Paper Point Samples Research Articles

Does Exposure to Burning and Heated Tobacco Affect the Abundance of Perio-Pathogenic Species in the Subgingival Biofilm?

This study investigated the impact of tobacco exposure, specifically through heating and burning, on periodontopathogens in the subgingival microbiome among clinically healthy individuals and those diagnosed with periodontitis. The sample comprised 66 subjects (26–56 years, median 38 yrs; 64% females) classified as non-smokers, classic cigarette smokers, and tobacco heating system (THS) smokers (each N = 22). Full-mouth periodontal examination was performed, and 330 paper-point samples from periodontal pockets were collected. Next-generation sequencing of 16S rRNA genes was conducted to identify the composition of subgingival microbiome. Periodontitis prevalence among the groups was ranked as THS (41%) < non-smokers (44%) < cigarette smokers (68%), without statistically significant differences between the groups. The number of perio-pathogenic species was higher in subjects with periodontitis compared to those without (median 7 vs. 6 species; p = 0.005) but without significant differences between exposure groups: non-smokers (6) = smokers (6) < THS (6.5). When combining exposure and periodontal status, each smoker group had more perio-pathogenic species than non-smokers: non-smokers without periodontitis (5) < smokers without periodontitis (5.5) < THS without periodontitis (6); non-smokers with periodontitis (6.5) < THS with periodontitis (7) = smokers with periodontitis (7). Multiple linear regression indicated periodontitis as the sole predictor of perio-pathogenic species quantity, irrespective of the type of tobacco consumption, sex, age, or oral hygiene (R2 = 0.163; p = 0.005). In conclusion, the quantity of perio-pathogenic species in the subgingival microbiome was more influenced by periodontitis than by exposure to tobacco smoke, regardless of whether it was heated or burned.

Periradicular tissue fluid-derived biomarkers for apical periodontitis: An in vitro methodological and in vivo cross-sectional study.

Periradicular tissue fluid (PTF) offers a source of diagnostic, prognostic and predictive biomarkers for endodontic disease. (1) To optimize basic parameters for PTF paper point sampling in vitro for subsequent in vivo application. (2) To compare proteomes of PTF from teeth with normal apical tissues (NAT) and asymptomatic apical periodontitis (AAP) using high-throughput panels. (1) To assess volume absorbance, paper points (n = 20) of multiple brands, sizes and sampling durations were inserted into PBS/1%BSA at several depths. Wetted lengths (mm) were measured against standard curves to determine volume absorbance (μL). To assess analyte recovery, paper points (n = 6) loaded with 2 μL recombinant IL-1β (15.6 ng/mL) were eluted into 250 μL: (i) PBS; (ii) PBS/1% BSA; (iii) PBS/0.1% Tween20; (iv) PBS/0.25 M NaCl. These then underwent: (i) vortexing; (ii) vortexing/centrifugation; (iii) centrifugation; (iv) incubation/vortexing/centrifugation. Sandwich-ELISAs determined analyte recovery (%) against positive controls. (2) Using optimized protocols, PTF was retrieved from permanent teeth with NAT or AAP after accessing root canals. Samples, normalized to total fluid volume (TFV), were analysed to determine proteomic profiles (pg/TFV) of NAT and AAP via O-link Target-48 panel. Correlations between AAP and diagnostic accuracy were explored using principal-component analysis (PCA) and area under receive-operating-characteristic curves (AUC [95% CI]), respectively. Statistical comparisons were made using Mann-Whitney U, anova and post hoc Bonferonni tests (α < .01). (1) UnoDent's 'Classic' points facilitated maximum volume absorbance (p < .05), with no significant differences after 60 s (1.6 μL [1.30-1.73]), 1 mm depth and up to 40/0.02 (2.2 μL [1.98-2.20]). For elution, vortexing (89.3%) and PBS/1% BSA (86.9%) yielded the largest IL-1β recovery (p < .05). (2) 41 (NAT: 13; AAP: 31) PTF samples proceeded to analysis. The panel detected 18 analytes (CCL-2, -3, -4; CSF-1; CXCL-8, -9; HGF; IL-1β, -6, -17A, -18; MMP-1, -12; OLR-1; OSM; TNFSF-10, -12; VEGF-A) in ≥75% of AAP samples at statistically higher concentrations (p < .01). CXCL-8, IL-1β, OLR-1, OSM and TNFSF-12 were strongly correlated to AAP. 'Excellent' diagnostic performance was observed for TNFSF-12 (AUC: 0.94 [95% CI: 0.86-1.00]) and the PCA-derived cluster (AUC: 0.96 [95% CI: 0.89-1.00]). Optimized PTF sampling parameters were identified in this study. When applied clinically, high-throughput proteomic analyses revealed complex interconnected networks of potential biomarkers. TNFSF-12 discriminated periradicular disease from health the greatest; however, clustering analytes further improved diagnostic accuracy. Additional independent investigations are required to validate these findings.

