Microbiological profile of early onset/aggressive periodontitis patients.

OBJECTIVES: The objectives of this study were to characterize the bacterial profile and to seek possible bacterial associations in the subgingival microbiota of early onset periodontitis/aggressive periodontitis patients by using two different techniques, culture and immunofluorescence. MATERIAL AND METHODS: The study group consisted of 66 systemically healthy individuals with evidence of early onset periodontitis - 41 females and 25 males aged 23-35 years (mean 31.1 +/- 3.1 years). Bacterial samples were collected from the deepest site in each quadrant, resulting in a total of 264 sites with a mean probing pocket depth of 6.6 +/- 1.5 mm. Samples were cultured anaerobically and in 10% CO(2) using selective and nonselective media, and isolates were characterized to species level. Indirect immunofluorescence using monoclonal antibodies was applied to detect Actinobacillus actinomycetemcomitans, Porphyromonas gingivalis, Tannerella forsythia (Bacteroides forsythus, Tannerella forsythensis), Prevotella intermedia/Prevotella nigrescens, Campylobacter rectus, Peptostreptococcus micros and Actinomyces israelii. RESULTS: 93.6% of sampled sites showed bleeding on probing and 23.5% were positive for suppuration. P. intermedia/P. nigrescens, P. gingivalis, and C. rectus were detected in 77.3-85.9% of samples using culture methods and in 85.6-91.3% using immunofluorescence. P. micros and A. actinomycetemcomitans were found, respectively, in 63.3% and 25.0% of all sites using culturing and in 58.7% and 27.7% sites using immunofluorescence. Significantly strong positive associations were observed between T. forsythia and C. rectus (odds ratio 109.46), and T. forsythia and P. gingivalis (odd ratio 90.26), whereas a negative association was seen between P. intermedia/P. nigrescens and A. actinomycetemcomitans (odds ratio 0.42). Coinfection by P. gingivalis, T. forsythia, P. intermedia/P. nigrescens and C. rectus was observed in 62.1% of the test sites, and in 89.4% of the studied subjects. The sensitivity of immunofluorescence for T. forsythia, C. rectus, P. intermedia/P. nigrescens and P. gingivalis was found to be very high (0.99-0.94) using culture as the reference detection method. The agreement between culture and immunofluorescence in detecting the presence or absence of the investigated species was 85.2-88.1% for P. gingivalis, P. intermedia/P. nigrescens, C. rectus, and T. forsythia, 75.9% for A. actinomycetemcomitans and 70.4% for P. micros. CONCLUSIONS: The microbial profile of the early onset/aggressive periodontitis population was complex. The agreement between the two detection methods was very high.

Anti-plaque agents in the prevention of biofilm-associated oral diseases.

The prevention of dental caries and periodontal diseases is targeted at the control of dental plaque. In this context, chemical agents could represent a valuable complement to mechanical plaque control. The active agents should prevent biofilm formation without affecting the biological equilibrium within the oral cavity. Depending on the goals of the preventive measures, various strategies may be considered. Anti-plaque agents with properties other than bactericidal or bacteriostatic activities may be used in primary prevention. In this approach, a modest antiplaque effect may be sufficient or even desirable, as it would decrease the side effects of the active agent. Antimicrobial agents are best indicated in secondary and tertiary prevention, as the objectives are to restore health and to prevent disease recurrence. The rational is to prevent or delay subgingival recolonization by pathogenic micro-organisms. The development of in vitro oral biofilm models certainly represents a major advance for studying and testing oral anti-plaque agents in recent years. The results of these studies have shown that chlorhexidine, hexetidine, delmopinol, amine fluoride/stannous fluoride, triclosan, phenolic compounds, among others, may inhibit biofilm development and maturation as well as affect bacterial metabolism.

Porphyromonas gingivalis gingipain-R enhances interleukin-8 but decreases gamma interferon-inducible protein 10 production by human gingival fibroblasts in response to T-cell contact.

