Genetic and phenotypic evaluation of Candida albicans strains isolated from subgingival biofilm of diabetic patients with chronic periodontitis.

Candida spp. are commensal microorganisms that are part of the microflora of different sites within the oral cavity. In healthy subjects, who have an unaltered immunological status, these yeasts do not cause disease. However, in immunosuppressed individuals whose condition may have been caused by diabetes mellitus, Candida spp. can express different virulence factors and may consequently become pathogenic. Studies have detected the presence of Candida spp. in periodontal sites of patients with chronic periodontitis, especially those that are immunologically compromised. However, the role of these microorganisms in the pathogenesis of periodontal disease is still unknown. The objectives of this study were: (1) to isolate and identify Candida albicans strains from subgingival sites of diabetic patients with chronic periodontitis; (2) to evaluate the following virulence factors; colony morphology, proteinase, phospholipase and hemolysin activities and cell surface hydrophobicity (CSH) under different atmospheric conditions; and (3) to determine the genetic patterns of these C. albicans isolates. Microbial samples were collected from subgingival sites and seeded on CHROMagar for subsequent identification of C. albicans by polymerase chain reaction (PCR). For the phenotypic tests, all strains of C. albicans were grown under reduced oxygen (RO) and anaerobiosis (ANA) conditions. Genotypes were defined by the identification through PCR of the transposable introns in the 25S rDNA. The results obtained relative to virulence factors were analyzed according to the atmospheric condition or genetic group, using Chi-square and Wilcoxon non-parametric tests. In this study, 128 strains were identified as C. albicans and of these, 51.6% were genotype B, 48.4% were genotype A and Genotype C was not found. Most of the strains were alpha-hemolytic in both atmospheric conditions, without a statistical difference. However, when comparing the genotypes, 46.1% of the genotype A strains were beta-hemolytic. In relation to colony morphology, 100% of the strains under ANA showed rough colonies, which were especially prevalent in genotype A isolates. In contrast, most of the colonies were smooth under RO. C. albicans strains did not produce proteinase and phospholipase activity in the total absence of oxygen. In RO, most strains had high proteinase activity and were positive by phospholipase tests (P < 0.05). Hydrophobicity was higher in anaerobiosis and was noted mainly for genotype A isolates. In conclusion, environmental oxygen concentration influenced the virulence factors of C. albicans strains isolated from subgingival sites of diabetic and periodontal patients. In addition, genotype A seems to be more virulent based on the phenotypic tests evaluated in this study.

Adhesion and invasion of Candida albicans from periodontal pockets of patients with chronic periodontitis and diabetes to gingival human fibroblasts.

The objectives of this study were to evaluate clinical isolates of Candida albicans, particularly their adhesion to and invasion of gingival human fibroblasts in culture and to measure nitric oxide concentration (NO) produced by fibroblasts in the presence of these yeasts. Sixteen strains of C. albicans isolated from patients with chronic periodontitis and diabetes mellitus type II were divided on the basis of phenotypic tests into two groups, i.e., highly or weakly hydrophobic. Primary cultures of human fibroblasts were isolated from gingival biopsies and after subsequent subcultures, the cells were seeded into culture plates and incubated for 24 h. C. albicans strains were inoculated into these plates and maintained for 2 and 4 h to assess their adhesion and invasion, respectively. The number of adherent or invasive yeasts was evaluated by assessing colony-forming units (CFU). The production of NO by fibroblasts was also quantified. The results showed that strains with high hydrophobicity had a greater ability to adhere and invade fibroblasts (p < 0.05, ANOVA and Tukey). The production of NO was higher for the most hydrophobic strains, but did not reach statistical difference with the weakly hydrophobic isolates. These data indicated that the hydrophobicity may play a role in the adhesion and invasion of C. albicans in fibroblast cultures.

Prospective study of potential sources of Streptococcus mutans transmission in nursery school children.

