Prevalence of ß-lactamase-producing bacteria in human periodontitis.

BACKGROUND AND OBJECTIVE: Beta-lactam antibiotics prescribed in periodontal therapy are vulnerable to degradation by bacterial ß-lactamases. This study evaluated the occurrence of ß-lactamase-positive subgingival bacteria in chronic periodontitis subjects of USA origin, and assessed their in vitro resistance to metronidazole at a breakpoint concentration of 4 µg/mL. MATERIAL AND METHODS: Subgingival plaque specimens from deep periodontal pockets with bleeding on probing were removed from 564 adults with severe chronic periodontitis before treatment. The samples were transported in VMGA III and then plated onto: (i) nonselective enriched Brucella blood agar (EBBA) and incubated anaerobically for 7 d; and (ii) selective trypticase soy-bacitracin-vancomycin (TSBV) and incubated for 3 d in air + 5% CO2 . At the end of the incubation periods, the bacterial test species were identified and quantified. Specimen dilutions were also plated onto EBBA plates supplemented with 2 µg/mL of amoxicillin, a combination of 2 µg/mL of amoxicillin plus 2 µg/mL of the ß-lactamase inhibitor clavulanic acid, or 4 µg/mL of metronidazole, followed by anaerobic incubation for 7 d. Bacterial test species presumptively positive for ß-lactamase production were identified by growth on EBBA primary isolation plates supplemented with amoxicillin alone and no growth on EBBA primary isolation plates containing both amoxicillin plus clavulanic acid. A subset of such isolates was subjected to nitrocefin-based chromogenic disk testing to confirm the presence of ß-lactamase activity. In vitro resistance to 4 µg/mL of metronidazole was noted when growth of test species occurred on metronidazole-supplemented EBBA culture plates. RESULTS: Two-hundred and ninety-four (52.1%) of the study subjects yielded ß-lactamase-producing subgingival bacterial test species, with Prevotella intermedia/nigrescens, Fusobacterium nucleatum and other Prevotella species most frequently identified as ß-lactamase-producing organisms. Of the ß-lactamase-producing bacterial test species strains recovered, 98.9% were susceptible in vitro to metronidazole at 4 µg/mL. CONCLUSION: The occurrence of ß-lactamase-positive subgingival bacterial species in more than half of the subjects with severe chronic periodontitis raises questions about the therapeutic potential of single-drug regimens with ß-lactam antibiotics in periodontal therapy. The in vitro effectiveness of metronidazole against nearly all recovered ß-lactamase-producing subgingival bacterial species further supports clinical periodontitis treatment strategies involving the combination of systemic amoxicillin plus metronidazole.

Subgingival microbiota in adult Down syndrome periodontitis.

BACKGROUND AND OBJECTIVE: The subgingival microbiota in Down syndrome and non-Down syndrome adults receiving periodic dental care was examined for 40 bacterial species using checkerboard DNA-DNA hybridization and the results were related to clinical periodontal attachment loss. MATERIAL AND METHODS: A total of 44 Down syndrome, 66 non-Down syndrome mentally retarded and 83 mentally normal adults were clinically evaluated. This involved, for each subject, the removal of subgingival specimens from three interproximal sites on different teeth; all subgingival samples per subject were then pooled and assessed for the presence and levels of 40 bacterial species using species-specific whole-genomic DNA probes and checkerboard DNA-DNA hybridization. Significant group differences in species proportions averaged across subjects were evaluated using the Kruskal-Wallis test, and associations between subgingival species and mean subject attachment loss within Down syndrome and non-Down syndrome subject groups were quantified using Pearson correlation and multiple linear regression analysis. RESULTS: Down syndrome subjects exhibited greater attachment loss than non-Down syndrome subjects (p=0.05). Most microbial species were present in Down syndrome subjects at levels similar to non-Down syndrome subjects, except for higher proportions of Selenomonas noxia, Propionibacterium acnes, Streptococcus gordonii, Streptococcus mitis and Streptococcus oralis in Down syndrome subjects compared with non-Down syndrome study subjects, higher proportions of Treponema socranskii in Down syndrome subjects compared with non-Down syndrome mentally retarded subjects, and higher proportions of Streptococcus constellatus in Down syndrome subjects compared with mentally normal subjects. Down syndrome adults classified with periodontitis revealed higher subgingival levels of T. socranskii than Down syndrome subjects with no periodontitis (p=0.02). Higher subgingival proportions of S. constellatus, Fusobacterium nucleatum ssp. nucleatum, S. noxia and Prevotella nigrescens showed significant positive correlations (r=0.35-0.42) and higher proportions of Actinomyces naeslundii II and Actinomyces odontolyticus showed negative correlations (r=-0.36 to -0.40), with increasing mean subject attachment loss in Down syndrome adults. CONCLUSION: Individuals with Down syndrome show higher levels of some subgingival bacterial species and specific associations between certain subgingival bacterial species and loss of periodontal attachment. These findings are consistent with the notion that certain subgingival bacteria may contribute to the increased level of periodontal disease seen in Down syndrome individuals and raise the question as to the reason for increased colonization in Down syndrome.