Proteases produced by Porphyromonas gingivalis, an oral pathogen, are considered important virulence factors and may affect the responses of cells equipped with proteinase-activated receptors. The aim of this study was to investigate the effect of the arginine-specific cysteine protease gingipain-R produced by P. gingivalis on chemokine production by human gingival fibroblasts (HGF) and the effect of gingipain-R treatment on the subsequent contact-dependent activation of HGF by T cells. HGF incubated in the presence of purified 47-kDa gingipain-R showed increased levels of interleukin-8 (IL-8) mRNA. Cyclooxygenase-2 (COX-2) mRNA was also induced. Further exposure of HGF to activated T cells resulted in the dose- and time-dependent enhancement of IL-8 transcription and release. T-cell membrane-bound tumor necrosis factor (TNF) was the ligand inducing IL-8 production by HGF, since TNF neutralization abrogated HGF responses to T-cell contact. The enhanced IL-8 release was due, at least in part, to prostaglandin-E(2) production, which was mostly blocked by indomethacin. Gingipain-R proteolytic activity was required since heat inactivation, specific synthetic protease inhibitors, and the natural substrate competitor histatin 5 abrogated its effects. The enhanced production of IL-8 in response to T-cell contact was specific since monocyte chemotactic protein-1 (MCP-1) production was unaffected while interferon-gamma-inducible protein-10 (IP-10) was inhibited. The sum of these activities may result in the recruitment of differential cell types to sites of inflammation since IL-8 preferentially recruits neutrophils and IP-10 attracts activated T cells and may be relevant to the pathogenesis of periodontitis.

Clinical and microbiological characteristics of smokers with early onset periodontitis.

Cigarette smoking is a potential risk factor which has recently been associated with periodontal disease progression. The objective of this study was to compare the microbial profile of smokers and non-smokers in a group of patients with early onset periodontitis. The study population consisted of 60 healthy individuals, 40 males and 20 females aged 22 to 35 yr, exhibiting early onset periodontitis. Thirty patients were smokers (30.9 cigarettes/d) and 30 non-smokers. Smokers had a higher proportion of deep pockets (PD >5 mm), especially in the maxilla anterior and premolar regions (p < 0.001) and presented a significantly greater mean probing depth and attachment loss (p <0.05) in diseased sites and a significantly greater alveolar bone loss (p <0.01) compared to non-smokers. Two pooled bacterial samples were obtained from each patient. Samples were collected from the deepest periodontal pockets of each quadrant. The samples were cultured anaerobically and in 10% CO2 plus air for bacterial isolation using selective and non-selective media. Isolates were characterized to species level by conventional biochemical tests and various identification kits. Smokers harboured a greater number of bacteria in total. Analysis of bacterial counts using the ANOVA (Mann-Whitney U-test) showed that Staphylococcus aureus, Peptostreptococcus micros, Campylobacter concisus, Escherichia coli, Bacteroides forsythus, C. gracilis, C. rectus, Porphyromonas gingivalis, Selenomonas sputigena, Candida albicans and Aspergillus fumigatus were found in significantly higher numbers and more frequently in smokers while Streptococcus intermedius, A. naeslundii, A. israelii and Eubacterium lentum were detected more frequently and in significantly higher proportions in non-smokers. The isolation of bacteria belonging to the exogenous flora such as E. coli, C. albicans, A. fumigatus and S. aureus in smokers' microbiota underscores the importance of the host that is adversely affected by cigarette smoking.

Scanning electron microscopic study of the effects of Er:YAG laser on root cementum.