Transmission of Streptococcus mutans, a major dental caries pathogen, occurs mainly during the first 2.5 years of age. Children appear to acquire S. mutans mostly from their mothers, but few studies have investigated non-familial sources of S. mutans transmission. This study prospectively analysed initial S. mutans oral colonization in 119 children from nursery schools during a 1.5-year period and tracked the transmission from child to child, day-care caregiver to child and mother to child. Children were examined at baseline, when they were 5-13 months of age, and at 6-month intervals for determination of oral levels of S. mutans and development of caries lesions. Levels of S. mutans were also determined in caregivers and mothers. A total of 1392 S. mutans isolates (obtained from children, caregivers and mothers) were genotyped by arbitrarily primed PCR and chromosomal RFLP. Overall, 40.3 % of children were detectably colonized during the study, and levels of S. mutans were significantly associated with the development of caries lesions. Identical S. mutans genotypes were found in four nursery cohorts. No familial relationship existed in three of these cohorts, indicating horizontal transmission. Despite high oral levels of S. mutans identified in most of the caregivers, none of their genotypes matched those identified in the respective children. Only 50 % of children with high levels of S. mutans carried genotypes identified in their mothers. The results support previous evidence indicating that non-familial sources of S. mutans transmission exist, and indicate that this bacterium may be transmitted horizontally between children during the initial phases of S. mutans colonization in nursery environments.

Th1 biased response to a novel Porphyromonas gingivalis protein aggravates bone resorption caused by this oral pathogen.

In previous studies we showed that biasing the immune response to Porphyromonas gingivalis antigens to the Th1 phenotype increases inflammatory bone resorption caused by this organism. Using a T cell screening strategy we identified eight P. gingivalis genes coding for proteins that appear to be involved in T-helper cell responses. In the present study, we characterized the protein encoded by the PG_1841 gene and evaluated its relevance in the bone resorption caused by P. gingivalis because subcutaneous infection of mice with this organism resulted in the induction of Th1 biased response to the recombinant PG1841 antigen molecule. Using an immunization regime that strongly biases toward the Th1 phenotype followed by challenge with P. gingivalis in dental pulp tissue, we demonstrate that mice pre-immunized with rPG1841 developed severe bone loss compared with control immunized mice. Pre-immunization of mice with the antigen using a Th2 biasing regime resulted in no exacerbation of the disease. These results support the notion that selected antigens of P. gingivalis are involved in a biased Th1 host response that leads to the severe bone loss caused by this oral pathogen.

Lonchocarpus sericeus lectin decreases leukocyte migration and mechanical hypernociception by inhibiting cytokine and chemokines production.

In this study, we tested the potential use of a lectin from Lonchocarpus sericeus seeds (LSL), to control neutrophil migration and inflammatory hypernociception (decrease of nociceptive threshold). Pretreatment of the animals intravenously (15 min before) with LSL inhibited neutrophil migration to the peritoneal cavity in a dose-dependent fashion confirmed by an inhibition of rolling and adhesion of leukocytes by intravital microscopy. We also tested the ability of the pretreatment with LSL to inhibit neutrophil migration on immunised mice, and it was observed that a strong inhibition of neutrophil migration induced by ovoalbumin in immunized mice. Another set of experiments showed that pretreatment of the animals with LSL, inhibited the mechanical hypernociception in mice induced by the i.pl. injection of OVA in immunized mice and of carrageenan in naïve mice, but not that induced by prostaglandin E(2) (PGE(2)) or formalin. This anti-nociceptive effect correlated with an effective blockade of neutrophil influx, as assessed by the hind paw tissue myeloperoxidase levels. In addition, we measured cytokines (TNF-alpha and IL-1beta) and chemokines (MIP-1alpha [CCL3] and KC [CXCL1]) from the peritoneal exudates and i.pl. tissue. Animals treated with LSL showed inhibition of cytokines and chemokines release in a dose-dependent manner. In conclusion, we demonstrated that the inhibitory effects of LSL on neutrophil migration and mechanical inflammatory hypernocicepetion are associated with the inhibition of the production of cytokines and chemokines.

