Clustering of subgingival microbial species in adolescents with periodontitis

It has become increasingly recognized that groups of microorganisms interact within the subgingival plaque of adult subjects with periodontitis. It is much less clear, however, whether the consortia of microorganisms associated with periodontitis are different in early and more advanced cases of periodontitis. To investigate this point further, subgingival plaque was collected from six sites in 87 adolescents with periodontitis and 73 controls and the samples were analyzed for the detection of 18 microbial species using the DNA-DNA hybridization technique. Actinomyces oris accounted for the highest proportion of the flora and was more predominant among controls. Prevotella nigrescens, Prevotella intermedia, Porphyromonas gingivalis, and Tannerella forsythia were present at higher levels among the subjects with periodontitis. Factor analyses identified one factor characterized by highly positive loadings for T. forsythia, Campylobacter rectus, P. gingivalis, P. intermedia, P. nigrescens, Parvimonas micra, and Treponema denticola, and another factor characterized by highly positive loadings of A. oris, Capnocytophaga ochracea, Eikenella corrodens, Streptococcus intermedius, Selenomonas noxia, Streptococcus oralis, Streptococcus sanguinis, and Veillonella parvula. Aggregatibacter actinomycetemcomitans and Streptococcus mutans did not load on any of the two factors, while Fusobacterium nucleatum loaded on both. These findings confirm the occurrence of clustering of subgingival bacteria according to case status also among young individuals.     

Relationship between periodontal pocket sulfide levels and subgingival species

BACKGROUND: Many species implicated in the pathogenesis of periodontal disease produce volatile sulfur compounds (VSC). This investigation examined the relationship between levels of sulfide and subgingival bacterial species in the same periodontal pockets. MATERIAL AND METHODS: Twenty chronic periodontitis subjects were measured clinically at six sites per tooth for plaque, gingivitis, bleeding on probing, suppuration, pocket depth and attachment level. Subgingival plaque samples, taken from the mesial aspect of each tooth, were individually analyzed for their content of 40 bacterial species using checkerboard DNA-DNA hybridization. Sulfide levels were measured at the same sites using a Diamond Probe/Perio 2000 system. Clinical and microbiological data were averaged for sulfide-positive and -negative sites separately in each subject and then averaged across subjects. Significance differences in clinical and microbial parameters between sulfide-positive and -negative sites were sought using the Wilcoxon signed ranks test. RESULTS: Mean total DNA probe counts (x10(5), +/-SEM) at sulfide-negative and -positive sites were 44.0 +/- 9.9 and 65.0 +/- 13.3, respectively (p < 0.01). Seventeen species were found at significantly higher levels in sulfide-positive than -negative sites. These included abundant producers of VSC such as members of the genera Fusobacterium, Campylobacter, Prevotella, Treponema and Eubacterium, and Bacteriodes forsythus, Selenomonas noxia and Propionibacterium acnes. Prevotella intermedia, Bacteriodes forsythus, Prevotella nigrescens, Fusobacterium nucleatum ss vincentii and Treponema denticola exhibited the greatest difference in mean counts between sulfide-negative and -positive sites. Orange and red complex species were at higher counts at shallow (< 4 mm) sulfide-positive than shallow sulfide-negative sites. Although not statistically significant, mean clinical parameters were somewhat higher at sulfide-positive than sulfide-negative sites. CONCLUSIONS: Intra-pocket sulfide levels reflect the levels of sulfide-producing species and may provide useful diagnostic information.     

慢性牙周炎龈下菌斑中五种牙周可疑致病微生物的分布

目的研究五种牙周可疑致病微生物在慢性牙周炎患者龈下菌斑的分布。方法选择27例慢性牙周炎患者,每位患者选取牙周袋最深的两个位点作为观察位点,采集龈下微生物样本,采用多重聚合酶链反应和反杂交的方法对伴放线菌嗜血菌、牙龈卟啉单胞菌、福赛斯坦纳菌、中间普雷沃菌和齿垢密螺旋体五种微生物进行半定量检测。结果在所检测的54个位点中,牙龈卟啉单胞菌、中间普雷沃菌、福赛斯坦纳菌和齿垢密螺旋体均有较高的检出率,分别为98.15%、92.59%、100%和98.15%;伴放线菌嗜血菌检出率较低,为20.37%。牙龈卟啉单胞菌和福赛斯坦纳菌的检出量明显高于其他三种微生物,其差异有统计学意义（P<0.05）。结论慢性牙周炎患者多存在牙龈卟啉单胞菌、福赛斯坦纳菌、中间普雷沃菌和齿垢密螺旋体的同时感染,且牙龈卟啉单胞菌和福赛斯坦纳菌的感染量较高。     