Use of Er:YAG laser has been proposed for the removal of microbial deposits and calculus present on teeth affected by periodontal disease. However, the influence of Er:YAG laser irradiation on root surfaces has not yet been fully investigated. The aim of the present study was to evaluate the effects of Er:YAG laser irradiation on root cementum by scanning electron microscopy (SEM). Specimens were obtained from extracted human periodontally-diseased teeth using a water-cooled high-speed bur. An Er:YAG laser beam was then applied at various powers ranging from 25 to 100 mJ/ pulse/sec. The laser irradiation was performed under water irrigation, with the tip held perpendicular to the root surface in the contact mode. Following laser exposure, specimens were fixed, dehydrated, and dried at critical-point in liquid CO2. After mounting on SEM plates and sputter-coating with gold, the cementum surface was examined by SEM. Observations of the root surface showed a relatively flat surface in control specimens. In Er:YAG exposed specimens, the laser beam created a circular, notched-edge, crater-like defect on the root. The bottom of the lesion showed an irregular and sharp-pointed surface. Subsequently, the specimens were fractured with a sharp scalpel perpendicularly to the surface. SEM observations of these specimens showed a 15 microm layer of damaged tissue within the laser-irradiated cementum. The tissue presented an amorphous appearance and the Sharpey's and matrix fiber bundles were not clearly distinguishable. These observations indicate that cementum tissue could be damaged by Er:YAG laser irradiation.

Current status in oral fluoride pharmacokinetics and implications for the prophylaxis against dental caries.

Fluoride plays a central role in the prevention of dental caries. There is evidence that its effect is mainly topical and that continuous presence of fluoride ions in low concentration at the plaque/enamel interface is essential. The present paper reviews the most important aspects of fluoride kinetics in the oral cavity and discusses their implications on preventive approaches to dental caries. As a continuous presence of fluoride ions in saliva is important for an optimum prophylactic effect, new formulations capable of delivering low levels of fluoride over prolonged periods of time have been developed. These systems consist either of intra-oral devices, or of restorative materials into which fluoride has been incorporated. Among all the preparations investigated, bioadhesive tablets and membrane-controlled reservoirs are the most promising.

Effects of an oral health program on selected clinical parameters and salivary bacteria in a long-term care facility.

The aim of this study was to evaluate clinically and microbiologically the effects of a preventive oral health program in a long-term care facility. A total of 116 dentate elderly residents agreed to participate, and half of them were included in an experimental group. Almost all of the residents were mentally or physically handicapped, and many were dependent on care-givers for daily living activities. Oral examination and microbiological sampling were performed at baseline and 18 months later. The experimental group benefited from a preventive program, including an oral hygiene course for the health care providers and regular recalls by dental hygienists of the residents. After 18 months, the plaque indices were statistically similar to those at baseline in both groups. Mutans streptococci counts and active root caries at 18 months were lower compared to baseline in the experimental group but did not change significantly in the control group. Thus, it seems that, while the preventive program failed to decrease plaque indices, it was effective in reducing mutans streptococci colonisation and caries prevalence.

Serum immunoglobulin G responses to various Actinobacillus actinomycetemcomitans serotypes in a young ethnographically heterogeneous periodontitis patient group.

Sera from young patients with periodontal diseases have been shown to often contain highly elevated antibody levels to Actinobacillus actinomycetemcomitans, in particular serotype b. Such responses were reportedly predominated by antibodies of the immunoglobulin G2 (IgG2) subclass. The aim of this study was to investigate an ethnically diverse group of 14 early-onset periodontitis and 15 rapidly progressive periodontitis patients for the occurrence of elevated antibody titers against the five known A. actinomycetemcomitans serotypes, and to compare the patient's IgG subclass response profiles. Enzyme-linked immunosorbent assays were used to measure both total IgG and subclass specific IgG titers. Twenty-four subjects had markedly elevate total IgG levels against at least one serotype. The frequencies of high responses against serotypes a, b, c, d and e were 7, 11, 6, 4, and 4, respectively. Elevated antibody responses were predominated by IgG2, regardless of the serotype to which the response was directed. The serotype specificity of the host responses was further investigated by competitive binding studies with serotype-specific monoclonal antibodies. Twelve sera were found to contain antibodies capable of strongly inhibit the binding of monoclonal antibodies against a single serotype; four other sera had antibodies against epitopes of two, and one serum against those of three serotypes. The findings document broad serotype diversity in an ethnically heterogeneous group of patients and indicate that strong antibody responses to A. actinomycetemcomitans are predominated by IgG2 regardless of the serotype of the infective agent.