Inhibition of bacterial adherence to saliva-coated through plant lectins.

In the present study, we evaluated the ability of lectin from Talisia esculenta (TEL) and a protein from Labramia bojeri seeds (Labramin) to inhibit adherence of microorganisms and exert antimicrobial effects. The minimum inhibitory and bactericidal concentrations of these proteins were determined using 5 species of bacteria: Streptococcus mutans UA159, Streptococcus sobrinus 6715, Streptococcus sanguinis ATCC10556, Streptococcus mitis ATCC903 and Streptococcus oralis PB182. In addition, an adherence assay was performed using these 5 bacterial species and sterile polystyrene microtiter plates coated with human saliva. Filtered protein solutions (6.25 to 100 mug/ml) were added to saliva-coated plates, and the plates were then incubated for 1 h at 37 degrees C. After incubation, the plates were washed, and a bacterial suspension (10(6 )CFU/ml) was then transferred to each plate, followed by incubation at 37 degrees C for 1 h (10% CO(2)). Adherence of bacteria to the acquired pellicle was visualized by staining with crystal violet, and absorbance was measured using a plate reader at 575 nm. Neither Labramin nor TEL, at any of the concentrations used, inhibited growth of any of the microorganisms. However, Labramin inhibited adherence of S. mutans and S. sobrinus. The present results indicate that Labramin is potentially useful as a biofilm-inhibiting drug.

Increase in probing depth is correlated with a higher number of Prevotella intermedia genotypes.

BACKGROUND: The aims of this study were to determine the genotypic diversity of Prevotella intermedia in subgingival plaque samples by using two techniques, arbitrarily primed polymerase chain reaction (AP-PCR) and heteroduplex analysis, and to assess the relationship of this diversity with increase in probing depth. METHODS: The subgingival plaque samples were obtained from 12 patients using paper points inserted into periodontal pockets (diseased sites) and healthy gingival sulci (healthy sites) of the same subjects. After isolation and identification, AP-PCR was performed for genotypic characterization of P. intermedia (80 isolates). The clinical samples with a positive result for P. intermedia were amplified by 16S rRNA-based PCR method, and the amplicons were subjected to heteroduplex analysis. RESULTS: The agreement between the two methods was very high; the AP-PCR and heteroduplex analysis showed that subjects harbored between one and five distinct genotypes of P. intermedia, with a positive association between numbers of genotypes by AP-PCR (P = 0.0042) or heteroduplex (P = 0.0099) and increase in probing depth. No matching of P. intermedia genotypes was observed between healthy and diseased sites of the same individual. Interindividual analyses demonstrated absence of identical clones and indicated a high level of genetic diversity in the species. CONCLUSION: A clear relationship was observed between a higher number of genotypes and increase in probing depth; these results suggest that environmental challenges in the periodontal pockets may modulate the microbiota by selecting genotypes best able to exploit the environment.

Frequency of four different mutacin genes in Streptococcus mutans genotypes isolated from caries-free and caries-active individuals.

The ability of Streptococcus mutans to produce mutacins, combined with the production of other virulence factors such as lactic acid, may contribute to the pathogenesis of this bacterium. In the present study, the detection of genes encoding mutacin types I/III, II and IV was performed by PCR with specific primers to each type in a total of 63 S. mutans genotypes isolated from caries-active and caries-free individuals. In the caries-free group, PCR screening for mutacin IV revealed that 31.8% of strains were positive for this mutacin. PCR for the other three mutacins tested (I/III and II) did not yield amplicons in any S. mutans strains in this group. The PCR with primers of mutacin IV showed 68.3% positive genotypes in the caries-active group, on the other hand, the amplicons of mutacins I/III revealed 41.5% positive strains that carried these genes. The chi square test showed significant differences in the number of positive strains to mutacin IV when comparing the caries-free and caries-active genotypes of S. mutans (P = 0.01). All tested S. mutans strains were negative by PCR for mutacin II. The low frequencies of detection of some mutacin genes suggest the existence of high diversity and polymorphism in the production of genetic determinants of mutacin-like substances. In addition, the production of a wide spectrum of mutacins can play an important biological role in colonization by S. mutans strains, mainly in the niche of high-complexity microbial communities.

