Glycan-foraging systems reveal the adaptation of Capnocytophaga canimorsus to the dog mouth.

UNLABELLED: Capnocytophaga canimorsus is known to form two kinds of cells on blood agar plates (coccoid and bacillary), evoking phase variation. When grown in coculture with animal cells these bacteria appeared only as bacilli, but in the presence of vancomycin they were round, indicating that coccoid shapes likely result from weakening of the peptidoglycan layer. Polysaccharide utilization locus 5 (PUL5) and sialidase mutant bacteria, unable to retrieve glycans from glycoproteins, grew less than wild-type bacteria and also appeared polymorphic unless GlcNAc was added, suggesting that C. canimorsus is unable to synthesize GlcNAc, an essential component of peptidoglycan. Accordingly, a genome analysis was conducted and revealed that C. canimorsus strain 5 lacks the GlmM and GlmU enzymes, which convert glucosamine into GlcNAc. Expression of the Escherichia coli GlmM together with the acetyltransferase domain of GlmU allowed PUL5 mutant bacteria to grow normally, indicating that C. canimorsus is a natural auxotroph that relies on GlcNAc harvested from the host N-glycoproteins for peptidoglycan synthesis. Mucin, a heavily O-glycosylated protein abundant in saliva, also rescued growth and the shape of PUL5 mutant bacteria. Utilization of mucin was found to depend on Muc, a Sus-like system encoded by PUL9. Contrary to all known PUL-encoded systems, Muc cleaves peptide bonds of mucin rather than glycosidic linkages. Thus, C. canimorsus has adapted to build its peptidoglycan from the glycan-rich dog's mouth glycoproteins. IMPORTANCE: Capnocytophaga canimorsus is a bacterium that lives as a commensal in the dog mouth and causes severe infections in humans. In vitro, it forms two kinds of cells (coccoid and bacillary), evoking phase variation. Here, we show that cell rounding likely results from weakening of the peptidoglycan layer due to a shortage of N-acetylglucosamine (GlcNAc). C. canimorsus cannot synthesize GlcNAc because of the lack of key enzymes. In its niche, the dog mouth, C. canimorsus retrieves GlcNAc by foraging glycans from salivary mucin and N-linked glycoproteins through two different apparatuses, Muc and Gpd, both of which are related to the Bacteroides starch utilization system. The Muc system is peculiar in the sense that the enzyme of the complex is a protease and not a glycosylhydrolase, as it cleaves peptide bonds in order to capture glycan chains. This study provides a molecular genetic demonstration for the complex adaptation of C. canimorsus to its ecological niche, the oral cavity of dogs.

Antibiotic content of selective culture media for isolation of Capnocytophaga species from oral polymicrobial samples.

UNLABELLED: In oral microbiome, because of the abundance of commensal competitive flora, selective media with antibiotics are necessary for the recovery of fastidious Capnocytophaga species. The performances of six culture media (blood agar, chocolate blood agar, VCAT medium, CAPE medium, bacitracin chocolate blood agar and VK medium) were compared with literature data concerning five other media (FAA, LB, TSBV, CapR and TBBP media). To understand variable growth on selective media, the MICs of each antimicrobial agent contained in this different media (colistin, kanamycin, trimethoprim, trimethoprim-sulfamethoxazole, vancomycin, aztreonam and bacitracin) were determined for all Capnocytophaga species. Overall, VCAT medium (Columbia, 10% cooked horse blood, polyvitaminic supplement, 3·75 mg l(-1) of colistin, 1·5 mg l(-1) of trimethoprim, 1 mg l(-1) of vancomycin and 0·5 mg l(-1) of amphotericin B, Oxoid, France) was the more efficient selective medium, with regard to the detection of Capnocytophaga species from oral samples (P < 0·001) and the elimination of commensal clinical species (P < 0·001). The demonstrated superiority of VCAT medium, related to its antibiotic content, made its use indispensable for the optimal isolation of Capnocytophaga species from polymicrobial samples. SIGNIFICANCE AND IMPACT OF THE STUDY: Isolation of Capnocytophaga species is important for the proper diagnosis and treatment of the systemic infections they cause and for epidemiological studies of periodontal flora. We showed that in pure culture, a simple blood agar allowed the growth of all Capnocytophaga species. Nonetheless, in oral samples, because of the abundance of commensal competitive flora, selective media with antibiotics are necessary for the recovery of Capnocytophaga species. The demonstrated superiority of VCAT medium made its use essential for the optimal detection of this bacterial genus. This work showed that extreme caution should be exercised when reporting the isolation of Capnocytophaga species from oral polymicrobial samples, because the culture medium is a determining factor.

