Denaturing gradient gel electrophoresis analysis to study bacterial community structure in pockets of periodontitis patients.

Bacteria are involved in the onset and progression of periodontitis. A promising molecular technique, denaturing gradient gel electrophoresis (DGGE), to study microbial population dynamics in the subgingival pocket is presented. Twenty-three samples were taken from the subgingival pockets of nine patients and six healthy family members. From four periodontitis patients, 12 samples were evaluated before, 1 day after and 3 months after treatment. Part of the 16S rRNA gene of all bacteria was amplified by PCR and separated by DGGE, creating banding patterns representative of the community structure. Shifts in composition and diversity of the microbial population could be determined semiquantitatively, and this showed that treatment resulted in a decrease in the diversity of the population. After 3 months a microbial population 33-47% different from the population before treatment had re-established. Intense bands representing Exiguobacterium aurantiacum were present in 13 out of 25 samples, indicating that this species may play a role in periodontal disease.

[Plastic periodontal surgery]. / Plastische parodontale chirurgie.

In this article some indications and procedures of mucogingival and plastic periodontal surgery are reviewed. Frenectomy, vestibulary extension and free gingival graft, which all are widely used among periodontitsts, are described. In the second part of this review different surgical methods for covering recession-like periodontal defects are discussed. The clinical results of coronally advanced flap, connective tissue graft, and guided tissue regeneration are compared. The preference of the Nijmegen periodontal clinic to use coronally advanced flap--with or without connective tissue graft--is explained.

Some effects of non-surgical therapy on gingival inflammatory cell subsets in patients with early-onset periodontitis associated with Actinobacillus actinomycetemcomitans.

BACKGROUND: Limited information is available as to whether local cellular immunity in early-onset periodontitis (EOP) subjects harboring Actinobacillus actinomycetemcomitans (Aa) differs from that in patients without Aa. In addition, the effect of scaling and root planing on various lymphocyte subsets is described rather sparsely. METHODS: In 10 subjects with early-onset periodontitis harboring Aa (EOP-Aa) and in 10 subjects without Aa (EOP-nonAa), clinical measurements were recorded and gingival biopsies were performed before and after scaling and root planing. The specimens were cut into serial sections; using the alkaline phosphatase-antialkaline phosphatase technique, monoclonal antibodies to CD20 (B cells), CD30 (plasma cells), and CD45RO (T-memory cells) were applied as well as polyclonal antibodies to alpha, gamma, and mu chains (Ig A, G, and M). Cells were counted from an area of 0.25 mm2 in areas showing the largest infiltration. RESULTS: Before therapy, mean counts of all cell phenotypes were found to be markedly enhanced in the EOP-Aa group compared to EOP-nonAa subjects. Following scaling and root planing, the numbers of all phenotypes decreased in both groups. However, comparing the data before and after therapy in the EOP-Aa group, the P value was <0.05 only for CD30-positive cells. In the EOP-nonAa group, the differences before and after therapy reached statistical significance (P<0.05) for all cell counts, except for IgM-positive cells. Furthermore, reduction of probing depth and gain of clinical attachment reached the 0.05 level of statistical significance only in EOP-nonAa subjects. CONCLUSIONS: In EOP subjects harboring Aa, inflammatory cell subsets were detected in 2- to 3-fold higher numbers compared to patients without Aa. Scaling and root planing resulted in a decrease of all cell phenotypes studied in individuals without Aa, whereas in subjects with Aa, the only significant decrease that was seen occurred in plasma cells.

Bacterial susceptibility to amoxicillin and potassium clavulanate in advanced periodontitis patients not responding to mechanical therapy.