Chemical or microbiological models of secondary caries development around different dental restorative materials.

This study evaluated artificial secondary caries around restorative materials, induced by means of chemical or microbiological models. The following materials were used randomly to restore 130 dental blocks: (1) zinc-oxide eugenol-free temporary filling: Coltosol (Coltène/Whaledent Inc.; n = 30), (2) silver amalgam: Permite C (SDI Limited, n = 20), (3) composite resin: Filtek Z250 (3M ESPE; n = 20), (4) glass-ionomer cement: Fuji II (GC America Inc.; n = 20), (5) resin-modified glass ionomer: Vitremer (3M ESPE; n = 20), and (6) polyacid modified resin: Dyract AP (Dentsply; n = 20). Ten specimens of Group 1 were kept in humidity, and had no carious formation (NC). Ten specimens of each group were submitted to pH cycling (CG, n = 60), and the others were immersed in a medium containing Streptococcus mutans and sucrose (BG, n = 60). Mineral content was determined by microhardness assessment, and lesion depth was measured in polarized light photomicrographs. In the chemical model (CG), mineral content values in the vicinities of restoration were high for Groups 5 (75.7 +/- 11.9), 4 (70.8 +/- 14.2), and NC (95.4 +/- 3.8); intermediate for Groups 1 (55.8 +/- 18.5), 6 (45.6 +/- 11.0), and 2 (44.3 +/- 11.2); and reduced for Group 3 (34.7 +/- 9.7). In the microbiological model (BG), results were similar to CG, although there was less demineralization. The highest lesion depths were found for Groups 3 (182.3 +/- 33.2) in CG and 6 (126.5 +/- 42.8) in BG, when compared to Group 5 (114.6 +/- 26.0 and 56.2 +/- 33.2, respectively). In both models of caries induction, ionomeric materials showed a superior cariostatic effect when compared to the other restorative materials.

In vitro evaluation of caries inhibition promoted by self-etching adhesive systems containing antibacterial agents.

This study evaluated the cariostatic effect of antibacterial self-etching adhesive systems, by means of an in vitro bacterial caries model. Seventy-five prepared bovine slabs were randomly divided into groups (n=15): (1) unbonded composite, no carious challenge (UNB-NC); (2) unbonded composite, carious challenge (UNB-C); (3) Clearfil SE Bond, no antibacterial agent (CSE); (4) Protect Bond, containing MDPB and fluoride (PB); and (5) Reactmer Bond, fluoride-releasing (RB). All preparations were restored with Filtek Z-250. Groups (2)-(5) were submitted to a medium containing Streptococcus mutans (ATCC-- 25175) for 5 days, and Group (1) was kept in a noninoculated medium. Insoluble polysaccharides present in tooth biofilms were quantified, Knoop hardness (KHN) was measured on the enamel adjacent to restorations, and standard 35-mm polarized light photomicrographs were taken as illustrations. Polysaccharide and Knoop hardness results were analyzed with the use of ANOVA, with a split-split-plot statistical design for KHN. Except for Group (1), all groups showed similar caries formation. Biofilm over PB restorations showed the smallest amounts of polysaccharides (14.37 microg/mg), and CSE showed the highest amounts (20.87 microg/mg). All self-etching systems tested were unable to inhibit secondary caries in a bacterial model simulating a high caries challenge, even though there was reduced glucan synthesis provided by the adhesive system containing MDPB and fluoride.

Detection of Tannerella forsythensis (Bacteroides forsythus) and porphyromonas gingivalis by polymerase chain reaction in subjects with different periodontal status.