Actinobacillus actinomycetemcomitans and Porphyromonas gingivalis subgingival presence, species-specific serum immunoglobulin G antibody levels, and periodontitis disease recurrence.

BACKGROUND AND OBJECTIVE: The biological and clinical effects of antibody against periodontal pathogenic bacteria are incompletely understood. This study evaluated the inter-relationships among periodontal levels of cultivable Actinobacillus actinomycetemcomitans and Porphyromonas gingivalis, species-specific serum immunoglobulin G (IgG) antibody levels, and periodontitis disease activity. MATERIAL AND METHODS: Forty-three adults who had previously been treated for periodontitis and who also harbored cultivable A. actinomycetemcomitans or P. gingivalis were evaluated semiannually for clinical disease recurrence over a 36-month period. Each patient provided subgingival microbial samples, for the recovery of A. actinomycetemcomitans and P. gingivalis, from the two deepest pockets in each dentition sextant. A. actinomycetemcomitans and P. gingivalis serum IgG antibody levels were assessed using enzyme-linked immunosorbent assay (ELISA), together with whole-cell sonicate extracts from A. actinomycetemcomitans serotypes a-c and P. gingivalis ATCC 33277. Data were analyzed using the Mantel-Haenszel chi-square and Fisher exact two-tailed tests. RESULTS: Eighteen (60.0%) of 30 A. actinomycetemcomitans-positive subjects, and 10 (76.9%) of 13 P. gingivalis-positive subjects, exhibited recurrent periodontal breakdown within 36 months of periodontal therapy. Nineteen (67.9%) of the 28 patients with active periodontitis had A. actinomycetemcomitans or P. gingivalis serum antibody levels below designated threshold values. In comparison, 10 (66.7%) of 15 culture-positive clinically stable subjects showed A. actinomycetemcomitans or P. gingivalis serum antibody levels above threshold values. The difference between specific antibody levels in periodontitis-active and periodontitis-stable patients was statistically significant (p = 0.032). CONCLUSIONS: Serum levels of IgG antibodies against A. actinomycetemcomitans or P. gingivalis in periodontitis-stable patients were higher than those in patients with active periodontitis. The results suggest that elevated levels of IgG antibody against A. actinomycetemcomitans and P. gingivalis have a detectable protective effect against periodontal infections with these microorganisms.

A new device for measuring density of jaw bones.

OBJECTIVES: The purpose of this project was to develop a lightweight, simple device to evaluate alveolar process bone density using normal intraoral and extraoral imaging procedures. METHODS: A simple lightweight device was constructed using barium sulfate as the major radiopaque component. The 5 x 32 x 12 mm(3) resin block has eight segments with known densities ranging from 1.304 (g/cm(3)) to 1.982 (g/cm(3)). The device was integrated into an XCP unit for standard intraoral radiographs and placed between the jaws for computer aided tomographic imaging. The relationship between the device segment densities and the optical densities of the exposed film was plotted. RESULTS: A linear inverse relationship was found between the device segment densities and optical densities when segment densities were between 1.304 (g/cm(3)) to 1.882 (g/cm(3)). However, the relationship was non-linear for segment densities above 1.882 (g/cm(3)). CONCLUSIONS: Normal human bone density is 1.85 (g/cm(3)), and this densitometer is useful for determination of material densities from 1.304 (g/cm(3)) to 1.882 (g/cm(3)). The device may be useful for precise bone density assessment.