Effect of Passive Ultrasonic Irrigation, Er,Cr:YSGG Laser, and Photon-Induced Photoacoustic Streaming against Enterococcus faecalis Biofilms in the Apical Third of Root Canals.

This study aimed to compare the antibacterial effectiveness of passive ultrasonic irrigation (PUI), Er,Cr:YSGG laser (WTL), and photon-induced photoacoustic streaming (PIPS) using an Er:YAG laser against Enterococcus faecalis biofilms in the apical third of root canals. Root canals of 70 single-rooted human teeth were instrumented and infected with E. faecalis for 3 weeks to form biofilms. The samples were randomly divided into five groups as follows: (i) PUI + 3% NaOCl (n = 16); (ii) Er,Cr:YSGG laser (n = 16); (iii) PIPS + 3% NaOCl (n = 16); (iv) positive control group (n = 10); and (v) negative control group (n = 10). The bacterial content in the root canal was sampled using (a) the paper-point sampling method before (S1) and after (S2) treatment and (b) pulverising the apical 5 mm of the root. The number of bacteria recovered from each group was counted as colony-forming units (CFUs). The amount of reduction between the groups was compared with the Kruskal-Wallis test and post-test Dunn's multiple comparisons tests. The significance level was set at 5% (p < 0.05). The samples from the paper-point sampling method showed that the amount of bacteria before (S1) and after treatment (S2) was significantly different between PIPS and WTL, as well as between the PUI and WTL groups. In contrast, no significant difference was found between the PIPS and PUI groups. From the pulverised samples, the results indicated no significant difference among all experimental groups in the amount of bacterial reduction in the apical 5 mm of the root. PUI and PIPS showed a significantly greater reduction in bacterial content within the main root canal compared with the WTL. There was no difference among all experimental groups in the apical third of the root.

In vitro antibacterial effects of photodynamic therapy against Enterococcus faecalis in root canals of deciduous teeth

ObjectiveThis study aimed at evaluating the in vitro antibacterial efficacy of photodynamic therapy (PDT) on planktonic E. faecalis and its biofilm in the root canal of infected deciduous teeth.MethodsForty root canals of maxillary deciduous anterior teeth were enlarged up to #35 K-file and inoculated with E. faecalis for 21 days. The root canals were randomly assigned into four groups (n = 10): The normal saline group (control), 1% NaClO group, PDT group, and the 1% NaClO + PDT group. Paper point samples were obtained at baseline (S1) and after treatment (S2). The colony-forming units (CFU) were counted, and the bacterial growth rate calculated. From each subgroup, 5 samples were randomly selected after treatment and a scanning laser confocal microscope (CLSM) used to determine the distribution of dead / living bacteria on the biofilm surface of each subgroup. A scanning electron microscope (SEM) was used to observe bacterial morphologies in the root canal walls of the remaining 5 samples in each subgroup. The Kruskal–Wallis test and Dunn test with boferroni adjustment were used to analyze the effect of the different treatment techniques on the E. faecalis in root canals.ResultsCompared to the saline group, PDT significantly reduced bacterial counts in the root canal (p < 0.05). The CFU counts were lowest (p < 0.05) in the 1% NaClO and in 1% NaClO + PDT groups. The rate of bacterial death on the surface of the biofilm in the PDT group was significantly increased after treatment (p < 0.05), and the rate of bacterial death was highest in 1%NaClO group and 1%NaClO + PDT group (p < 0.05).ConclusionPDT has an antibacterial activity against E. faecalis in the root canal of deciduous teeth. Its activity against planktonic E. faecalis is better than the activity on the intact biofilm. The antibacterial activity of PDT on E. faecalis in root canals of deciduous teeth is lower compared to that of 1% NaClO.