BACKGROUND: The aim of this study was to determine the prevalence of Tannerella forsythensis (formerly Bacteroides forsythus) and Porphyromonas gingivalis in subgingival plaque samples by using polymerase chain reaction (PCR), and to assess the relationship of these bacteria with different categories of periodontal disease and health. METHODS: Subjects were distributed into 3 groups according to their periodontal diagnosis: group 1, periodontally healthy (N = 10); group 2, periodontitis with probing depth < or = 5 mm (N = 10); group 3, periodontitis with probing depth > 5 mm (N = 10). The subjects in groups 2 and 3 had healthy and diseased periodontal sites. Subgingival plaque samples were obtained using paper points inserted into periodontal pockets (diseased sites) and into healthy gingival sulci (healthy sites) of the same subject. RESULTS: The distribution of bacteria differed in healthy and diseased sites. T. forsythensis (B. forsythus) was not detected in any sample from healthy sites in any group but was detected in 70% and 100% of diseased sites in groups 2 and 3, respectively. P. gingivalis was detected in only one sample from a healthy site (group 2), and in the diseased sites, its prevalence was 40% (group 2) and 90% (group 3). In addition, T. forsythensis (B. forsythus) and P. gingivalis were both detected in 30% and 90% of the diseased sites in groups 2 and 3, respectively. CONCLUSION: These results indicate a possible association between periodontal disease and the presence of T. forsythensis (B. forsythus) and/or P. gingivalis.

Microbial diversity similarities in periodontal pockets and atheromatous plaques of cardiovascular disease patients.

BACKGROUND AND OBJECTIVE: The immune and infectious alterations occurring in periodontitis have been shown to alter the development and severity of cardiovascular disease. One of these relationships is the translocation of oral bacteria to atheroma plaques, thereby promoting plaque development. Thus, the aim of this study was to assess, by 16s cloning and sequencing, the microbial diversity of the subgingival environment and atheroma plaques of patients concomitantly suffering from periodontitis and obstructive coronary artery atherosclerosis (OCAA). METHODS: Subgingival biofilm and coronary balloons used in percutaneous transluminal coronary angioplasty were collected from 18 subjects presenting with generalized moderate to severe periodontitis and OCAA. DNA was extracted and the gene 16S was amplified, cloned and sequenced. RESULTS: Significant differences in microbial diversity were observed between both environments. While subgingival samples mostly contained the phylum Firmicutes, in coronary balloons, Proteobacteria (p<0.05) was predominant. In addition, the most commonly detected genera in coronary balloons were Acinetobacter, Alloprevotella, Pseudomonas, Enterobacter, Sphingomonas and Moraxella, while in subgingival samples Porphyromonas, Filifactor, Veillonella, Aggregatibacter and Treponema (p<0.05) were found. Interestingly, 17 identical phylotypes were found in atheroma and subgingival samples, indicating possible bacterial translocation between periodontal pockets and coronary arteries. CONCLUSION: Periodontal pockets and atheromatous plaques of cardiovascular disease patients can present similarities in the microbial diversity.

The combination of amoxicillin and metronidazole improves clinical and microbiologic results of one-stage, full-mouth, ultrasonic debridement in aggressive periodontitis treatment.

