhibitory effects against plaque accumulation compared with control (p<0.05), although it showed no elution of unpolymerized MDPB. The plaque-inhibitory effect of the experimental composite was found to depend upon the ability to inhibit the attachment, glucan synthesis, and growth of bacteria on its surface as no significant differences in the surface characteristics were obtained between control and experimental composites (p>0.05). SIGNIFICANCE It was indicated that the experimental composite containing bactericide-immobilized filler has the possibility to be used clinically with an effective anti-plaque property.

The Localization of Periodontal-disease-associated Bacteria in Human Periodontal Pockets

Some Gram-negative anaerobes are associated with the incidence and progression of periodontal disease. In periodontal pockets, however, the localization of those bacteria is unknown. We investigated the localization of 5 bacterial species in human periodontal pockets. Fifteen teeth with a part of periodontal pockets from 10 adult periodontitis patients were obtained, and the localization of bacteria was examined immunohistochemically. Positive reactions with anti-Prevotella nigrescens antibody were located at the epithelium-associated plaque area in the middle pocket zones. In the middle and deep pocket zones, Fusobacterium nucleatum and Treponema denticola were especially localized in the unattached plaque area, but Eikenella corrodens was observed in the tooth-attached plaque area. Actinobacillus actinomycetemcomitans, detected in 2 of 15 samples examined, was found in the unattached plaque area, in the middle pocket zone. The present findings indicated that the 5 bacterial species examined localized at distinct regions in human periodontal pockets.

Identification and Localization of Extraradicular Biofilm-Forming Bacteria Associated with Refractory Endodontic Pathogens

ABSTRACT Bacterial biofilms have been found to develop on root surfaces outside the apical foramen and be associated with refractory periapical periodontitis. However, it is unknown which bacterial species form extraradicular biofilms. The present study aimed to investigate the identity and localization of bacteria in human extraradicular biofilms. Twenty extraradicular biofilms, used to identify bacteria using a PCR-based 16S rRNA gene assay, and seven root-tips, used to observe immunohistochemical localization of three selected bacterial species, were taken from 27 patients with refractory periapical periodontitis. Bacterial DNA was detected from 14 of the 20 samples, and 113 bacterial species were isolated. Fusobacterium nucleatum (14 of 14), Porphyromonas gingivalis (12 of 14), and Tannellera forsythensis (8 of 14) were frequently detected. Unidentified and uncultured bacterial DNA was also detected in 11 of the 14 samples in which DNA was detected. In the biofilms, P. gingivalis was immunohistochemically detected in all parts of the extraradicular biofilms. Positive reactions to anti-F. nucleatum and anti-T. forsythensis sera were found at specific portions of the biofilm. These findings suggested that P. gingivalis, T. forsythensis, and F. nucleatum were associated with extraradicular biofilm formation and refractory periapical periodontitis.

Influence of Resin Monomers on Growth of Oral Streptococci

Ethyleneglycol dimethacrylate monomers have been previously reported to stimulate the growth of certain caries-associated bacteria on the basis of turbidity measurements. To elucidate the detail of this effect, we examined the influence of resin monomers on the growth of Streptococcus sobrinus and Streptococcus sanguis by determination of bacterial numbers (colony-forming units), morphological observation, and chemical analysis. Although the absorbance values in the stationary phase of bacterial suspension were increased in the presence of ethyleneglycol monomers, no significant differences were observed for bacterial numbers throughout the incubation period. Scanning electron microscopy observation revealed the formation of sparse vesicular material surrounding bacterial cells when incubated with ethyleneglycol monomers, and these products were proved to be resin polymers. The results demonstrate that the apparent biomass increase during incubation with ethyleneglycol monomers is due not to promotion of bacterial multiplication, but to the polymerization of resin monomers to form vesicular structures attached to cells.

A Porphyromonas gingivalis Mutant Defective in a Putative Glycosyltransferase Exhibits Defective Biosynthesis of the Polysaccharide Portions of Lipopolysaccharide, Decreased Gingipain Activities, Strong Autoaggregation, and Increased Biofilm Formation

ABSTRACT The Gram-negative anaerobic bacterium Porphyromonas gingivalis is a major pathogen in periodontal disease, one of the biofilm-caused infectious diseases. The bacterium possesses potential virulence factors, including fimbriae, proteinases, hemagglutinin, lipopolysaccharide (LPS), and outer membrane vesicles, and some of these factors are associated with biofilm formation; however, the precise mechanism of biofilm formation is still unknown. Colonial pigmentation of the bacterium on blood agar plates is related to its virulence. In this study, we isolated a nonpigmented mutant that had an insertion mutation within the new gene PGN_1251 (gtfB) by screening a transposon insertion library. The gene shares homology with genes encoding glycosyltransferase 1 of several bacteria. The gtfB mutant was defective in biosynthesis of both LPSs containing O side chain polysaccharide (O-LPS) and anionic polysaccharide (A-LPS). The defect in the gene resulted in a complete loss of surface-associated gingipain proteinases, strong autoaggregation, and a marked increase in biofilm formation, suggesting that polysaccharide portions of LPSs influence attachment of gingipain proteinases to the cell surface, autoaggregation, and biofilm formation of P. gingivalis.