Mechanical non-surgical treatment of peri-implantitis: a single-blinded randomized longitudinal clinical study. II. Microbiological results.

BACKGROUND: Peri-implantitis is common in patients with dental implants. We performed a single-blinded longitudinal randomized study to assess the effects of mechanical debridement on the peri-implant microbiota in peri-implantitis lesions. MATERIALS AND METHODS: An expanded checkerboard DNA-DNA hybridization assay encompassing 79 different microorganisms was used to study bacterial counts before and during 6 months following mechanical treatment of peri-implantitis in 17 cases treated with curettes and 14 cases treated with an ultrasonic device. Statistics included non-parametric tests and GLM multivariate analysis with p<0001 indicating significance and 80% power. RESULTS: At selected implant test sites, the most prevalent bacteria were: Fusobacterium nucleatum sp., Staphylococci sp., Aggregatibacter actinomycetemcomitans, Helicobacter pylori, and Tannerella forsythia. 30 min. after treatment with curettes, A. actinomycetemcomitans (serotype a), Lactobacillus acidophilus, Streptococcus anginosus, and Veillonella parvula were found at lower counts (p<0.001). No such differences were found for implants treated with the ultrasonic device. Inconsistent changes occurred following the first week. No microbiological differences between baseline and 6-month samples were found for any species or between treatment study methods in peri-implantitis. CONCLUSIONS: Both methods failed to eliminate or reduce bacterial counts in peri-implantitis. No group differences were found in the ability to reduce the microbiota in peri-implantitis.

Effects of selected immunouppressive drugs on prostaglandin release, protein synthesis and cell proliferation in human gingival fibroblasts and on the growth of plaque bacteria.

BACKGROUND: Immunosuppressants play an essential role in transplantation therapy. In view of the side effects, e.g. gingival overgrowth, the present in vitro study was performed in order to investigate the effect of selected immunosuppressants on metabolic activities of gingival fibroblasts. Furthermore, the effect on the growth of six oral microorganisms was investigated. METHODS: Human gingival fibroblasts were incubated in the presence of azathioprine (Aza), cyclosporin A (CsA), tacrolimus (Tac) or mycophenolatmofetil (Myc). PGE subset 2 release was determined by means of a specific competitive enzyme immunoassay, using monoclonal antibodies specific for PGE subset 2 (clone E2R1). The protein content was measured spectrophotometrically. A redox indicator system was employed to assess the proliferation activity. In an additional trial the growth of six strains of oral bacteria (A. viscosus T14V, S. oralis H1, S. mutans 10449, C. gingivalis DR2001, A. actinomycetemcomitans Y4, and M. micros 33270) in the presence of the immunosuppressants was measured. RESULTS: In comparison with the controls, the PGE subset 2 release was increased by 39.3% following incubation with Aza, and by 77.0% with CsA. The protein concentrations (1 g immunosuppressant / ml medium) were reduced by 26.0% for Aza and 17.0% for Myc. Furthermore, a drug-dependent inhibition in the cell proliferation rate was noted after an incubation period of 6 hours (Aza 70.7%, CsA 78.2%, Myc 69.8%, Tac 64.0%). The most pronounced growth-inhibiting effects were observed for CsA at values ranging from 21.0% (S. mutans 10449) to 48.6% (A. viscosus T14V) growth inhibition. CONCLUSIONS: The present study with common immunsuppresants demonstrated both a medication- and dose-dependent alteration in the metabolic activity of gingival fibroblasts. Furthermore, growth-inhibitory effects on the selected bacterial strains could be observed.

Microbiological, immunological and genetic factors in family members with periodontitis as a manifestation of systemic disease, associated with hematological disorders.