BACKGROUND: The aim of the present study is to assess clinical, microbiologic, and immunologic benefits of amoxicillin/metronidazole (AM) when performing full-mouth ultrasonic debridement (FMUD) in generalized aggressive periodontitis (GAgP) treatment. METHODS: Twenty-four GAgP patients were divided into two groups: the FMUD group (n = 12), which received FMUD plus placebo, and the FMUD+AM group (n = 12), which received FMUD and 375 mg amoxicillin plus 250 mg metronidazole for 7 days. The following clinical outcomes were tested: plaque and bleeding on probing indices, pocket probing depth (PD), relative gingival margin position (GMP), and relative clinical attachment level (CAL). Total amount of Porphyromonas gingivalis (Pg), Aggregatibacter actinomycetemcomitans (Aa), Tannerella forsythia (Tf), and gingival crevicular fluid (GCF) concentration of interleukin (IL)-10 and IL-1ß were also determined. All clinical, microbiologic, and immunologic parameters were assessed at baseline and at 3 and 6 months post-therapy. The ANOVA/Tukey test was used for statistical analysis (α = 5%). RESULTS: Amoxicillin/metronidazole used as an adjunct to the FMUD protocol added clinical and microbiologic benefits to GAgP treatment (P <0.05). FMUD+AM groups presented an additional PD reduction in initially deep PDs at the 3-month follow-up (3.99 ± 1.16 mm and 3.09 ± 0.78 mm for FMUD+AM and FMUD, respectively; P <0.05), a lower number of residual pockets at the 3- and 6-month follow-ups, and a statistical reduction in amounts of Aa (P <0.05). Analysis of Tf and Pg amounts, as well as IL-10 and IL-1ß GCF concentrations failed to demonstrate a difference between the groups (P >0.05). CONCLUSION: It may be concluded that amoxicillin/metronidazole improves clinical and microbiologic results of FMUD in GAgP treatment.

Impact of supragingival therapy on subgingival microbial profile in smokers versus non-smokers with severe chronic periodontitis.

BACKGROUND: The aim of this study was to assess subgingival microbiological changes in smokers versus non-smokers presenting severe chronic periodontitis after supragingival periodontal therapy (ST). METHODS: Non-smokers (n=10) and smokers (n=10) presenting at least nine teeth with probing pocket depth (PPD) (≥5 mm), bleeding on probing (BoP), and no history of periodontal treatment in the last 6 months were selected. Clinical parameters assessed were plaque index (PI), BoP, PPD, relative gingival margin position (rGMP) and relative clinical attachment level (rCAL). Subgingival biofilm was collected before and 21 days after ST. DNA was extracted and the 16S rRNA gene was amplified with the universal primer pair, 27F and 1492R. Amplified genes were cloned, sequenced, and identified by comparison with known 16S rRNA sequences. Statistical analysis was performed by Student's t and Chi-Square tests (α=5%). RESULTS: Clinically, ST promoted a significant reduction in PI and PPD, and gain of rCAL for both groups, with no significant intergroup difference. Microbiologically, at baseline, data analysis demonstrated that smokers harbored a higher proportion of Porphyromonas endodontalis, Bacteroidetes sp., Fusobacterium sp. and Tannerella forsythia and a lower number of cultivated phylotypes (p<0.05). Furthermore, non-smokers featured significant reductions in key phylotypes associated with periodontitis, whereas smokers presented more modest changes. CONCLUSION: Within the limits of the present study, ST promoted comparable clinical improvements in smokers and non-smokers with severe chronic periodontitis. However, in smokers, ST only slightly affected the subgingival biofilm biodiversity, as compared with non-smokers.

Periodontal conditions and prevalence of putative periodontopathogens and Candida spp. in insulin-dependent type 2 diabetic and non-diabetic patients with chronic periodontitis--a pilot study.