Aggregation of human platelets by gingipain-R from Porphyromonas gingivalis cells and membrane vesicles.

The hypothesis that there is an association between periodontitis and cardiovascular disease suggests new lines of research on the mechanism whereby oral bacteria might exert systemic effects. This study was conducted to ascertain and quantitate the effect of Porphyromonas gingivalis on human platelets in vitro. A second related objective was to purify and identify the aggregating vector. Aggregation was measured by platelet turbidometry and gingipain-R was purified from P. gingivalis membrane vesicles by Sepharose 2B and hydroxyapatite chromatography. The in vitro aggregation of platelets requires that at least 1.0 x 10(4) cells be stirred with 1.35 x 10(8) platelets. The specific activity is substantially increased in the membrane vesicles that are shed by this bacterium. Aggregability was due to gingipain-R activity, a potent cysteine protease that was found to be highly concentrated in the membrane vesicle fraction. The enzyme was purified 18-fold in high yield from the membrane vesicles, and consists of two noncovalently linked proteins that migrate at 49 and 44 kDa on SDS-PAGE. Aggregation of platelets by gingipain-R was shown to be dose-dependent, and inhibited by leupeptin and arginine, but not by anti-thrombin III. This is the first report enumerating the specific number of cells and lowest concentration of membrane vesicles necessary to evoke a full human platelet response, and the first report to assign this activity to gingipain-R.

Effect of an essential oil-containing antiseptic mouthrinse on induction of platelet aggregation by oral bacteria in vitro.

BACKGROUND: With an increasing body of data suggesting an association between periodontitis and cardiovascular disease, studies have been conducted to elucidate potential mechanisms by which oral bacteria might exert systemic effects. 2 oral bacteria, Streptococcus sanguis and Porphyromonas gingivalis, have been shown to induce platelet aggregation in vitro. This study was conducted to determine the effect of treatment with an essential oil mouthrinse (Listerine Antiseptic) on the platelet-aggregating activity of these organisms. METHOD: Bacteria were grown under standard culture conditions. S. sanguis ATCC strain 10556 was exposed for 3 min to the essential oil mouthrinse at either full strength or a 1:1 dilution, while P. gingivalis FDC strain 381 was exposed to the essential oil mouthrinse at a 1:10 dilution. Positive control cells were treated with Hanks balanced salt solution (HBSS). Aggregation was measured using a recording platelet aggregometer. The assay of each organism in its respective mouthrinse dilution(s) or HBSS was repeated 5 times. RESULTS: In all cases, the HBSS-treated organisms induced platelet aggregation, with mean(+/-S.E.) lag times of 12.30 (+/-1.36) min and 11.36 (+/-0.58) min for P. gingivalis and S. sanguis, respectively. In contrast, treatment with the essential oil mouthrinse completely inhibited the platelet aggregating activity of P. gingivalis and of S. sanguis exposed to the 1:1 mouthrinse dilution in all assays; the aggregating activity of S. sanguis treated with full-strength mouthrinse was completely inhibited in 4 of 5 assays, and inhibited by 75% in the 5th, for a mean inhibition of 95 +/- 1.5%. CONCLUSION: This study provides additional evidence that the essential oil mouthrinse can interfere with bacterial cell surface-associated activities which may have clinical relevance.

Management of gingival overgrowth associated with generalized enamel defects in a child.

Gingival overgrowth is usually associated with systemic conditions or treatment (e.g. blood dyscrasias, anti-epileptic or immunosuppressive agents). A child is presented, who had enlarged gingiva associated with a generalized enamel defect (amelogenesis imperfecta (AI), hypoplastic type) and document the periodontal and restorative management of this case.

Clinical and microbiological effects of adjunctive antibiotics in treatment of localized juvenile periodontitis. A controlled clinical trial.