BACKGROUND, AIMS: Between 4 and 8% of periodontitis patients are reported to respond poorly to conventional therapy. In these cases, adjunctive use of systemic antibiotics might be a reasonable therapeutic approach. The purpose of this study was to evaluate the effects of systemic amoxicillin/clavulanate as adjunct to periodontal surgery on the predominant subgingival microorganisms in patients not responding to mechanical therapy. Furthermore, the bacterial susceptibility to amoxicillin/clavulanate was analyzed before and after therapy in order to assess the clinical validity of pre-therapeutic susceptibility testing. METHODS: In 10 periodontitis subjects with no subgingival detection of Actinobacillus actinomycetemcomitans, the predominant subgingival organisms were identified using the identification system Rapid ID 32 A as well as antibiotic susceptibility was tested utilizing the E test. RESULTS: Porphyromonas gingivalis and Prevotella oralis were detected in 7/10 subjects and could no more recovered after therapy. Fusobacterium nucleatum and Peptostreptococcus micros were present in 5/10 patients before treatment, but could be detected in 6/10, resp. 3/10 after therapy. In 4/10 subjects harboring F. nucleatum and in 3/10 with P. micros, those organisms were not targeted by amoxicillin/clavulanate, although post-treatment testing revealed their alleged susceptibility (MICs varied from 0.023 to 0.032 microg/ml, resp. from 0.125 to 2.0 microg/ ml). CONCLUSIONS: The results of this study suggest that the outcomes of conventional methods of susceptibility testing have to be interpreted very carefully when being used for treatment of plaque-related diseases. Furthermore, since the endpoint of systemic antibiotic treatment as adjunct to conventional therapy is elimination of F. nucleatum or P. micros in patients harboring these organisms, the use of amoxicillin/clavulanate appears not to be justified.

Some effects of non-surgical therapy on gingival inflammatory cell subsets in patients with adult and early-onset periodontitis.

BACKGROUND: Limited information is available to determine if there is a distinction in local cellular immunity between adult and early-onset periodontitis. Furthermore, the effect of scaling and root planing on various lymphocyte subsets is sparsely described. METHODS: Clinical measurements were recorded and gingival biopsies were performed before and after scaling and root planing in 10 subjects with adult (AP) and in 10 with early-onset periodontitis (EOP). The specimens were cut into serial sections and, using the alkaline phosphatase-anti-alkaline phosphatase technique, monoclonal antibodies to CD20 (B-cells), CD30 (plasma cells), and CD45RO (T-memory cells) were applied, as well as polyclonal antibodies to alpha, gamma, and mu chains (Ig A, G, and M). Areas showing the largest infiltration cells were counted. RESULTS: Mean counts of all cell phenotypes in the AP versus the EOP group did not show any significant differences before therapy (P >0.05). Following scaling and root planing, numbers of all phenotypes decreased in both groups. Comparing the data before and after therapy, P values were >0.05 in the AP group, except for IgA-positive cells. In the EOP group, the differences before and after therapy reached statistical significance (P<0.05) for all cell counts, except for IgM-positive cells. Furthermore, reduction of probing depth and gain of clinical attachment reached the 0.05 level of statistical significance only in EOP subjects. CONCLUSIONS: Local cellular immunity in patients with adult onset periodontitis does not appear to differ from the immune response in patients with early onset periodontitis. Scaling and root planing causes a decrease in the inflammatory cells subsets tested, however, this decline seems to be more pronounced in EOP than in AP subjects.

Fluoroquinolones in the treatment of Actinobacillus actinomycetemcomitans-associated periodontitis.

BACKGROUND: Periodontitis patients harboring Actinobacillus actinmycetemcomitans (Aa) are prime candidates for systemic antibiotic therapy. Besides tetracycline and the combination of metronidazole and amoxicillin the fluoroquinolones are also believed to have antibacterial activity against Aa. The aim of the present study was to evaluate systemic ofloxacin therapy as adjunct to flap surgery. METHODS: Twenty-five adult periodontitis patients with subgingival detection of Aa were treated with 2x200 mg/d ofloxacin for 5 days as adjunct to open flap surgery (test). Another 10 patients received only flap surgery (control). Probing depth (PD) and clinical attachment level (CAL) was recorded and subgingival plaque samples were cultivated on TSBV agar for detection of Aa at baseline as well as 3 and 12 months following therapy. RESULTS: At 3 and 12 months following therapy mean PD at monitored sites in the test group changed from 6.8 mm (+/-1.3) to 3.6 mm (+/-1.0), 3.8 mm (+/-1.1) and CAL from 7.5 mm (+/-1.4) to 5.4 mm (+/-1.4), 5.5 mm (+/-1.3). In the control group PD changed from 6.5 mm (+/-0.7) to 4.0 mm (+/-1.7), 4.1 mm (+/-1.6) and CAL from 7.5 mm (+/-1.0) to 6.3 mm (+/-1.7), 6.4 mm (+/-1.8). P was <0.05 for CAL between groups. Three and 12 months following adjunctive systemic ofloxacin therapy, Aa was suppressed below detectable levels in 22 of 22, test patients, whereas Aa could not be recovered in only 2 of the 10 controls. (P<0.0001). CONCLUSIONS: Systemic ofloxacin as adjunct to open flap surgery is able to suppress A. actinomycetemcomitans below detectable level in patients harboring this organism at baseline.