The microflora, immunological profiles of host defence functions, and human leukocyte antigen (HLA) findings are reported for a mother, son and daughter who were diagnosed as having 'periodontitis as a manifestation of systemic diseases, associated with hematological disorders'. Examinations were made of the bacterial flora from the periodontal pocket, neutrophil chemotaxis, neutrophil phagocytosis, and the genotypes (DQB1) and serotypes (DR locus) of HLA class II antigens. Phenotypic analyses of the peripheral lymphocytes were also conducted. The subgingival microflora from the mother was dominated by Gram-negative rods, especially Porphyromonas endodontalis, Prevotella intermedia/Prevotella nigrescens and Fusobacterium nucleatum. Subgingival microflora samples from the son and daughter were dominated by Gram-positive cocci and Gram-positive rods. Through the use of polymerase chain reaction, Campylobacter rectus and Capnocytophaga gingivalis were detected in all subjects, whereas Porphyromonas gingivalis, P. intermedia, and Treponema denticola were not detected in any subjects. All three subjects showed a remarkable level of depressed neutrophil chemotaxis to N-formyl-methionyl-leucyl-phenylalanine, although their phagocyte function levels were normal, in comparison to healthy control subjects. Each subject had the same genotype, HLA-DQB1*0601, while the mother had HLA-DR2 and HLA-DR8, and the son and daughter had HLA-DR2 only. In summary, the members of this family showed a similar predisposition to periodontitis with regard to certain host defence functions. It is suggested that the depressed neutrophil chemotaxis that was identified here could be a significant risk factor for periodontitis in this family.

Long-term follow-up of osseointegrated titanium implants using clinical, radiographic and microbiological parameters.

The aim of this study was to longitudinally follow up osseointegrated titanium implants in partially dentate patients by clinical, radiographic and microbiological parameters in order to evaluate possible changes in the peri-implant health over time. Fifteen individuals treated with titanium implants, ad modum Brånemark, and followed for ten years were included in the study. Before implant placement ten years previously, the individuals had been treated for advanced periodontal disease and thereafter been included in a maintenance care program. The survival rate of the implants after ten years was 94.7%. The bone loss was 1.7 mm when using the abutment-fixture junction as a reference point. Of the individuals, 50% were positive for plaque at the implants. Bleeding on sulcus probing was present at 61% of the implant surfaces. Ten years previously, the individuals had been carriers of putative periodontal pathogens, such as Porphyromonas gingivalis, Prevotella intermedia, Actinobacillus actinomycetemcomitans, Capnocytophaga spp. and Campylobacter rectus, and were also carriers of these species at the current examination. The results of the present study suggest that the presence of these putative periodontal pathogens at implants may not be associated with an impaired implant treatment. These species are most likely part of the normal resident microbiota of most individuals and may therefore be found at random at both stable and progressing peri-implant sites.

Effects of instrumentation, irrigation and dressing with calcium hydroxide on infection in pulpless teeth with periapical bone lesions.

AIM: The aim of this study was to evaluate the fate of microorganisms in root canals of teeth with infected pulps and periapical bone lesions with and without the use of calcium hydroxide medication. METHODOLOGY: Endodontic samples were cultured and microorganisms were counted and identified in 43 teeth before (sample 1) and after (sample 2) treatment during the first visit and before (sample 3) and after (sample 4) treatment during the second visit. In the first visit teeth were instrumented and half of the teeth were filled with a thick slurry of calcium hydroxide in sterile saline. The other teeth were obturated with gutta-percha and AH-2 6 sealer. After 4 weeks the teeth with calcium-hydroxide were accessed again and after microbiological sampling they were obturated with gutta-percha and AH-26 sealer. RESULTS: The mean total colony forming unit (CFU) counts of positive samples dropped significantly as a result of canal preparation during the first visit from 1.0 x 10(6) to 1.8 x 10(3) (between samples 1 and 2) but increased to 9.3 x 10(3) in the period between the two visits (sample 2 and 3). There was no difference in mean total CFU counts of positive samples between the end of the first (sample 2) and the end of the second visit (sample 4). The most frequently isolated species were Prevotella intermedia, Capnocytophaga spp.. Actinomyces odontolyticus. Propionibacterium acnes and Peptostreptococcus micros. CONCLUSIONS: Although a calcium hydroxide paste was placed in the prepared canals, the number of positive canals had increased in the period between visits. However, the number of microorganisms had only increased to 0.93% of the original number of CFU (sample 1). It is concluded that a calcium hydroxide and sterile saline slurry limits but does not totally prevent regrowth of endodontic bacteria.

Relationship of periodontopathic bacteria with early-onset periodontitis in Down's syndrome.