OBJECTIVES: The aims of this study were to evaluate periodontal conditions and identify the presence of Aggregatibacter actinomycetemcomitans, Porphyromonas gingivalis and Tannerella forsythia, and four different species of Candida (C. albicans, C. dubliniensis, C. glabrata and C. tropicalis) in periodontal pockets and furcation sites of insulin-dependent type 2 diabetic and non-diabetic patients with generalised chronic periodontitis. DESIGN: Clinical parameters, including oral status assessed using plaque index, gingival index, probing depth, gingival recession and clinical attachment level and systemic conditions with fasting glucose level or glycosylated haemoglobin were measured in diabetic and non-diabetic patients with chronic periodontitis. Samples of subgingival biofilm were obtained from the periodontal pockets and furcation sites and submitted to phenol-chloroform DNA extraction and PCR analysis using specific primers. RESULTS: Clinical conditions of diabetic and non-diabetic patients were similar, without statistical differences in both periodontal indexes and glucose levels (p>0.05). Diabetics had a higher prevalence of Candida spp., mainly C. albicans and C. dubliniensis, and a lower frequency of T. forsythia, when compared to non-diabetic patients, for both periodontal sites. C. glabrata and C. tropicalis were not found in periodontal pockets and furcation sites of non-diabetic patients. CONCLUSION: The results demonstrated a strong colonisation of Candida spp. in the periodontal sites of diabetic patients that have generalised chronic periodontitis with a higher prevalence of C. dubliniensis followed by C. albicans.

Candida spp. in periodontal disease: a brief review.

Although the main reservoir of Candida spp. is believed to be the buccal mucosa, these microorganisms can coaggregate with bacteria in subgingival biofilm and adhere to epithelial cells. Such interactions are associated with the capacity of Candida spp. to invade gingival conjunctive tissue, and may be important in the microbial colonization that contributes to progression of oral alterations caused by diabetes mellitus, some medications, and immunosuppressive diseases such as AIDS. In addition, immune deficiency can result in proliferation of Candida spp. and germination of forms that are more virulent and have a higher capacity to adhere to and penetrate cells in host tissues. The virulence factors of Candida spp. increase host susceptibility to proliferation of these microorganisms and are likely to be important in the study of periodontal disease. Herein, we briefly review the literature pertaining to the role of Candida spp. in periodontal disease, and consider the main virulence factors, the host immune response to these microorganisms, and the effect of concomitant immunosuppressive conditions.

Halistanol sulfate A and rodriguesines A and B are antimicrobial and antibiofilm agents against the cariogenic bacterium Streptococcus mutans

In the present investigation we report the antibacterial activity of halistanol sulfate A isolated from the sponge Petromica ciocalyptoides, as well as of rodriguesines A and B isolated from the ascidian Didemnum sp., against the caries etiologic agent Streptococcus mutans. The transcription levels of S. mutans virulence genes gtfB, gtfC and gbpB, as well as of housekeeping genes groEL and 16S, were evaluated by sqRT-PCR analysis of S. mutans planktonic cells. There were no alterations in the expression levels of groEL and 16S after antimicrobial treatment with halistanol sulfate A and with rodriguesines A and B, but the expression of the genes gtfB, gtfC and gbpB was down-regulated. Halistanol sulfate A displayed the most potent antimicrobial effect against S. mutans, with inhibition of biofilm formation and reduction of biofilm-associated gene expression in planktonic cells. Halistanol sulfate A also inhibited the initial oral bacteria colonizers, such as Streptococcus sanguinis, but at much higher concentrations. The results obtained indicate that halistanol sulfate A may be considered a potential scaffold for drug development in Streptococcus mutans antibiofilm therapy, the main etiologic agent of human dental caries. .

PPAR-gamma agonist rosiglitazone prevents inflammatory periodontal bone loss by inhibiting osteoclastogenesis.