The aim of this study was to assess the clinical and microbiologic effects of the combination of amoxicillin and metronidazole therapy as an adjunct to mechanical treatment in the management of localized juvenile periodontitis. Twenty-five localized juvenile periodontitis (LJP) patients from a Brazilian population were randomly allocated into an experimental group receiving mechanical treatment and antibiotics, and a control group receiving mechanical treatment and placebo. Clinical and radiographic assessments, as well as microbiologic sampling for Actinobacillus actinomycetemcomitans, were performed at baseline and one year after the end of the treatment. At the termination of the study A. actinomycetemcomitans could be isolated from the oral cavity of all patients in the control group who harbored the bacterium at baseline and in 4 out of 8 patients in the experimental group. Both treatment modalities resulted in significant benefit on an individual basis. The experimental group, however, displayed better results than did the control group regarding gingival index (GI), probing depth (PD), clinical attachment level (CAL), and radiographic analysis of crestal alveolar bone mass, but not with respect to plaque index (PI). No serious adverse effects of the antibiotic treatment were observed in the present study.

Socio-behavioral determinants of oral hygiene practices among USA ethnic and age groups.

In this study, socio-behavioral determinants of oral hygiene practices were examined across several dentate ethnic and age groups. Oral hygiene scale scores were constructed from toothbrushing and dental floss frequencies self-reported by population-based samples of middle-aged (35-44 years) and older (65-74 years) dentate adults representing Baltimore African-American and White, San Antonio Hispanic and non-Hispanic White, and Navajo and Lakota Native American persons participating in the WHO International Collaborative Study of Oral Health Outcomes (ICS-II) survey. Female gender, education, certain oral health beliefs, household income, and the presence of a usual source of care were revealed with multivariate analysis to show a significant positive relationship with higher oral hygiene scale scores (indicating better personal oral hygiene practices). Other socio-behavioral variables exhibited a more varied, ethnic-specific pattern of association with oral hygiene scale scores.

Human Herpesviridae in acute necrotizing ulcerative gingivitis in children in Nigeria.

Herpesviruses have been implicated in the pathogenesis of human periodontitis. The present study investigated whether herpesviruses are present in the lesions of acute necrotizing ulcerative gingivitis. Sixty-two Nigerian children, aged 3-14 years, were studied. Twenty-two children had acute necrotizing ulcerative gingivitis and were also malnourished, 20 exhibited no acute necrotizing ulcerative gingivitis but were malnourished and 20 were free of acute necrotizing ulcerative gingivitis and in a good nutritional state. Polymerase chain reaction methods were used to determine the presence of human cytomegalovirus (HCMV), Epstein-Barr virus type 1 and type 2 (EBV-1, EBV-2), herpes simplex virus (HSV), human herpes virus 6 (HHV-6), human papilloma virus and human immunodeficiency virus type 1 in crevicular fluid specimens collected by paper points. Of the 22 acute necrotizing ulcerative gingivitis patients, 15 (68%) revealed viral infection and 8 (36%) viral coinfection. Thirteen (59%) acute necrotizing ulcerative gingivitis patients demonstrated HCMV, 6 (27%) EBV-1, 5 (23%) HSV and 1 (5%) HHV-6. Only 2 (10%) subjects from each group not affected by acute necrotizing ulcerative gingivitis showed viral presence, and no control subject revealed viral coinfection. These findings suggest that HCMV and possibly other herpesviruses contribute to the onset and/or progression of acute necrotizing ulcerative gingivitis in malnourished Nigerian children.

PIP: Recent laboratory studies have implicated herpesviruses in the pathogenesis of human periodontitis. This present study examines whether herpesviruses are present in the lesions of acute necrotizing ulcerative gingivitis. A total of 62 Nigerian children, aged 3-14 years were studied: 22 had acute necrotizing ulcerative gingivitis and were also malnourished, 20 showed no acute necrotizing ulcerative gingivitis but were malnourished, and 20 were free of acute necrotizing ulcerative gingivitis and in a good nutritional state. A polymerase chain reaction assay was used to determine the presence of human cytomegalovirus (HCMV), Epstein-Barr virus type 1 and type 2 (EBV-1, EBV-2), herpes simplex virus (HSV), human herpes virus 6 (HHV-6), human papilloma virus, and HIV-1 in crevicular fluid samples obtained by paper points. Of the 22 patients with acute necrotizing ulcerative gingivitis, 15 (68%) showed viral infection and 8 (36%) showed a viral coinfection. In addition, 13 (59%) of these patients demonstrated HCMV, 6 (27%) EBV-1, 5 (23%) HSV, and 1 (5%) HHV-6. Only 2 (10%) subjects from each group not affected by acute necrotizing ulcerative gingivitis revealed viral presence, and none of the control group demonstrated viral coinfection. The findings suggest that HCMV and possibly other herpesviruses contribute to the onset and/or progression of acute necrotizing ulcerative gingivitis in malnourished children in Nigeria.