BACKGROUND: Down's syndrome (DS) patients often develop severe early-onset marginal periodontitis in early adulthood; however, there is little information available on the microbiology of DS periodontitis. METHODS: Subgingival plaque specimens were taken from 67 DS young adults and 41 age-matched systemically healthy individuals with mental disabilities (MD). The prevalence of 10 possible periodontopathic bacterial species, Actinobacillus actinomycetemcomitans, Porphyromonas gingivalis, Bacteroides forsythus, Treponema denticola, Prevotella intermedia, Prevotella nigrescens, Capnocytophaga ochracea, Capnocytophaga sputigena, Campylobacter rectus, and Eikenella corrodens, were investigated in their subgingival plaque samples using a polymerase chain reaction method. The detection of P. gingivalis fimA genotypes was also performed in P. gingivalis-positive samples. RESULTS: Although DS subjects generally develop an earlier and more extensive periodontal breakdown than those with MD, no significant differences were observed in the bacterial profiles. The profiles of subjects with periodontitis were significant in DS, but not in MD. The prevalence of P. gingivalis, B. forsythus, and P. intermedia were significant in the DS periodontitis group, compared to DS gingivitis group. Moreover, the occurrence of P. gingivalis with the type II fimA gene was significantly related to periodontitis in both DS and MD, with odds ratios of 6.32 and 12.03, respectively. CONCLUSIONS: These results suggest that early-onset periodontitis in DS is mainly due to the more susceptible host for the causative microbial agents including P. gingivalis with type II fimA.

Effect of three mouthrinses, containing amine/stannous fluoride, herbal extracts or Emser salt on the growth of oral bacteria--an in vitro study.

BACKGROUND/AIMS: Clinical studies have shown the efficacy of mouthrinses in reducing plaque accumulation and inflammation of oral tissues. The aim of this in vitro study was to compare the effect of three mouthrinses: Meridol, an organic amine/ stannous fluoride solution; Parodontax, containing herbal ingredients; and an 0.8 % Emser salt solution, on the growth of oral bacteria and dental plaque. METHODS: Growth of Actinomyces viscosus T14V, Capnocytophaga ochracea 25, C. sputigena 4, Actinobacillus actinomycetemcomitans (A.a.) Y4, and pooled supragingival plaque in the presence of the various mouthrinses, applied to paper discs, was tested in an agar diffusion test. In a second series of tests, the 4 bacterial strains were exposed to the agents for about 3 min to simulate rinsing, then the agent was removed, and the bacteria were inoculated into fresh nutrient broth. After 48 h bacterial growth was measured in a spectrophotometer and compared with the controls. RESULTS: In the agar diffusion test only Meridol, the organic amine/stannous fluoride-containing solution, could inhibit bacterial growth, except for A. a. Y4. When the bacteria where in contact with the agents for only a few minutes these results were confirmed. Neither Paradontax nor Emser salt inhibited the growth of the bacteria, and A. a. Y4 proved to be resistant to all three agents. Growth of the other three strains was inhibited by Meridol 92-99% (undiluted), 85-96% (1:5) and 83-98% (1:10). CONCLUSIONS: We conclude that only Meridol contains ingredients capable of inhibiting the growth of oral bacteria in vitro. The efficacy of the other two mouthrinses in reducing plaque accumulation in vivo has to be explained by other mechanisms.

Severe, rapidly progressing peri-implantitis.

The replacement of teeth by means of titanium implants is a safe and predictable procedure in most patients. Some studies show that a small number of patients lose the majority of their implants. Unfortunately, it is impossible to identify these patients prior to surgery. It is important to report such cases so that we may, in the future, be able to identify them before surgery. The present case showed a severe and rapid exfoliation of titanium implants. Out of 8 implants inserted in the anterior lower jaw of this patient, only two maintained osseointegration over a 2-year period.

Growth and hydrolytic enzyme production of Capnocytophaga gingivalis on different protein substrates.

Capnocytophaga gingivalis was grown with proteins (albumin, collagen, mucin and hemoglobin) as carbon and energy sources in chemostat culture. The mu max (0.34 h-1) and biomass yield (0.96 g.l-1) were as high with hemoglobin (3 g.l-1) as with glucose (3 g.l-1) (20). Albumin, collagen and mucin also supported an increased mu max, or yield or both, in comparison with basal (tryptone/thiamine) medium. In steady-state, trypsin-like protease specific activity increased 3- to 5-fold in the presence of albumin, collagen and hemoglobin: whereas the greatest increase (21-fold) in alpha-glucoside activity was in the presence of mucin. There were significant, but less substantial changes in other hydrolytic enzymes (aminopeptidase, acid and alkaline phosphatases). The bulk of the detected hydrolytic activity (> 66%) was associated with the cells. The data indicate that C. gingivalis regulates its production of hydrolytic enzymes in response to environmental conditions.