A modified dentine infection model with fluorescent lipopolysaccharide and lipopolysaccharides sampling technique to compare XP-Endo finisher and passive ultrasonic irrigation.

The lipopolysaccharides-dentine-infection (LPS-dentine-infection) models and sampling techniques frequently used to evaluate LPS disinfection have limitations. In this study, a LPS-dentine-infection model was devised using fluorescent conjugate LPS. Secondly, a sampling technique using cryogenic grinding for intraradicular LPS analysis was evaluated. Thirdly, the effectiveness of the XP-endo Finisher (XP-EF) was compared with passive ultrasonic irrigation (PUI) in removing LPS from root canal system. Sixty-nine mandibular premolars were submitted to dentine pretreatment and inoculated with fluorescent LPS conjugate (Alexa Fluor® 594). Twenty-three teeth were analysed under confocal laser scanning microscopy (CLSM) to validate this modified LPS-dentine-infection model. Forty-six teeth were randomly divided into two experimental groups: XP-EF (n=23) and PUI (n=23). All teeth were instrumented with XP-endo shaper (XPS; FKG Dentaire) and 2.5% NaOCl. The root canals were sampled with paper points before (s1) and after (s2) instrumentation and after supplemental treatment (s3) with XP-EF and PUI. After s3, all roots were cryogenically ground for intraradicular LPS analysis (s4). Limulus amebocyte lysate assay was used for LPS quantification. The Friedman test was used for differences in LPS among four time-points (s1, s2, s3, and s4). Dunn's test was used for pairwise testing of time-points. The significance level was set at 5% (p < .05). Fluorescent LPS conjugate was detected in 100% of the samples under CLSM with a penetration depth of approximately 400 μm into dentine. Chemo-mechanical preparation using XPS files significantly reduced LPS levels (p < .05). Both the XPS and PUI improved the LPS disinfection (p < .05), with no difference between them (p > .05). LPS was recovered from all samples after cryogenic grinding. The residual amount of LPS detected using the cryogenically sampling technique at s4 was approximately three times greater than with the paper-point sampling technique at s3. This study established a modified LPS-dentine-infection model using fluorescent conjugate LPS, and validated a LPS sampling technique for using cryopulverization intraradicular LPS analysis. Moreover, both the XP-EF and PUI further improved LPS disinfection from the root canals, and the innovative XP-EF was as effective as PUI.

Prevalence Of Epstain–Bar Virus Among Patients With Periodontitis In Al–Najaf Al–Ashraf / Iraq

Periodontitis, a complex chronic inflammatory disease caused by subgingival infection, is among the most prevalent microbial diseases in humans which characterized by periodontal damage, alveolar bone resorption, pain, and eventual tooth loss. Epstein–Barr Virus (EBV) has widely infected &gt;90% adults in the world and is associated with many human diseases, so that this study aimed to investigate the association between EBV and periodontitis.&#x0D; Patients and samples: Subgingival paper point samples were collected from 100 patients with periodontitis and 30 healthy people. All samples undergo direct DNA extraction to amplified EBVs DNA using PCR technique.&#x0D; The results indicated a high percentage of EBV infections (18%) in patient suffering from periodontitis while there was no EBV infection were detected in healthy persons. A high percentage of EBV was detected in female (56%) in comparison with male (44%).&#x0D; The results improved the association between EBV and periodontitis suggesting that EBV may serve as a pathogenic factor leading to periodontitis among patients.

