Microbiological evaluation of primary endodontic infections in teeth with and without sinus tract

Aim To examine the microbiological status of primary endodontic infections in teeth with and without a sinus tract. Methodology Samples were collected by means of a size 15 H‐type file and two sterile paper points from 30 cases of primary endodontic infections with (n = 15) or without (n = 15) a sinus tract. The presence of 40 bacterial species was determined by the checkerboard DNA‐DNA hybridization method. Results The species found at the highest levels and prevalence were Fusobacterium nucleatum sp. vincentii, Porphyromonas gingivalis, Veillonella parvula, Enterococcus faecalis, Campylobacter gracilis and Neisseria mucosa. Total bacterial counts were similar between teeth with (44 × 10⁵) and without (50 × 10⁵) a sinus tract (t‐test: P > 0.05). E. faecalis, Streptococcus anginosus, Capnocytophaga sputigena and Capnocytophaga gingivalis had significantly higher counts in the absence of sinus tract (Mann–Whitney test, P < 0.05). Higher levels of P. gingivalis and Fusobacterium nucleatum sp. nucleatum were observed in cases with a sinus tract. Leptotrichia buccalis (OR = 1.83; CI 95%) and Porphyromonas endodontalis (OR = 2.15; CI 95%) were associated with an increased chance of subjects having a sinus tract. Conclusions Primary endodontic infections were associated with a large variety of bacterial species. Specific differences between the composition of the microbiota of primary root canal infections were observed in cases with or without a sinus tract.     

Comparative analysis of endodontic pathogens using checkerboard hybridization in relation to culture

Background/Aim:  The purpose of this study was to detect bacterial species and to quantify the total number of bacteria from samples of infected root canals before (S1) and after chemo-mechanical preparation using 2% chlorhexidine (CHX) gel as auxiliary chemical substance (S2) and after 7 days of intracanal dressing (S3) to compare microbial changes.
Method:  Twenty-four teeth were selected for this study. Chemo-mechanical preparation was performed using 2% CHX gel, then three different intracanal medicaments [M1: Ca(OH)₂ paste; M2: 2% CHX gel; and M3: Ca(OH)₂ paste plus 2% CHX gel] were used for 7 days. Checkerboard DNA–DNA hybridization was performed to detect 40 bacterial species. Aerobic and anaerobic culture techniques were used to determine the bacterial community by counting the colony-forming units (CFU).
Results:  The species most frequently identified by checkerboard in S1 were: Fusobacterium nucleatum ssp . polymorphum , Treponema socranskii ssp. socranskii , Parvimonas micra and Enterococcus faecalis. In S2 and S3 a total of eight different species were identified; and only one of them was gram-positive ( E. faecalis ). Microorganisms were not identified after use of M2 for 7 days. The quantification obtained on agar plates ranged from 4 × 10⁵ to 2.6 × 10⁶ CFU/ml in S1, mean CFU was reduced by 99.96% in S2, and there was no statistical difference between the CFU in S2 and S3.
Conclusion:  The antibacterial effect of the mechanical preparation supplemented by the use of an antibacterial auxiliary substance greatly reduced the microorganisms in the main root canal.     

Coptidis rhizoma inhibits growth and proteases of oral bacteria

OBJECTIVE: The aim of this study was to investigate the antibacterial effect of Coptidis Rhizoma (CR), a traditional medicinal plant, on oral bacteria. MATERIALS AND METHODS: CR extract was prepared by boiling CR in water for 2 h. Alkaloids contained in CR extract were assayed by high performance liquid chromatography (HPLC). Antibacterial activity of CR extract was estimated from the lowest concentration that did not permit bacterial growth (minimum inhibitory concentration, MIC) and the concentrations that inhibited 50% of bacterial proteolytic activity (IC50). RESULTS: CR extract inhibited the growth of Actinomyces naeslundii, Porphyromonas gingivalis, Prevotella intermedia, Prevotella nigrescens and Actinobacillus actinomycetemcomitans at MIC of 0.031-0.25 mg ml(-1), whereas it had less inhibitory effect (MIC: 0.5-2 mg ml(-1)) on the growth of Streptococcus and Lactobacillus. The major active component of CR extract was berberine (Ber), an alkaloid, and its inhibiting specificity to bacterial growth was similar to that of CR extract. CR extract and Ber were bacteriostatic at the MICs against most of the bacteria, and bacteriocidal at the concentrations higher than the MICs. Ber inhibited the activities of collagenase from P. gingivalis and A. actinomycetemcomitans. CONCLUSION: CR extract and Ber had an inhibitory effect on periodontopathogenic bacteria. These results suggest the possibility of their clinical application for the treatment of periodontal diseases.     