Supportive care of the periodontal patient.

The ultimate goal of supportive periodontal care is to maintain health of the dental and oral soft tissues. It represents a preventive measure for individuals who have never experienced periodontal problems. On the other hand, supportive care is a continuation of therapy for the treated periodontal patient, once health has been reestablished. It aims at optimizing the results of therapy and prevent further destruction following active treatment. Attempts are being made to individualize and tailor supportive periodontal care according to the patient's profile and needs. Recent trends also show increased use of antimicrobials as adjuncts to mechanical procedures for controlling the etiologic agents.

Potential of diagnostic microbiology for treatment and prognosis of dental caries and periodontal diseases.

Most evidence suggests that only a finite number of bacteria are responsible for dental caries and periodontal diseases. This knowledge led to the development of microbial tests which can identify suspected pathogens. Current evaluation of the diagnostic power of microbial tests has shown that they have a low sensitivity and a low prognostic value. Despite these shortcomings, there are valid indications for microbiological-based diagnosis. Salivary microbial tests for the detection of mutans streptococci and lactobacilli may be useful, for example, in young children, oligosialic patients, and orthodontic patients. These tests can be used to monitor the success of chemopreventive measures or compliance with dietary recommendations. Microbial diagnosis, may also be valuable in the treatment of early-onset periodontitis or in subjects who respond poorly to periodontal therapy. The use of microbial tests to monitor the efficacy of chemotherapy or mechanical treatment is of particular interest.

Treponema denticola induces actin rearrangement and detachment of human gingival fibroblasts.

Spirochetes are associated with destructive periodontal diseases, and one cultivatable oral species, Treponema denticola, binds to mammalian cells and perturbs metabolism. To evaluate the cytoskeletal responses and attachment functions of human gingival fibroblasts (HGF) exposed to T. denticola, monolayers of HGF were incubated with T. denticola strains ATCC 35405, e, and e' in serum-free medium. HGF retracted pseudopods, rounded up, and ultimately detached from the substratum. Scanning electron microscopy showed spirochetes in close contact with HGF surfaces; occasionally, bacteria were partially submerged between folds in the HGF membrane. Blebbing and numerous microvilli formed on the cell surface as the HGF retracted. By confocal microscopy, spirochetes were detected in contact with the HGF surface but were never found on the ventral surface of fibroblasts between the substratum and cell. Morphological alterations were associated with and preceded by actin assembly, as measured by microscopic fluorimetry: there was a 263% increase in actin fluorescence over controls within 30 min. Detachment of fibroblasts from the substratum was related to incubation time and was dependent on the concentration of T. denticola. Detachment was observed for all strains tested and was also dependent on the viability of T. denticola: UV light, heat, and metronidazole treatment markedly reduced the HGF detachment response. Detachment was also significantly reduced by the protease inhibitor phenylmethylsulfonyl fluoride. HGF viability was not significantly affected by coincubation with spirochetes, as measured by lactate dehydrogenase release. Thus, T. denticola induces rapid cytoskeletal remodelling followed by cell detachment, which might be stimulated by a bacterially associated protease but is not likely directly mediated by proteolytic degradation at the cell-substratum adhesive contact points.

Effects of orthodontic bands on microbiologic and clinical parameters.

The aim of this study was to evaluate the effect of placement of orthodontic bands on the gingival tissues and the microbial composition of dental plaque. Ten subjects undergoing orthodontic treatment completed the study. In each subject four sites were examined: two test sites with orthodontic bands and two control sites free of bands. Clinical and bacterial examinations were performed before the beginning of the treatment and 5, 7, 47, 72, and 90 days after placement of the orthodontic appliances. Plaque index (Pl I) and bleeding scores increased significantly on banded teeth as compared with control sites. Probing depth remained within normal values for both test and control groups. The composition of dental plaque determined by dark-field microscopy showed significant shifts in the test sites after banding. Changes consisted of an increase in the percentage of spirochetes, motile rods, filaments, and fusiforms; conversely, a decrease in cocci was noted. During the same period no significant changes in the bacterial distribution were observed in the control group.