Comparison of the Antibacterial Efficacy of Ibuprofen Versus Triple Antibiotic Paste Against Enterococcus Faecalis in Single Rooted Teeth

Aim: The aim of the present study was to compare the antibacterial efficiency of the anti-inflammatory non-antibiotic ibuprofen versus triple antibiotic paste as an intra-canal medication against Enterococcus faecalis. Methodology: Forty four single rooted teeth were decoronated and instrumented up to F4 ProTaper rotary files. Apical foramen was sealed with acrylic resin, and all the external surfaces were made impermeable with nail varnish, except for the coronal access. Roots were autoclaved at 121°C for 20 minutes, placed in Eppendorf tubes and contaminated with E. faecalis for 14 days. Teeth were divided into two groups of 22 teeth; (Group1: ibuprofen, Group2: triple antibiotic paste (TAP). Colony forming unit (CFU) counts were taken before placing the intracanal medications by a paper point sampling (CFU1). The tested medicaments were mixed with distilled water (1:1 w/v), placed inside the root canals, then canals were temporarily sealed, and incubated at 37°C for 7 days. After intracanal medication removal, the second count (CFU2) was taken as (CFU1) and the antibacterial action of each medication was determined by calculating the percentage of bacterial reduction when comparing CFU2 to CFU1, and then the antibacterial action of the tested medications was compared with each other. Results: There was no statistically significant difference between (T.A.P) and (Ibuprofen) groups in the percentage of bacterial reduction where (p=0.073). Both medications caused a reduction in the bacterial count from CFU1 to CFU2. Conclusion: Ibuprofen proved to have an antibacterial effect comparable to that of TAP.

Comparison of DNA Extracted from Pediatric Saliva, Gingival Crevicular Fluid and Site-Specific Biofilm Samples.

(1) Introduction: Due to the non-invasive nature of saliva, many methods have been used to isolate and collect DNA from saliva samples for microbial screening. Many oral microbes also inhabit the oral biofilm, which may represent significantly different microbial constituents that may contribute to oral health and disease, including caries and periodontal disorders. Moreover, the biofilm may vary within the same patient at different sites. Few studies have evaluated the comparison between DNA isolated from saliva and DNA from site-specific biofilm, with virtually no studies addressing this analysis among pediatric patients. (2) Methods: An existing repository of paper point derived biofilm, gingival crevicular fluid (GCF), and unstimulated saliva samples previously collected from pediatric patients (n = 47) was identified. DNA was isolated from biofilm sites (tongue, upper buccal molar, mandibular lingual incisor), and GCF and saliva were used for quantitative DNA comparison using a phenol:chloroform extraction. A quantitative and qualitative analysis was performed using the NanoDrop 2000 spectrophotometer using absorbance readings at A230 nm, A260 nm and A280 nm. (3) Results: These data demonstrated the successful isolation of DNA from all of the patient samples, with the highest concentrations observed among unstimulated saliva (4264.1 ng/μL) and the lowest derived from GCF (1771.5 ng/μL). No differences were observed between males and females or minorities and non-minority patients. In addition, comparison of the overall concentrations of DNA obtained from adult samples was slightly higher than, but not significantly different from, the concentrations obtained from pediatric samples (p = 0.2827). A real-time quantitative qPCR screening revealed that all of the samples evaluated harbored bacterial and human DNA of sufficient quantity and quality for a molecular screening greater than the limit of detection (ΔRn = 0.01). (4) Conclusions: Many methods are currently available to provide the sampling and screening of saliva and specific sites within the oral cavity, but the validation and comparison of simple and low-cost methods, that include paper point sampling and unstimulated saliva collection, may suggest these methods and protocols provide sufficient DNA quality and quantity for molecular screening and other comparison applications. In addition, although heterogeneity will be a constant and consistent feature between patient samples, standardized methods that provide similar and consistent DNA from various oral sites may provide needed consistency for screening and molecular analysis.

Rapid Chairside Microbial Detection Predicts Endodontic Treatment Outcome.