Utility of 5 major putative periodontal pathogens and selected clinical parameters to predict periodontal breakdown in patients on maintenance care.

The predictive utility of 5 major putative periodontopathic microbial species, "superinfecting" organisms, and several clinical periodontal parameters were assessed relative to periodontitis recurrence over a 12-month period in 78 treated adult patients participating in a 3-month maintenance care program. At baseline, pooled subgingival microbial samples were collected from each patient, and whole-mouth evaluations of probing depth, relative periodontal attachment level, furcation involvement, and indices of plaque and gingival inflammation were carried out. 67 (85.9%) subjects were culture-positive at baseline for presence of either Actinobacillus actinomycetemcomitans, Porphyromonas gingivalis, Prevotella intermedia, Campylobacter rectus or Peptostreptococcus micros, with 48 (61.5%) subjects yielding one or more of these species at or above designated threshold proportions of > or = 0.01% for A. actinomycetemcomitans, > or = 0.1% for P. gingivalis, > or = 2.5% for P. intermedia, > or = 2.0% for C. rectus, and > or = 3.0% for P. micros. Subgingival yeasts were recovered from 12 subjects, staphylococci from 7, and enteric rods/pseudomonads from 6; however, no subjects revealed > or = 1.0% baseline proportions of these "superinfecting" organisms in subgingival specimens. Periodontitis recurrence in subjects was defined as any periodontal site exhibiting either a probing depth increase of > or = 3 mm from baseline, or a probing depth increase of > or = 2 mm from baseline together with a loss in relative periodontal attachment of > or = 2 mm from baseline. 15 (19.2%) study subjects showed periodontitis recurrence within 6 months of baseline, and 25 (32.1%) within 12 months. The mere baseline presence of the 5 major test species and "superinfecting" organisms were not significant predictors of periodontitis recurrence over 12 months. However, a 2.5 relative risk for periodontitis recurrence over 12 months was found for subjects yielding one or more of the 5 major test species at or above the designated baseline threshold proportions (p = 0.022, Mantel-Haenszel chi 2 test). The positive predictive value for periodontitis recurrence of a microbiologic analysis encompassing the 5 major test species at or above the designated threshold proportions improved with increasing time from baseline, up to approximately 42% at 12 months. Baseline variables jointly providing in multiple regression analysis the best predictive capability for periodontitis recurrence in subjects over a 12-month period were recovery of one or more of the 5 major test species at or above designated threshold proportions, the proportion of sites per subject with > or = 5 mm probing depth, and the mean whole-mouth probing depth. These findings indicate that one or more of 5 major putative periodontal pathogens in elevated subgingival proportions together with increased probing depth predispose adults on maintenance care to recurrent periodontitis.

Efficacy of CPITN sextant scores for detection of periodontitis disease activity.

The relationship between CPITN sextant scores and periodontitis recurrence at individual tooth sites was evaluated in a longitudinal study in 83 treated adult periodontitis patients receiving systematic 3-month maintenance care. At baseline and semi-annual examinations over 36 months, CPITN scores were assigned to each dentition sextant using probing depths and gingival index scores, and relative periodontal attachment level was assessed at individual tooth sites using an occlusal reference stent. Periodontitis recurrence was defined as any periodontal site exhibiting either a probing depth increase of > or = 3 mm from baseline, or a probing depth increase of > or = 1 mm from baseline together with a loss of relative periodontal attachment of > or = 2 mm from baseline. 49 (59.0%) subjects developed periodontitis recurrence in 147 (29.8%) sextants at 181 (2.2%) individual periodontal sites during the 36-month study period. Baseline CPITN scores of 4 were more common in disease-active subjects than clinically-stable subjects (p = 0.003, t-test), and were associated with a statistically significant 1.66 relative risk of periodontitis recurrence within 36 months. CPITN sextant scores of 3 or 4 showed low specificity and low positive predictive values as indicators of periodontitis recurrence at > or = 1 individual sites within the affected sextant. In comparison, low CPITN sextant scores (0-2) provided high specificity (96.2-100%), high positive predictive values (99.5-100%), and a summary odds ratio of 24.2 as an indicator of clinical stability at all periodontal sites within a given dentition sextant. Changes in sextant scores for CPITN over 6-month periods showed no relationship with periodontitis recurrence at individual periodontal sites. This study suggests that while CPITN is inadequate for detection of periodontitis recurrence, low CPITN scores provide rapid presumptive identification of clinically-stable sextants in adult periodontitis patients on maintenance care.