Microbiota of successful osseointegrated dental implants.

BACKGROUND: The long-term survival of dental implants depends, in part, on control of bacterial infection in the peri-implant region. Periodontal pathogens colonized implants symptomatic through infection, whereas the microbiota of successful implants was similar to that of periodontal health. This study examined the impact on the peri-implant microbiota of crown restorations; implant type; length of time of loading; history of implant or periodontal infections; and whether implants replaced single or multiple teeth. It was of particular interest to evaluate implant colonization by species in a newly described red complex of periodontal pathogens, Porphyromonas gingivalis and Bacteroides forsythus. METHODS: This study sampled 43 partially edentulous subjects with successfully osseointegrated titanium root-form dental implants. Eighty-one (81) non-submerged and 20 submerged asymptomatic implants, 83 crowned, and 36 uncrowned teeth were sampled from peri-implant or subgingival sites. The microbiota of samples was evaluated using whole genomic DNA probes in a checkerboard assay to 23 subgingival species. RESULTS: Implants were colonized principally by oral streptococci, capnocytophagae, Veillonella parvula, Peptostreptococcus micros, and Fusobacterium nucleatum. The periodontal species, P. gingivalis, B. forsythus, Prevotella intermedia, Prevotella nigrescens, and Campylobacter rectus were detected in a few subjects. The microbiota around crowned implants and crowned teeth was similar. Streptococcus oralis, P. intermedia, and Selenomonas noxia were elevated in samples from uncrowned teeth compared to crowned teeth and implants. Microbial complexity increased as loading time increased, but colonization by periodontal pathogens, including red complex species, was higher in subjects with previous periodontal disease. No differences were observed in the microbiota of 1- and 2-stage implants, or between implants supporting single or multiple restorations. CONCLUSIONS: While presence of crowns had only a minor impact on the peri-implant microbiota, microbial changes were observed the longer the implants had been in function and in those patients with a history of periodontal or peri-implant infections. A history of periodontitis had a greater impact on the peri-implant microbiota than implant loading time. The major influence on the peri-implant microbiota was, however, the microbiota on remaining teeth. P. gingivalis and B. forsythus, red complex periodontal pathogens, colonized several implants, although all implants were successfully osseointegrated.

Microbiological features and crevicular fluid aspartate aminotransferase enzyme activity in early onset periodontitis patients.

Gingival crevicular fluid (GCF) reflects the immune and inflammatory reactions and the specific host-microbe interactions that lead to periodontal diseases. Aspartate aminotransferase enzyme (AST) is one of the components of GCF that is released as a result of cell death. In this study, periodontal sites (4 sites/patient) with a probing depth of > or =5 mm in early onset periodontitis (EOP) patients were first examined for the AST levels in GCF by the Periogard periodontal tissue monitor. To be eligible for the study, each of the patients had at least 1 AST positive site with clinical inflammatory changes (AST+, CIC+) and 1 AST negative site with no or minimum clinical inflammatory changes (AST-, CIC-). In 15 EOP patients who met the entry criteria, 30 AST+, CIC+ sites (1st group) and 19 AST-, CIC- sites (2nd group) were evaluated for microbiological variables. Certain microbial species, including Porphyromonas gingivalis, Actinobacillus actinomycetemcomitans and Prevotella intermedia were detected more frequently (p<0.001, p<0.001 and p<0.05, respectively) in the 1st group, while gram-positive facultative organisms such as Actinomyces species were found more often (p<0.001) in the 2nd group. Parallel to the AST levels, the 2nd group had a lower number of total bacteria and proportion of obligate anaerobic and capnophilic micro-organisms than the first group (p<0.05 and p<0.05, respectively). Within the scope of this study, AST activity and microbiological data were found in agreement in the examined groups. These findings are encouraging and indicate the need for further studies to evaluate the ability of the AST test to differentiate the microbial flora of progressing sites and those that are inflamed, but not progressing.

The effect of supragingival plaque control on the progression of advanced periodontal disease.