牙周病基础治疗前后龈下菌群的动态观察

目的：动态观察龈下细菌治疗前后在口腔内定植的变化，为牙周病病因学研究和治疗方案确定提供依据。方法：选取26例慢性牙周炎患者治疗前、治疗后1周、1个月和3个月时同一位点的龈下菌斑，测量牙周探诊深度，提DNA，洲浓度。扩增全细菌16SrRNA基因片段，变性梯度凝胶电泳分离，选择特异性的DNA条带，回收、测序。结果：测序结果表明治疗后消失的两个DNA条带与牙龈卟啉单胞菌有98％和99％的同源性；治疗后新出现的两个DNA条带与卟啉菌属的一种有99％的同源性。结论：牙龈卟啉单胞菌是慢性牙周炎的重要可疑致病菌。变性梯度凝胶电泳适用于分析大量微生物标本分布和类型。     

5种牙周龈下可疑致病微生物与慢性牙周炎局部不同牙周状态间的关系

目的 观察5种龈下微生物检出水平与慢性牙周炎局部牙周状态的关系。方法 选择20例慢性牙周炎患者的80个位点及10例牙周健康者的20个位点为观察位点,采集龈下微生物样本,记录牙周探诊深度（PD）,根据所测位点的PD进行分组。PD≤4 mm为A组,4 mm＜PD≤6 mm为B组,PD>6 mm为C组,健康对照组为H组。通过聚合酶链反应（PCR）和DNA探针反杂交技术半定量检测各组伴放线菌嗜血菌、牙龈卟啉单胞菌、福赛斯坦纳菌、齿垢密螺旋体和中间普氏菌的检出率和检出水平。结果 B、C组牙龈卟啉单胞菌、福赛斯坦纳菌、齿垢密螺旋体和中间普氏菌的检出率和检出水平均高于H组,A组牙龈卟啉单胞菌的检出率和检出水平也高于H组,C组福赛斯坦纳菌和齿垢密螺旋体检出水平高于B组,以上差异均有统计学意义（P<0.05）;伴放线菌嗜血菌在各组间的检出率及检出水平都无明显差异。结论 随着牙周袋的加深,牙龈卟啉单胞菌、福赛斯坦纳菌、齿垢密螺旋体和中间普氏菌体的阳性检出率和检出水平都有随之增加的趋势;牙龈卟啉单胞菌与慢性牙周炎的早期炎症关系较为密切,而福赛斯坦纳菌和齿垢密螺旋体与中重度慢性牙周炎炎症位点的严重程度有关。     

Smoking and subgingival microflora in periodontal disease

AIM: The present investigation was undertaken to analyze the influence of smoking on the periodontal disease associated subgingival microflora. The population included 33 smokers and 31 non-smokers in the age range 36-86 years. METHODS: Microbial samples were obtained from 4 sites per patient. The checker-board DNA-DNA hybridization technology was used for detection of the bacterial species P. gingivalis, P. intermedia, P. nigrescens, B. forsythus, A. actinomycetemcomitans, F. nucleatum, T. denticola, P. micros, C. rectus, E. corrodens, S. noxia and S. intermedius. RESULTS: Using score 1 as cutoff, contrasting colonized versus non-colonized patients, 8 out of 12 species were detected in > or = 90% of both smokers and non-smokers. Using score 4 as cutoff, contrasting heavily colonized patients versus non-colonized and less heavily colonized patients, the detection rates decreased in both smokers and non-smokers. No significant differences in detection rates were observed between smokers and non-smokers. Logistic regression analysis indicated that neither smoking, probing depth nor gingival bleeding influenced the occurrence of the species analyzed. The lack of a smoking exposure dose-response further supported the indication of a limited influence of smoking. CONCLUSION: Smoking exerts little, if any, influence on the subgingival occurrence of several of the bacteria most commonly associated with periodontal disease.     