Effects of Er:YAG Laser Irradiation on Biofilm-forming Bacteria Associated with Endodontic Pathogens In Vitro

With the development of dental laser delivery systems that can enter into the root canals, it is possible to use Er:YAG lasers to remove the residual biofilm associated with infected root canals. We examined their effects against biofilms made of Actinomyces naeslundii, Enterococcus faecalis, Lactobacillus casei, Propionibacterium acnes, Fusobacterium nucleatum, Porphyromonas gingivalis, or Prevotella nigrescens in vitro. After Er:YAG laser irradiation with energy densities ranging between 0.38-0.98 J/cm(2), the biofilm samples on hydroxyapatite disks were quantitatively and morphologically evaluated. The Er:YAG laser was effective against biofilms of 6 of the bacterial species examined, with the exception of those formed by L. casei. After irradiation, the numbers of viable cells in the biofilms were significantly decreased, whereas atrophic changes in bacterial cells and reductions in biofilm cell density were seen morphologically. Er: YAG lasers might be suitable for clinical application as a suppressive and removal device of biofilms in endodontic treatments.

Effects of the tea catechin epigallocatechin gallate on Porphyromonas gingivalis biofilms.

The aim of this study was to investigate the effects of tea catechin epigallocatechin gallate (EGCg) on established biofilms and biofilm formation by Porphyromonas gingivalis, a major pathogen of periodontal disease.

Essential Role for the gtfA Gene Encoding a Putative Glycosyltransferase in the Adherence of Porphyromonas gingivalis

ABSTRACT Porphyromonas gingivalis, an oral bacterium, might play a role in the pathogenesis or progression of adult periodontitis. In this study, we isolated from P. gingivalis a putative glycosyltransferase gene, designated gtfA, which had a consensus domain for glycosyltransferase in its N terminus. GtfA consisted of 248 amino acids and its predicted molecular mass was 28 kDa; however, as the molecular mass of endogenous GtfA protein was around 40 kDa, this suggested that GtfA had undergone some posttranslational modifications. To reveal the role of the gtfA gene in P. gingivalis, we established gtfA-deficient strains by allelic replacement. Morphologically, gtfA-deficient P. gingivalis lacked mature fimbriae. gtfA-deficient P. gingivalis also showed a very low ability for autoaggregation, and its ability to attach to epithelial cells was severely impaired. Thus, the results indicate that the gtfA gene is required for P. gingivalis autoaggregation as well as attachment to epithelial cells. These results suggest that GtfA might have an important role in the pathogenicity of P. gingivalis by regulating adhesion.

Antibiofilm Effects of Azithromycin and Erythromycin on Porphyromonas gingivalis

ABSTRACT Antibiotic resistance of biofilm-grown bacteria contributes to chronic infections, such as marginal and periapical periodontitis, which are strongly associated with Porphyromonas gingivalis. Concurrent azithromycin (AZM) administration and mechanical debridement improve the clinical parameters of periodontal tissue in situ. We examined the in vitro efficacy of AZM against P. gingivalis biofilms. The susceptibilities of adherent P. gingivalis strains 381, HW24D1, 6/26, and W83 to AZM, erythromycin (ERY), ampicillin (AMP), ofloxacin (OFX), and gentamicin (GEN) were investigated using a static model. The optical densities of adherent P. gingivalis cells were significantly decreased by using AZM and ERY at sub-MIC levels compared with those of the controls in all the strains tested, except for the effect of ERY on strain W83. AMP and OFX inhibited P. gingivalis adherent cells at levels over their MICs, and GEN showed no inhibition in the static model. The effects of AZM and ERY against biofilm cells were investigated using a flow cell model. The ATP levels of P. gingivalis biofilms were significantly decreased by AZM at concentrations below the sub-MICs; however, ERY was not effective for inhibition of P. gingivalis biofilm cells at their sub-MICs. Furthermore, decreased density of P. gingivalis biofilms was observed three-dimensionally with sub-MIC AZM, using confocal laser scanning microscopy. These findings suggest that AZM is effective against P. gingivalis biofilms at sub-MIC levels and could have future clinical application for oral biofilm infections, such as chronic marginal and periapical periodontitis.