The aim of the present trial was to study the effect of meticulous supragingival plaque control on (i) the subgingival microbiota, and (ii) the rate of progression of attachment loss in subjects with advanced periodontal disease. An intra-individual group of sites exposed to non-surgical periodontal therapy served as controls. 12 patients with advanced periodontal disease were subjected to a baseline examination (BL) including assessments of oral hygiene status, gingival condition (BoP), probing depth, clinical attachment level and subgingival microbiota from pooled samples from each quadrant. The assessments were repeated after 12, 24 and 36 months. Following BL, a split mouth study was initiated. The patients received oral hygiene instruction, supragingival scaling and case presentation. 2 quadrants in each patient were identified as "test" and the remaining 2 as "control" quadrants. Subgingival therapy was performed in all bleeding sites in the control quadrants. Oral hygiene instructions and plaque control exercises were repeated once every 2 weeks during the initial 3 months of the study. Thereafter the plaque control program was repeated once every 3 months for the duration of the 3 years. Sites demonstrating loss of clinical attachment > or =2 mm in the test quadrants were treated subgingivally. The results showed that in both test and control quadrants repeated oral hygiene instructions and supragingival plaque removal procedures resulted in low plaque scores throughout the study. The gingival bleeding scores and the frequency of periodontal pockets > or =4 mm was, however, significantly higher in the test quadrants than in the control quadrants. At the end of the 3 year study, the control quadrants showed significantly more reduced (> or =2 mm) pockets than the test quadrants, 265 versus 96. The number of sites in the test quadrants showing probing attachment loss > or =2 mm was more than 4x greater than in the control quadrants (59 versus 13). The microbiological findings indicate a more pronounced reduction only for P. gingivalis in the control quadrants. None of the other 4 marker bacteria consistently reflected or predicted the clinical parameters. The present study shows that only supragingival plaque control fails to prevent further periodontal tissue destruction in subjects with advanced periodontal disease.

Subgingival distribution of periodontopathic bacteria in adult periodontitis and their susceptibility to minocycline-HCl.

The purpose of this study was to investigate the distribution of several periodontopathic bacteria in adult periodontitis, their in vitro susceptibility to minocycline-HCl, and whether the efficacy of the drug changes with a decrease in bacterial susceptibility. Twenty-one patients (43 to 75 years old) with 62 periodontal lesions from pockets > or =4 mm participated in the study. After subgingival sampling, an ointment containing 2% minocycline-HCl was applied locally to the selected pockets once a week for 4 weeks. The lesions were clinically examined after 1 and 4 weeks of administration. The distribution of the subgingival microorganisms included Capnocytophaga sputigena (37.1%), Prevotella intermedia (22.6%), Porphyromonas gingivalis (22.6%), Fusobacterium nucleatum (20.1%), Actinobacillus actinomycetemcomitans (9.7%), and Eikenella corrodens (4.8%). The distribution was complex, with 76.8% of the sites containing 1 to 3 bacterial spieces. The minimum inhibitory concentration (MIC) of minocycline-HCl for each organism showed that most were inhibited by a minocycline-HCl concentration equal to or less than the MIC for reference strains. However, some clinical strains of Prevotella intermedia seemed to exihibit low susceptibility to minocycline-HCl. There were no significant differences among sites with strains exhibiting low or normal susceptibility to minocycline-HCl. The concentration of the drug applied to deep periodontal pockets inhibited the growth of most of the microorganisms investigated in this study.

C-telopeptide pyridinoline cross-links (ICTP) and periodontal pathogens associated with endosseous oral implants.

Detection of periodontal or peri-implant sites exhibiting progressing disease or those at risk of deterioration has proven difficult. Pyridinoline cross-linked carboxyterminal telopeptide of type I collagen (ICTP), a marker specific for bone degradation found in gingival crevicular fluid (GCF), has been associated with both bone and attachment loss in periodontitis and may be useful for predicting disease activity. The aim of this cross-sectional study was to examine the relationship between ICTP levels and subgingival species around implants and teeth from 20 partially and 2 fully edentulous patients. GCF and plaque samples were collected from the mesiobuccal site of each implant and tooth. Radioimmunoassay techniques were utilized to determine GCF ICTP levels. Plaque samples were analyzed utilizing checkerboard DNA-DNA hybridization. Traditional clinical parameters were assessed. Seventy-one implants and 370 teeth from 22 subjects were examined. ICTP levels and subgingival plaque composition were not significantly different between implants and teeth. Implant sites colonized by Prevotella intermedia, Capnocytophaga gingivalis, Fusobacterium nucleatum ss vincentii, and Streptococcus gordonii exhibited odds ratios of 12.4, 9.3, 8.1, and 6.7, respectively of detecting ICTP. These results suggest a relationship between elevated ICTP levels at implant sites and some species associated with disease progression. Longitudinal studies are necessary to determine whether elevated ICTP levels may predict the development of peri-implant bone loss.