Background. The aim of this longitudinal, one-year cohort study was to explore the hypothesis that fluorescence sampling of the root canal space prior to obturation could predict the outcome of root canal treatment (RCT). Methods. Sixty-five teeth underwent primary RCT and were followed up clinically and radiographically. The outcome was determined radiographically with periapical radiographs (PR) and cone beam computed tomography (CBCT) scans. Results. Success at 12 months was predictable based on the fluorescence score. When the fluorescence score (defined as the percentage of signal over total signal including background) was lower than 67, there was a 4.5 times (Odds ratio (OR) = 0.028; 95% confidence interval (CI): 0.003, 0.291, p = 0.001) greater chance of success (90% overall). When the readings were above this threshold, the success rate was 20%. Conclusion. A chairside sampling method is able to predict the outcome of RCT, through the use of paper point sampling and fluorescence staining. This has reduced the prevalence of persistent infections by guiding the optimum time for obturation. ClinicalTrials.gov trial NCT03660163.

Enterococci in the oral cavity of periodontitis patients from different urban socioeconomic groups

Enterococci are the transient constituents of the oral microbiome and have been now increasingly related to oral and systemic diseases. They have now become the most notable nosocomial pathogens and also been linked to etiology of periodontitis. This study evaluates the prevalence of Enterococci in the chronic periodontitis and healthy Indian cohort in different urban socioeconomic groups. In this retrospective study, individuals in the age range of 18.75 years were included. Seventy individuals had mild-to-moderate periodontitis and 30 healthy persons were taken as controls. A questionnaire was administered. Paper point samples of gingival crevicular fluid were obtained, pooled, and sent to microbiology laboratory in a transport media. Forty-six isolates were identified as enterococci for statical analyzes Pearson's Chi-square test used and. P < 0.001 was considered to be significant. The prevalence of enterococci was seen mostly in the individuals from lower socioeconomic class, having poor oral hygiene, and smokers. This was significantly different from those of upper class (P < 0.001). The predominant species isolated was Enterococcus faecalis. Enterococci particularly E. faecalis followed by Enterococcus faecium could play a crucial role in the severity or progression of periodontitis particularly in a favorable oral environment.

Quantitative Comparison of Oral Site-specific DNA Isolates Reveals Differential Outcomes

Introduction: More and more evidence has accumulated that suggests salivary sampling may provide direct analysis of oral conditions and microbial constituents, but may also be useful in the diagnosis and early detection of other chronic diseases. Although multiple methods of oral sampling currently exist, some methods are prohibitively expensive or based upon technologies not ubiquitously available at public health centers or state-funded colleges. This study provides a comparative analysis of DNA concentrations and quality from five specific oral sites derived using sterile paper points, including the gingival crevice between the upper central incisors, biofilm of the upper first molar, lingual incisor, and the dorsum of the tongue for comparison with unstimulated saliva collection.&#x0D; Methods: This study analyzed previously collected unstimulated saliva and paper point samples. In brief, DNA was isolated from each using TRIzol (phenol: Chloroform) extraction and DNA quantification and quality was measured using a NanoDrop spectrophotometer at 260 and 280 nm.&#x0D; Results: Analysis of Paper Point (PP) biofilm sampling sites from upper first molar, lower incisor, and dorsum of the tongue revealed similar average DNA concentrations, ranging between 14,342 ng/uL and 14,402 ng/uL (p=0.9851). Although variations were observed between different patients, samples from different oral sites within the same patient were strikingly similar, R=0.8355. Comparison of DNA isolated from fluids, gingival crevicular fluid (GCF) and unstimulated saliva revealed average DNA concentrations that were similar to the biofilm sampling sites (14,686 ng/uL and 13,743 ng/uL, respectively), which were not significantly different from one another (p=0.7893). DNA concentrations ranged considerably between patients (low = 4,410 ng/uL; high = 48,783 ng/uL), but were most similar with different samples (GCF, saliva) from the same patient (Pearson’s R=0.6979). In addition, DNA purity measured by A260:A280 nm absorbance did not reveal any significant difference among sampling sites (range 1.62 – 1.70; p=0.427).&#x0D; Discussion: Although many methods are available to provide oral sampling, simple and low-cost methods such as paper point sampling, unstimulated saliva collection and buccal swabs may represent tools that provide sufficient DNA quality and quantity for molecular screening. In addition, although heterogeneity between patient samples will always be present – samples from various oral sites within the same patient may provide roughly equivalent DNA samples for further screening and molecular analysis.