Periodic subgingival antimicrobial irrigation of periodontal pockets

The present investigation was undertaken to study the clinical effect of professionally performed periodic subgingival irrigation per se and as an adjunct to scaling and root planing. 10 patients suffering from moderate-severe periodontal disease participated in the study. Following an initial 3-month period of supervised supragingival plaque control, a total of 102 periodontal sites with probing pocket depth greater than or equal to 6 mm and "bleeding on probing" were selected and subjected to a Baseline examination comprising assessments of oral hygiene and gingival conditions, probing depths and probing attachment levels. The pockets in the various jaw quadrants were randomly assigned to one of the following treatment groups: (1) periodic subgingival irrigation with hydrogen peroxide, (2) periodic subgingival irrigation with chlorhexidine, (3) periodic subgingival irrigation with saline and (4) no subgingival treatment. During the first part of the study (baseline-32 weeks), no mechanical debridement of the subgingival area was performed. The irrigation treatment was carried out by the operator 3 times per week during weeks 1 + 2 and 5 + 6 of the trial. In the 2nd part of the trial (32-52 weeks), the sites were subjected to scaling and root planing combined with professional irrigation during weeks 32-38. The previously non-irrigated control sites were not subjected to adjunctive irrigation when mechanically debrided. During the entire study, the patients were recalled for professional tooth cleaning once every 4 weeks. Re-examinations were carried out at 4, 6, 32, 40 and 52 weeks. The results revealed that repeated professional irrigation of unscaled periodontal pockets with chlorhexidine or hydrogen peroxide resulted in a temporarily reduced frequency of bleeding sites, but not in any clinically significant changes in probing assessments. A similar improvement of bleeding scores was observed in the saline-irrigated control group. Scaling and root planing, in combination with an optimal supragingival plaque control, resulted in a marked resolution of the clinical symptoms of periodontal disease. Adjunctive irrigation with chlorhexidine or hydrogen peroxide did not improve the healing result above and beyond that obtained after mechanical debridement alone or in combination with saline irrigation. Hence, the study failed to demonstrate that professionally performed periodic subgingival irrigation with chlorhexidine or hydrogen peroxide, used alone or in combination with thorough mechanical debridement, has a significant therapeutic effect.     

Effect of sampling strategy on the false-negative rate for detection of selected subgingival species

Subgingival plaque samples were obtained from the mesial surface of each tooth (maximum 28 samples per subject) in 62 subjects with prior evidence of destructive periodontal disease. The resulting 1596 samples were evaluated for their content of 14 selected taxa using a colony lift method and DNA probes. The present investigation compared the ability of 6 sampling strategies to detect a species known to be present in a subject as determined by the 28-site sampling procedure. On average, a species was not detected in 68% of the positive subjects if only the upper right first molar was sampled; 55% of subjects if both upper first molars were sampled; 36% of subjects if the 4 first molars were sampled; 28% of subjects if the 6 Ramfjord teeth were sampled; 60% of subjects if the deepest pocket was sampled and 25% of subjects if the 4 deepest pockets were sampled. The error rate was greatest for species that were infrequently detected in plaque samples such as Actinobacillus actinomycetemcomitans serotype b. This species was not detected in 49% of positive subjects, when the 6 Ramfjord teeth were sampled and 38% of subjects when the 4 deepest pockets were sampled. The data indicated that multiple plaque samples are needed to minimize false-negative rates.     

Repeated subgingival oxygen irrigations in untreated periodontal patients

Abstract The aim of this study was to assess the effect of repeated subgingival oxygen irrigations in previously untreated deep periodontal pockets. 112 pockets ≥4 mm were selected in 14 subjects. Probing attachment level and bleeding on probing were recorded, as well as the presence of disease-associated microorganisms within the pocket. Subsequently, the pockets were irrigated with gaseous oxygen 1 × a week during a continuous 8-week period. Irrigations with nitrogen served as control. The re-evaluation of all clinical and microbiological parameters at the end of study revealed that repeated oxygen insufflations resulted in a significant clinical improvement of the periodontal baseline conditions superior to the one found in the control.     