Apical extrusion of intracanal bacteria following use of two engine-driven instrumentation techniques

AIM: To evaluate the number of bacteria extruded apically from extracted teeth ex vivo after canal instrumentation using the two engine-driven techniques utilizing nickel-titanium instruments (ProTaper and System GT). METHODOLOGY: Forty extracted single-rooted human mandibular premolar teeth were used. Access cavities were prepared and root canals were then contaminated with a suspension of Enterococcus faecalis and dried. The contaminated roots were divided into two experimental groups of 15 teeth each and one control group of 10 teeth. Group 1, ProTaper group: the root canals were instrumented using ProTaper instruments. Group 2, System GT group: the root canals were instrumented using System GT instruments. Group 3, control group: no instrumentation was attempted. Bacteria extruded from the apical foramen during instrumentation were collected into vials. The microbiological samples from the vials were incubated in culture media for 24 h. Colonies of bacteria were counted and the results were given as number of colony-forming units. The data obtained were analysed using the Kruskal-Wallis one-way analysis of variance and Mann-Whitney U-tests, with alpha = 0.05 as the level for statistical significance. RESULTS: There was no significant difference as to the number of extruded bacteria between the ProTaper and System GT engine-driven systems (P > 0.05). CONCLUSIONS: Both engine-driven nickel-titanium systems extruded bacteria through the apical foramen.     

Infection-mediated early-onset periodontal disease in P/E-selectin-deficient mice

BACKGROUND: Retrospective and correlation studies suggest that early-onset periodontal disease may be due to a deficiency in phagocyte function, a pathogenic oral biofilm, and/or dysregulated gingival cytokine expression. Increased susceptibility to periodontal disease is therefore thought to result from multiple risk factors. METHODS: We tested this hypothesis prospectively using P/E-selectin adhesion molecule deficient mice that mimic the human syndrome leukocyte adhesion deficiency II. RESULTS: Our studies demonstrate that, in comparison to wild type animals, P/E-/- mice exhibit: spontaneous, early onset alveolar bone loss which is significant by 6 weeks of age; a 10-fold elevation in bacterial colonization of their oral cavities; and elevated gingival tissue levels of the bone resorptive cytokine IL-1alpha. Alveolar bone loss is completely prevented by prophylactic antibiotic therapy. CONCLUSIONS: These experiments provide the first prospective evidence for the multiple risk factor hypothesis of periodontal disease, and validate the first animal model for early onset periodontitis in which both the microbiota and host response can be systematically manipulated. P/E-/- animals should be useful in testing the virulence of putative periodontal pathogens, in determining the role of host resistance factors in periodontitis, in exploring the proposed relationship(s) between infection mediated alveolar bone loss and systemic health disorders, and exploring their genetic relationships.     

The effects of a triclosan/copolymer dentifrice on oral bacteria including those producing hydrogen sulfide

A clinical procedure was developed to examine the effects of short-term and extended use of a triclosan/copolymer dentifrice and a commercial fluoride dentifrice on oral bacteria, including those producing hydrogen sulfide. Healthy adults volunteered for this double-blind, crossover design clinical study and provided saliva samples for culturing on enriched and indicator media to enumerate all salivary bacteria and those producing hydrogen sulfide (odorigenic), respectively. Subjects brushed with an assigned dentifrice for 7 d and were sampled on day 8 to assess the long-term effects on bacteria. Extended use of the triclosan/copolymer dentifrice resulted in a 49% and 66% reduction of salivary and odorigenic bacteria, respectively, compared with the fluoride dentifrice. To examine short-term effects, subjects subsequently brushed with their assigned dentifrice and were sampled at 2 h and 4 h post-brushing. At 2 h and 4 h post-brushing, the triclosan/copolymer dentifrice resulted in a 62% and 52% decrease for salivary bacteria and 79% and 72% decrease for odorigenic bacteria, respectively, vs. the fluoride dentifrice. The results indicate a significant decrease of all salivary bacteria and hydrogen sulfide-producing odorigenic bacteria following use of the triclosan/copolymer dentifrice and explain previous results on the efficacy of this dentifrice on oral malodor.     