Intraoral persistence of Actinobacillus actinomycetemcomitans in periodontally healthy subjects following treatment of diseased family members.

A. actinomycetemcomitans (Aa) can be transmitted among family members. The purpose of the present study was, to evaluate, whether A. actinomycetemcomitans is able to persist for a prolonged period of time in periodontally healthy subjects following elimination of this pathogen from diseased family members. In 14 periodontitis patients harboring Aa as well as in their 37 family members, the clinical (PPD, PAL, P1I, GI) and microbiological status (TSBV) was evaluated. After study patients received mechanical, surgical and antibiotic (3x375 mg amoxicillin+3x250 mg metronidazole/7 d) treatment, they, as well as their family members, were reassessed 3, 6, 9 and 12 months following therapy. From 13 spouses, 4 (31%) had periodontitis associated with Aa, 4 (31%) had periodontitis and no Aa, 2 (15%) were periodontally healthy and harbored Aa, 3 (23%) were healthy with no detection of Aa. From 24 off-springs, one (4%) had periodontitis associated with Aa, 8 (33%) were healthy and harbored Aa, 15 (63%) were healthy with no detection of Aa. In the 2 spouses and in 7 of the 8 offsprings being healthy, but harboring Aa, this pathogen still persisted at the 12-month appointment, after it had been eliminated from the diseased family members. Only one child became negative for Aa after 12 months. The results of this study indicate that A. actinomycetemcomitans is able to persist in the oral habitats of periodontally-healthy subjects for at least 12 months, after this pathogen had been eliminated from diseased family members. This might suggest that also in healthy family members, A. actinomycetemcomitans as a source of transmission should be eliminated to prevent reinfection.

Antibiotic susceptibility of putative periodontal pathogens in advanced periodontitis patients.

In the present study, the antibiotic susceptibility of most prevalent micro-organisms in advanced periodontitis patients was evaluated. In 56 patients, pooled subgingival plaque samples were taken from the deepest site of each quadrant and were cultivated anaerobically. From each patient, the 4 most frequently encountered types of bacterial colonies were subcultured and identified (Rapid ID 32 A). From all bacterial species identified in the 224 subcultures, the 4 most prevalent were used for susceptibility testing to tetracycline, metronidazole and amoxicillin/clavulanate using the E Test. The most prevalent microorganisms were Fusobacterium nucleatum (38/214), Peptostreptococcus micros (33/214), Prevotella oralis (33/214) and Porphyromonas gingivalis (32/214). Regarding antibiotic susceptibility it could be shown that minimal inhibitory concentration (MIC) in all cases was below antibiotic concentrations achievable in gingival crevicular fluid. However, antibiotic resistance was seen in 3 to 29% of the investigated microorganisms.

Microbiological and immunohistological findings in a patient with Papillon-Lefèvre syndrome.

The following communication is a case history of an 11 year-old female patient suffering from Papillon-Lefèvre syndrome. Since a massive occurrence of A. actinomycetemcomitans had been found in the subgingival microflora of the periodontal pockets, the patient was treated with repeated subgingival scaling, with an adjunct Amoxicillin and Metronidazol treatment. A bacteriological examination of the girl's family proved that several brothers and sisters as well as one parent also carried. A. actinomycetemcomitans, showing 3 different strains of this bacterium within the family. An immunohistological examination of the gingival tissue showed a massive inflammatory infiltrate which was dominated by plasma cells. The histological investigation of the first molars did not show morphological abnormalities of the root cementum. Posttreatment clinical and radiographical improvement of the periodontal conditions is reported despite the recurrent finding of A. actinomycetemcomitans.

[Papilla-preserving flap procedure in periodontal implantation of alloplastic materials. Comparative study]. / Die papillenerhaltende Lappen-Technik bei der parodontalen Implantation alloplastischer Materialien im Vergleich.

40 periodontal defects with probing depths of 5 mm in 10 patients were treated with a conventional flap or with the "papilla preservation flap". In addition, half of the defects received granular hydroxyapatite. In terms of pocket depth and probing attachment-level no significant differences were seen between the 4 groups after 3 and 6 months. Regarding the development of gingival recession, however, the papilla preservation technique has significant advantages and showed better aesthetic results.