Periodic subgingival antimicrobial irrigation of periodontal pockets

The purpose of this study was to evaluate the microbiological effects of repeated subgingival irrigation of deep periodontal pockets as a single measure of treatment as well as combined with mechanical debridement, and to study the concomitant radiographical changes of the alveolar bone. 2-3 interproximal sites per jaw quadrant in 10 patients showing a probing depth of greater than or equal to 6 mm and bleeding on pocket probing were selected for the study. The pockets in the various quadrants were randomly assigned to professionally performed subgingival irrigation with 0.2% chlorhexidine gluconate, 3% hydrogen peroxide or saline or to non-irrigation. During a first phase of treatment, the pockets were periodically irrigated (every 2nd-3rd day during weeks 1-2 and 5-6) and no subgingival mechanical debridement was performed. During a second phase, subgingival scaling and root planning were carried out with adjunctive subgingival irrigation of the pockets. During the entire trial, the patients' plaque control was carefully supervised. Sampling of the subgingival microflora was performed before and after the first and second treatment phases and 3 months after the termination of the active treatment. Dark-field assessment and cultivation of the bacterial samples were performed. The radiographical examination was carried out at the start of each treatment phase and 3 months after the termination of phase II and the radiographs were analysed by the use of a subtraction technique. The results demonstrated that periodic subgingival antimicrobial irrigation per se had only limited and transient effects on the subgingival microflora.(ABSTRACT TRUNCATED AT 250 WORDS)     

The effect of subgingival debridement on periodontal disease parameters and the subgingival microbiota

Abstract The aim of the present investigation was to analyse the effect of subgingival scaling and root planing in subjects who prior to treatment exercised meticulous supragingival plaque control. 300 subjects were examined at baseline and after 1 and 2 years without treatment. After the year 2 examination, 62 subjects were randomly selected for therapy. They were given detailed instruction in proper self-performed toothcleaning measures and were carefully monitored during the subsequent 2 years. Following the year-4 examination, 2 quadrants, 1 maxillary and 1 mandibular in each subject, were randomly selected for additional therapy. The teeth in the selected quadrants were exposed to subgingival scaling and root planing. The subgingival therapy was repeated until a site no longer bled on gentle probing. This basic therapy was completed within a 2-month period. All subjects were re-examined after another 12-month interval. The examinations at year 4 and 5 included assessment of plaque, gingivitis, probing pocket depth and analysis of samples obtained from the subgingival microbiota at 134 selected sites. The findings from the present study demonstrated: (i) that subgingival scaling and root planing were effective in eliminating subgingival plaque and gingivitis; (ii) that professional therapy resulted in a pronounced reduction of probing depth at sites which at year 4 had a probing depth >3 mm; (iii) that in non-scaled quadrants, the extension of self-performed plaque control resulted in a continued improvement of the periodontal conditions at sites which at year 4 were < 5 mm deep.     

Distribution of selected bacterial species on intraoral surfaces

BACKGROUND/AIM: To examine the proportions of 40 bacterial species in samples from 8 oral soft tissue surfaces and saliva in systemically healthy adult subjects and to compare these microbiotas with those of supra- and subgingival plaque. METHODS: Microbial samples were taken from 8 oral soft tissue surfaces of 225 systemically healthy subjects using a "buccal brush". Saliva was taken by expectoration. Forty-four of these subjects provided additional supra- and subgingival plaque samples. Samples were individually evaluated for their content of 40 bacterial species using checkerboard DNA-DNA hybridization. The percentage of total DNA probe count was determined for each species, at each sample location and averaged across subjects. The significance of differences among the proportions of the 40 test species at different sample locations was sought in the 225 and 44 subjects separately using the Quade test and adjusted for multiple comparisons. Cluster analysis was performed using the proportions of the 40 species at the different sample locations using the minimum similarity coefficient and an average unweighted linkage sort. The proportions of each species were averaged across subjects in the resulting cluster groups and the significance of differences was tested using the t-test and ANOVA. RESULTS: Microbial profiles differed markedly among sample locations in the 225 subjects, with 34 of 40 species differing significantly. Proportions of Veillonella parvula and Prevotella melaninogenica were higher in saliva and on the lateral and dorsal surfaces of the tongue, while Streptococcus mitis and S. oralis were in significantly lower proportions in saliva and on the tongue dorsum. Cluster analysis resulted in the formation of 2 clusters with >85% similarity. Cluster 1 comprised saliva, lateral and dorsal tongue surfaces, while Cluster 2 comprised the remaining soft tissue locations. V. parvula, P. melaninogenica, Eikenella corrodens, Neisseria mucosa, Actinomyces odontolyticus, Fusobacterium periodonticum, F. nucleatum ss vincentii and Porphyromonas gingivalis were in significantly higher proportions in Cluster 1 and S. mitis, S. oralis and S. noxia were significantly higher in Cluster 2. These findings were confirmed using data from the 44 subjects providing plaque samples. The microbial profiles of supra- and subgingival plaque differed from the other sample locations, particularly in the increased proportions of the Actinomyces species. Species of different genera exhibited different proportions on the various intraoral surfaces, but even within the genus Streptococcus, there were differences in colonization patterns. S. oralis, S. mitis and S. constellatus colonized the soft tissues and saliva in higher proportions than the samples from the teeth, while the other 4 streptococcal species examined colonized the dental surfaces in proportions comparable to the soft tissue locations and saliva. CONCLUSIONS: Proportions of bacterial species differed markedly on different intraoral surfaces. The microbiota of saliva was most similar to that of the dorsal and lateral surfaces of the tongue. The microbiotas of the soft tissues resembled each other more than the microbiotas that colonized the teeth both above and below the gingival margin.     