Effects of triclosan and triclosan monophosphate on maximum specific growth rates, biomass and hydrolytic enzyme production of Streptococcus sanguis and Capnocytophaga gingivalis in continuous culture.

Dental plaque species, Streptococcus sanguis and Capnocytophaga gingivalis, were grown in continuous culture with progressively increasing concentrations of triclosan or its phosphorylated derivative, triclosan monophosphate (TMP). For both organisms, the maximum specific growth rates decreased with increasing concentrations of triclosan or TMP until complete inhibition of growth occurred, which for S. sanguis was at 20 mg/L and 50 mg/L, and for C. gingivalis was at 10 mg/L and 5 mg/L for triclosan and TMP respectively. For both species, biomass levels remained approximately constant or, in some cases, increased slightly at low levels of triclosan or TMP. However, biomass levels then decreased significantly as the triclosan or TMP concentrations approached lethal levels. For S. sanguis, levels of hydrolytic enzymes (acid phosphatase, leucine aminopeptidase and esterase) generally remained approximately constant or increased with increasing concentrations of triclosan or TMP until close to inhibitory levels where enzyme levels were reduced. The ratio of extracellular soluble enzymes to cell-bound enzymes remained constant or increased slightly with increasing levels of triclosan or TMP. For C. gingivalis, production of hydrolytic enzymes (neutral phosphatase, leucine aminopeptidase and trypsin-like protease) remained constant or were reduced when grown with low levels of triclosan and TMP but in some cases increased with higher levels of agents. The proportion of extracellular soluble activity increased significantly when concentrations of agent neared inhibitory levels. The results taken together show that the physiology of cells is significantly altered and that hydrolytic enzymes are released from the cells when these are grown in the presence of increasing concentrations of triclosan or TMP. Enzyme release is more pronounced in the Gram-negative C. gingivalis and indicates that triclosan or TMP can cause membrane perturbation with subsequent release of membrane-located (S. sanguis) or periplasmic (C. gingivalis) hydrolytic enzymes. S. sanguis was more sensitive to triclosan than TMP while C. gingivalis was more sensitive to TMP. This suggests that, in C. gingivalis, TMP may diffuse into the cell wall more easily than triclosan and then be converted to triclosan by phosphatase activity within the cell wall complex, where it may give rise to high localized concentrations and subsequent cell damage.

Capnocytophaga gingivalis: effects of glucose concentration on growth and hydrolytic enzyme production.

In chemostat culture, the microaerophilic, CO2 requiring, gingival-plaque-associated bacterium Capnocytophaga gingivalis responded to the addition of glucose (1-6 g I-1) by doubling its growth rate and increasing its biomass yield fivefold. The data suggest that the glucose is catabolized by a fully aerobic route. Rather than repressing hydrolytic enzymes which might be associated with pathogenic properties, glucose enhanced the specific activity of aminopeptidase, trypsin-like protease, acid and alkaline phosphatase and alpha-glucosidase in comparison with a control culture grown in a tryptone/thiamin medium. Thus, the supply of glucose could be of importance in maximizing the pathogenic potential of this organism.

Effects of sublethal exposure to an antiseptic mouthrinse on representative plaque bacteria.

Although the mechanism responsible for the clinical antiplaque efficacy of oral antiseptics is generally considered to be primarily one of bactericidal activity, it has been suggested that oral antiseptics may have additional effects on bacteria exposed to sublethal levels. Studies reported herein, investigated the effects of sublethal levels of an essential oil-containing antiseptic mouthrinse (Listerine Antiseptic, Warner-Lambert Co., Morris Plains, NJ) on selected activities of representative plaque microorganisms using in vitro models. These studies demonstrated that sublethal exposure to the tested oral antiseptic can have significant effects in reducing intergeneric coaggregation, increasing bacterial generation time, and extracting endotoxin from Gram-negative bacteria. These in vitro activities can be correlated with features of plaque formation and pathogenicity seen in vivo; however, additional studies will be necessary to confirm that these mechanisms are, in fact, operative clinically.