Utility of radiographic crestal lamina dura for predicting periodontitis disease-activity.

The relationship between radiographic crestal lamina dura and periodontitis disease-activity was studied longitudinally in 51 treated adult patients on a systematic 3-month maintenance program. The presence or absence of crestal lamina dura at 1809 interproximal sites was scored from periapical and bitewing radiographs taken at baseline of a 36-month maintenance care period. Semi-annual clinical evaluations by 2 independent examiners were carried out on each patient, with disease recurrence defined as sites revealing a > or = 3 mm increase in probing depth from baseline, or a > or = 2 mm increase in probing depth together with a > or = 2 mm loss of relative attachment level from an occlusal reference stent. Over the 36-month study period, 23 (45%) patients exhibited disease recurrence at 55 (3%) interproximal tooth sites scored for baseline crestal lamina dura. Absence of detectable baseline crestal lamina dura yielded high sensitivity (87-100%), but low specificity (17%) and low positive predictive values (0.8-3.2%), for localized periodontitis recurrence. In contrast, no sites exhibiting an intact baseline crestal lamina dura demonstrated periodontitis recurrence up to 24 months from baseline (100% positive predictive values). Presence of radiographic crestal lamina dura was positively associated with clinical periodontal stability (summary odds ratio for sites = 2.6, P = 0.0004), and negatively associated with periodontitis recurrence (summary odds ratio for sites = 0.4, P = 0.0004), for the 36-month study period. Evaluation of radiographic crestal lamina dura status appears valuable for assessing the risk of periodontitis disease-activity at inter-proximal tooth sites in patients on maintenance care programs.

Comparison of two pressure-sensitive periodontal probes and a manual periodontal probe in shallow and deep pockets.

Three periodontal probes--a manual probe and two computerized, pressure-sensitive probes--were studied to determine their relative recording accuracy. In the first part of the study, replicate probing measurements were taken with the three probes at shallow (less than 5 mm) and deeper (greater than or equal to 5 mm) periodontal sites. In the second part of the study, probing depths were determined on all teeth except third molars by the three probing techniques at a single appointment. Mean values between replicate probings for each instrument did not differ statistically, except for duplicate measurements with one of the electronic probes in pockets with depths greater than 5 mm. All three probes demonstrated higher standard deviations with increasing depth, which indicates decreased reproducibility of probing depth measurements. Results of this study suggest that an electronic, pressure-sensitive probe yields more reproducible probing depth measurements than a conventional manual periodontal probe.

Campylobacter rectus in human periodontitis.

Campylobacter rectus (formerly Wolinella recta) in periodontitis lesions was studied relative to age and sex distribution, relationship to disease-active periodontitis, response to periodontal debridement and in vitro antimicrobial susceptibility. Subgingival C. rectus was collected with paper points, transported in VMGA III and plated onto nonselective enriched brucella blood agar and Hammond's selective medium for C. rectus, both incubated anaerobically. C. rectus was recovered from 80% of 1654 periodontitis patients. Although the organism showed similar age and sex occurrence, its proportional recovery in culture-positive adults was inversely related to increasing age (r = 0.999, P < 0.001). The organism was positively associated (summary odds ratio = 2.95) with disease activity in a 24-month longitudinal study of 93 adult periodontitis patients on maintenance therapy. C. rectus decreased from 8.2% to 0.7% following local periodontal debridement of 20 culture-positive adult periodontitis patients. The organism exhibited high in vitro susceptibility to therapeutic levels of tetracycline hydrochloride, metronidazole, penicillin G and ciprofloxacin. These findings further delineate the epidemiology and potential pathogenic role of C. rectus in human periodontitis.