The effect of plaque control after scaling and root planing on the subgingival microflora in human periodontitis.

The aim of the present study was to evaluate the effect of a strict supragingival plaque control regimen on bacterial repopulation following scaling and root planing. 7 patients with moderate to severe inflammatory periodontal disease received a full-mouth scaling and subgingival curettage. Using a split-mouth design, 2 sites of opposite quadrants were submitted to professional supragingival plaque control 3 X a week while the contralateral sites served as controls. Clinical and bacterial examination were performed on days 7, 14, 28, 49, 56, 63 and 70 following therapy. All clinical parameters (P1I, GI, probing depth, attachment levels) showed significant improvement after scaling and root planing. Shifts in the subgingival bacterial population observed by dark field were also noticed following curettage: there was a decrease in the proportions of spirochetes and motile rods and an increase in that of coccoid cells. However, bacterial distribution tended to return to base line values towards the end of the observation period. No difference in the pattern of bacterial recolonization of the subgingival area could be detected between the sites under strict supragingival plaque control and the control sites.

The effect of supragingival plaque control on the composition of the subgingival microflora in human periodontitis.

The aim of this study was to evaluate the effect of supragingival plaque control on the composition of the subgingival microflora. 8 subjects with moderate to severe periodontitis were chosen for the study. Sites with periodontal destruction (GI greater than 2; probing depth greater than 6.5 mm; vertical alveolar bone loss on radiographs) were submitted to professional plaque control 3 X a week for 3 weeks. Contralateral sites received no prophylaxis and served as controls. Patients maintained usual oral hygiene during the observation period: it consisted exclusively of tooth brushing once or twice a day with no use of interdental cleaning aids. Clinical examination and bacterial sampling were performed every week. At the end of the study, PlI scores for the experimental sites showed a marked diminution compared with the control sites. No variations were observed in GI or probing depth in test or control sites during the study. The composition of subgingival plaque in both groups showed no significant variations during that period.

Periodontal status associated with chronic neutropenia.

A case of chronic neutropenia in a 12-year-old boy is reported. The patient presented with severe gingival inflammation and alveolar bone loss. Immunologic analysis of the patient's serum revealed the presence of precipitating antibodies against antigenic components of Actinobacillus actinomycetemcomitans Y4 and 652. It was also found that the serum neutralized the leukotoxic activity of Actinobacillus actinomycetemcomitans Y4. The etiology and the pathogenesis of periodontal disease in neutropenic patients are discussed in view of these findings.

Serum neutralizing activity against Actinobacillus actinomycetemcomitans leukotoxin in juvenile periodontitis.

A relatively high incidence of infection by Actinobacillus actionomycetemcomitans can be shown in subgingival plaque samples obtained from patients with juvenile periodontitis. These organisms possess a potent leukotoxin(s) which rapidly destroys isolated human polymorphonuclear leukocytes (PMNs) and monocytes. If such leukotoxins operate in vivo, they could deprive the gingival crevice area of an essential antibacterial defense mechanism. We have found that sera from juvenile periodontitis patients consistently (greater than 90%) contain antibodies which neutralize Actinobacillus actinomycetemcomitans leukotoxin(s). On the other hand, sera from normal individuals or patients with other types of periodontal disease usually amplified rather than inhibited the leukotoxic reaction. Many patients with juvenile periodontitis have demonstrable defects in PMN or monocyte chemotaxis and this may place them at risk to gingival infection by Actinobacillus actinomycetemcomitans. The immune response against these organisms could be a crucial determinant in the course of juvenile periodontitis. While this disease is relatively rare, it does cause immeasurable emotional, physical and economic hardship for patients and their families. The identification of Actinobacillus actinomycetemcomitans as a potential pathogen in this disorder may eventually lead to specific forms of therapy to prevent and eliminate infection by this organism in these patients.