Microbial complexes in supragingival plaque

Background/aims:  To examine microbial communities in supragingival biofilm samples.
Methods:  Supragingival plaque samples were taken from 187 subjects at baseline ( n  = 4745). Fifty-five subjects provided supragingival plaque samples at 1–7 days after professional tooth cleaning ( n  = 1456); 93 subjects provided 8044 samples between 3 and 24 months post-therapy. All samples were individually analyzed for their content of 40 bacterial species using checkerboard DNA–DNA hybridization. Microbial associations among species were sought using cluster analysis and community ordination techniques for the three groups separately.
Results:  Six complexes were formed for the baseline samples. Similar complexes were formed for the samples taken 3–24 months post-therapy. However, distinct changes were observed in microbial communities in samples taken during the 7 days of plaque redevelopment. The complexes related to clinical parameters of periodontal disease.
Conclusion:  There were specific microbial complexes in supragingival plaque that were similar to those found in subgingival plaque samples with a few minor differences. The relation of previously unclustered taxa to the complexes was also described.     

In vitro evaluation of antimicrobial property of silver nanoparticles and chlorhexidine against five different oral pathogenic bacteria

Introduction

Numerous antimicrobial agents are used to eliminate oral biofilm. However due to emergence of multi drug resistant microorganisms, the quest to find out biologically safe and naturally available antimicrobial agents continues.

Antimicrobial barrier of an in vitro oral epithelial model

OBJECTIVE: Oral epithelia function as a microbial barrier and are actively involved in recognizing and responding to bacteria. Our goal was to examine a tissue engineered model of buccal epithelium for its response to oral bacteria and proinflammatory cytokines and compare the tissue responses with those of a submerged monolayer cell culture. DESIGN: The tissue model was characterized for keratin and beta-defensin expression. Altered expression of beta-defensins was evaluated by RT-PCR after exposure of the apical surface to oral bacteria and after exposure to TNF-alpha in the medium. These were compared to the response in traditional submerged oral epithelial cell culture. RESULTS: The buccal model showed expression of differentiation specific keratin 13, hBD1 and hBD3 in the upper half of the tissue; hBD2 was not detected. hBD1 mRNA was constitutively expressed, while hBD2 mRNA increased 2-fold after exposure of the apical surface to three oral bacteria tested and hBD3 mRNA increased in response to the non-pathogenic bacteria tested. In contrast, hBD2 mRNA increased 3-600-fold in response to bacteria in submerged cell culture. HBD2 mRNA increased over 100-fold in response to TNF-alpha in the tissue model and 50-fold in submerged cell culture. Thus, the tissue model is capable of upregulating hBD2, however, the minimal response to bacteria suggests that the tissue has an effective antimicrobial barrier due to its morphology, differentiation, and defensin expression. CONCLUSIONS: The oral mucosal model is differentiated, expresses hBD1 and hBD3, and has an intact surface with a functional antimicrobial barrier.     

Killing of Fusobacterium nucleaturn,Porphyromonas gingivalis and Prevotella intermedia by protegrins

Protegrins are broad spectrum antibiotic peptides isolated from porcine leukocytes. In this study, we (i) examine the sensitivity of Gram-negative, anaerobic periodontal pathogens to synthetic protegrins; (ii) determine the relative potencies of protegrin congeners against these bacteria; and (iii) compare the potency of protegrins with other antibiotic peptides, including magainin MSI-78, tachyplesin I, cecropin P1, human defensins HNP-1-3, and clavanin A. Synthetic l - and d -enantiomers of protegrin 1 (PG-1 and D-PG-1, respectively), and L-enantiomers of protegrins 2, 3 and 5 (PG-2, PG-3 and PG-5) were tested against Fusobacteriurn nucleatum, and black-pigmented organisms including Porphyromonas gingivalis and Prevotella intermedia. Strains of both F. nucleatum and the black-pigmented organisms were sensitive to PG-1, and exhibited mean ED₉₉ of 2.2-2.3 μg/ml and 3.4-9.9 μg/ml, respectively. The D-form was statistically more potent than the L-form against these oral anaerobes, and although this difference in potency is unlikely to be of decisive therapeutic significance, the d -form may be of value given ability to resist microbial and host-derived proteases. PG-1 was more potent than magainin, tachyplesin, cecropin, defensins and clavanin under test conditions. Hypertonic saIt concentrations and heat-inactivated serum were found to be inhibitory to the bactericidal activity of PG-1. PG-1 was found to induce morphologic alterations in the ultrastructural appearance of F. nucleatum consistent with damage to the bacterial membranes. We conclude that protegrins may be useful antimicrobial agents in therapy against Gram-negative anaerobic bacteria believed to be involved in chronic, adult forms of periodontal infections.