The synthesis of nano silver-graphene oxide system and its efficacy against endodontic biofilms using a novel tooth model

ObjectiveThe deleterious caustic effects of sodium hypochlorite (NaOCl) as a root canal irrigant makes it imperative that alternative methods are developed for root canal disinfection. The purpose of this study was to examine the antimicrobial efficacy of silver nanoparticles (AgNPs) synthesized on an aqueous graphene oxide (GO) matrix (Ag-GO), with different irrigant delivery methods to enhance the disinfection regimen, using a novel ex vivo infected tooth model. MethodsAgNPs were prepared by reducing AgNO3 with 0.01M NaBH4 in presence of GO. Elemental analysis was performed with scanning electron microscopy/energy dispersive X-ray spectroscopy (SEM/EDS) and scanning transmission electron microscopy (STEM) was used for size and morphology analysis of GO and Ag-GO. Nutrient stressed, multi-species biofilms were grown in prepared root canals of single-rooted teeth. The irrigants used were sterile saline, 1% and 2.5% NaOCl, 2% chlorhexidine gluconate (CHX), 17% EDTA and an aqueous suspension of 0.25% Ag-GO. The antimicrobial efficacy of the irrigants were performed with paper point sampling and measurement of microbial counts. The biofilm disruption in dentine tubule surfaces was analysed with confocal laser scanning microscopy (CLSM). The acquisition of total biovolume (μm3/μm2) and biofilm viability was performed using software BioImage_L. Two-way analysis of variance (ANOVA) with post hoc Tukey tests was used for data analysis with level of statistical significance set at P<0.05. ResultsSEM/EDS analysis confirmed impregnation of Ag within the GO matrix. TEM images showed polygonal GO sheets and spherical AgNPs of diameter 20–50nm, forming a network on the surface of GO sheets. The use of ultrasonic activation enhanced the efficacy of Ag-GO compared to 1% NaOCl, 2% CHX, 17% EDTA and sterile saline (P<0.05). The microbial killing efficacy of 2.5% NaOCl was superior compared to the experimental groups. The maximum biofilm disruption, in dentine tubule surfaces, was achieved by 2.5% NaOCl, however Ag-GO caused a significant reduction of total biovolumes compared to the rest of the experimental groups (P<0.05%). SignificanceThe successful documentation of the microbial killing and biofilm disruption capacity of Ag-GO is a promising step forward to explore its unique properties in clinical applications and biomaterials in dentistry.

A Promising Probiotic Irrigant: An In Vitro Study.

AIM:The present study aimed to investigate the inhibitory effect of Lactobacillus rhamnosus (B-445) as a probiotics irrigant on the growth of Enterococcus faecalis.METHODS:Forty-two extracted single human canal anterior teeth were prepared with rotary instrumentation and sterilised. Teeth were divided into 3 groups according to the type of irrigant, N = 14. Three experimental groups were inoculated with E. faecalis and cultured for 21 days before use; Group 1 was 2.5% NaOCl (positive control), Group 2 was saline (negative control), Group 3 was the experimental probiotic irrigant. Paper point sampling of the canals of each group was obtained before irrigation (S1), immediately after irrigation (S2) and after 24 hours (post irrigation samples) (S3) to determine remaining colony forming units for E. faecalis. Also, Colony counts for L. rhamnosus in Group 3 after immediate irrigation, as well as 24 hours post irrigation, was performed to determine the survival profile of these bacteria in infected root canal with E. faecalis.RESULTS:The NaOCl irrigant group had the lowest mean value of (log 10 CFU/mL) of E. faecalis after immediate irrigation and after 24 hrs post irrigation followed by the probiotic group, while the highest mean value was the saline group (P ≤ 0.001). The survival profile for L. rhamnosus in Group 3 after immediate irrigation and post-irrigation were slightly higher than for E. faecalis (P ≤ 0.001).CONCLUSION:Lactobacillus rhamnosus which revealed a potential inhibitory effect on the growth of Enterococcus faecalis, could be used as a new natural, safe probiotic irrigant agent.