Rosiglitazone (RGZ), an oral anti-hyperglycemic agent used for non-insulin-dependent diabetes mellitus, is a high-affinity synthetic agonist for peroxisome proliferator-activated receptor-gamma (PPAR-gamma). Both in vitro and in vivo experiments have also revealed that RGZ possesses anti-inflammatory properties. Therefore, in the present study, we investigated the anti-inflammatory effects of RGZ in a rat model of periodontal disease induced by ligature placed around the mandible first molars of each animal. Male Wister rats were divided into four groups: 1) animals without ligature placement receiving administration of empty vehicle (control); 2) animals with ligature receiving administration of empty vehicle; 3) animals with ligature receiving administration with oral RGZ (10 mg/kg/day); and 4) animals with ligature receiving administration of subcutaneous RGZ (10 mg/kg/day). Thirty days after induction of periodontal disease, the animals were sacrificed, and mandibles and gingival tissues were removed for further analysis. An in vitro assay was also employed to test the inhibitory effects of RGZ on osteoclastogenesis. Histomorphological and immunohistochemical analyses of periodontal tissue demonstrated that RGZ-treated animals presented decreased bone resorption, along with reduced RANKL expression, compared to those animals with ligature, but treated with empty vehicle. Corresponding to such results obtained from in vivo experiments, RGZ also suppressed in vitro osteoclast differentiation in the presence of RANKL in MOCP-5 osteoclast precursor cells, along with the down-regulation of the expression of RANKL-induced TRAP mRNA. These data indicated that RGZ may suppress the bone resorption by inhibiting RANKL-mediated osteoclastogenesis elicited during the course of experimental periodontitis in rats.

Genetic diversity and exoenzyme activities of Candida albicans and Candida dubliniensis isolated from the oral cavity of Brazilian periodontal patients.

OBJECTIVE: Mucosal surfaces are the primary oral reservoirs of Candida species, but these species can also be found in subgingival biofilm. The present study investigated the genetic diversity and production of exoenzymes of C. albicans and C. dubliniensis isolated from the oral cavity of systemically healthy patients with periodontitis. DESIGN: Fifty-three patients were analysed. Samples were collected from three oral cavity sites (periodontal pocket, gingival sulci and oral mucosa), plated and, after isolation, suspect strains of C. albicans and C. dubliniensis were identified by PCR. The genetic diversity of the isolates was evaluated by RAPD and the activities of the secreted aspartyl proteinases and phospholipases were evaluated by the agar plate method. RESULTS: Twenty-one patients showed positive results for Candida spp. There were no statistically significant differences between genders, or between sites. C. albicans was the most frequently found specie, while C. dubliniensis was isolated from the periodontal pocket of only one patient. Sixteen genotypes were detected among the C. albicans isolates, and one among the C. dubliniensis isolates. The similarity coefficient (S(SM)) values among the C. albicans genotypes ranged from 0.684 to 1.0 with an average of 0.905+/-0.074. All isolates produced high levels of Saps and most of them produced high levels of phospholipases. No relationship was found between the genotypes and the pattern of enzymatic production. There was no association between specific genotypes and their site of isolation. CONCLUSIONS: The results of the present study suggest that genetically homogeneous strains of C. albicans are present in the oral cavity of patients with periodontitis and that these strains are capable of producing high levels of exoenzyme.

Th1 immune response promotes severe bone resorption caused by Porphyromonas gingivalis.

Bacterial infections of the dental pulp result in soft tissue and alveolar bone destruction. It has been suggested that Th1 responses promote disease, whereas Th2 responses are protective. However, other studies have challenged this notion. To address this question, bone destruction was evaluated in mice immunized to develop strong and polarized Th1- or Th2-biased responses to the oral pathogen Porphyromonas gingivalis. Th1 bias was confirmed by the presence of high titers of serum IgG2a and the production of high levels of interferon (IFN)-gamma and no interleukin (IL)-4 by lymph node cells stimulated with P. gingivalis antigens. In contrast, Th2-biased animals had high titer IgG1 and no IgG2a, and their lymph node cells produced high levels of IL-4 but no IFN-gamma. Subsequent infection of the dental pulp with P. gingivalis caused extensive inflammation and alveolar bone destruction in Th1-biased mice, whereas Th2-biased mice and controls developed minimal lesions. Inflammatory granulomas in Th1-biased mice were heavily infiltrated with osteoclasts and had high local expression of IFN-gamma, IL-1alpha, and IL-1beta. Little or no IFN-gamma/IL-1alpha/IL-1beta and no obvious osteoclasts were detected in lesions of Th2-biased and control groups. These results directly demonstrate that specific Th1 responses promote severe infection-stimulated alveolar bone loss.