Effects of subgingival chlorhexidine irrigation on periodontal inflammation

Abstract The purposes of this study were to investigate the effect of direct application of chlorhexidine to periodontal pockets and the practicability of patient self-therapy using a technique of subgingival irrigation. Patients received no other oral hygiene instruction. After initial assessment of parameters, patients were given scaling and polishing and then instruction in the irrigation of designated pockets with chlorhexidine or a placebo using a disposable syringe and blunt needle. During the 28-day irrigation period with chlorhexidine there was a highly significant reduction in periodontal inflammation which was maintained at levels significantly below the baseline values for a further 28-day period without irrigation. There was a deterioration in the periodontal state of those patients who had used the placebo. The irrigation technique itself caused no discernible injury in this group of routine periodontal patients. Also, staining in the chlorhexidine group was minimal. It is concluded that subgingival irrigation with chlorhexidine is effective in reducing periodontal inflammation and in controlling subgingival plaque. Intermittent treatment of this kind by the patient at home might reduce to more manageable levels the frequency of hygiene visits and the need for rigorous interdental oral hygiene.     

The subgingival periodontal microbiota of the aging mouth

Different mechanisms have been hypothesized to explain the increase in prevalence and severity of periodontitis in older adults, including shifts in the periodontal microbiota. However, the actual impact of aging on the composition of subgingival biofilms remains unclear. In the present article, we provide an overview of the composition of the subgingival biofilm in older adults and the potential effects of age on the oral microbiome. In particular, this review covers the following topics: (i) the oral microbiota of an aging mouth; (ii) the effects of age and time on the human oral microbiome; (iii) the potential impact of inflammaging and immunosenescence in the host–oral microbiota interactions; and (iv) the relationship of the aging oral microbiota and Alzheimer's disease. Finally, we present analyses of data compiled from large clinical studies that evaluated the subgingival microbiota of periodontally healthy subjects and patients with periodontitis from a wide age spectrum (20–83 years of age).     

Differences in subgingival microflora of Korean and German periodontal patients

Purpose

The goal of this study was to characterize the microbiological profile in samples of subgingival plaque taken from periodontal patients with different ethnic origin.

Methods

178 patients (n = 90 from South Korea and n = 88 from Germany; age: 45.4 ± 10.4 years) were diagnosed with severe generalized periodontitis. In all patients the deepest pocket of each quadrant was sampled for subgingival plaque. The four samples per patient were pooled and subsequently analysed with a 16s-RNA-gene probe test.

Results

Prevalence of Aggregatibacter actinomycetemcomitans was significantly higher in German patients (47.7%) compared to Korean patients (26.7%) (p < 0.01, χ²-test). For Tannerella forsythia and porphyromonas gingivalis, differences between Germans and Koreans were not as pronounced. A statistically significant difference could also be found for Treponema denticola (Germans: 95.5%, Koreans: 81.1%, p < 0.01, χ²-test).After logarithmic transformation, bacterial counts differed for all microorganisms under investigation between Germans and Koreans, even after using a General Linear Model/Analysis of Covariance (GLM/ANCOVA) to adjust for gender, age, smoking status, pocket probing depths (PPD) of sampled teeth, and gingival bleeding index (GBI).

Conclusion

Depending on their ethnic origin, the microbiological profile of pooled subgingival plaque sample seems to differ significantly between patients of Caucasian and Asian ethnic origin.