Antimicrobial efficacy of medium chain fatty acids as root canal irrigants: An in vitro study

Enterococcus faecalis and Candida albicans are the microbes that are most resistant to elimination by disinfecting agents and are the causative agents for reinfection of the root canal treated teeth. Medium chain fatty acids (MCFAs), which are the main components of coconut oil, are proven antimicrobial agents. Thus, the aim of this study was to evaluate their antimicrobial efficacy against E. faecalis and C. albicans. Ninety extracted single-rooted mandibular premolar teeth were decoronated, biomechanically prepared, autoclaved, and divided into three groups (n = 30): Group A (inoculated with E. faecalis), Group B (inoculated with C. albicans), and Group C (control group). Each group was again subdivided into three groups (n = 10) and irrigated with lauric acid (LA), decanoic acid (DA), and octanoic acid, respectively, for 5 min. Paper point samples were taken from canal walls and transferred into Brain Heart Infusion broth and potato dextrose broth and placed in an incubator at 37°C. The appearance of tubidity was checked at 24, 48, 72, and 96 h using direct contact test. The data were then statistically analyzed using the analysis of variance and Tukey honestly significant difference post hoc tests. Among the three MCFAs, LA showed the maximum inhibitory against E. faecalis at 24 h and the inhibitory activity decreased considerably at 48, 72, and 96 h. DA was the most effective against C. albicans with a maximum inhibition at 48 h. DA also showed significant substantivity at 72 and 96 h. Within the limitations of this study, it can be concluded that MCFAs show promising antimicrobial efficacy against E. faecalis and C. albicans.

Effectiveness of different endodontic irrigants against Enterococcus faecalis: an ex vivo study.

Aim: To evaluate the effectiveness of different endodontic irrigants against Enterococcus faecalis (ATCC 29212).Methods: Seventy bovine mandibular incisors were prepared, inoculated with a bacterial strain for 60 days and divided into the following groups: positive control; negative control; 2.5% NaOCl; 17% EDTA; 0.2% chitosan; 2.5% NaOCl + 0.2% chitosan; and 2.5% NaOCl + 17% EDTA. The irrigation protocol was performed using an experimental peristaltic pump device, with the irrigating solutions circulating within the apparatus at a constant flow for 10 min. Paper-point samples were then collected from the root canals and immersed in 7 mL of brain heart infusion broth, followed by incubation at 37°C for 48 h. Bacterial growth was assessed by turbidity of the culture medium.Results: E. faecalis was present in all samples after the use of different irrigants.Conclusion: The different irrigants tested were not effective in completely eliminating dentin bacterial contamination with E. faecalis.

Comparison of anti-bacterial efficiency of ibuprofen, diclofenac, and calcium hydroxide against Enterococcus faecalis in an endodontic model: An in vitro study.

Context:One of the important goals of root canal therapy is disinfection aided by irrigation and intracanal medicaments. Commonly used nonsteroidal anti-inflammatory agents have shown anti-bacterial activity, thus, when used as intracanal medicaments, nonantibiotic agents have the potential to provide anti-inflammatory, local analgesic, and anti-bacterial activity.Aim:The aim of this study is to evaluate and compare the anti-bacterial efficiency of anti-inflammatory nonantibiotics ibuprofen and diclofenac, and routinely used intracanal dressing calcium hydroxide (Ca[OH]2), against Enterococcus faecalis, in an endodontic model.Materials and Methods:A total of 76 single-rooted mandibular premolar teeth were decoronated and instrumented up to F4-ProTaper rotary. Apical foramen was sealed with composite resin, and all external surfaces made impermeable with nail varnish, except for coronal access. Roots were autoclaved (121°C for 20 min), placed in Eppendorf tubes, and contaminated with E. faecalis for 14 days. Colony-forming unit (CFU) counts were taken before (CFU-1), and after intracanal medication (CFU-2) by paper point sampling. Test medicaments (Group-1: ibuprofen, Group-2: diclofenac, Group-3: Ca[OH]2) were mixed with distilled water (1:1 w/v), placed into root canals, temporarily sealed, and incubated (37°C; 7 days). Group-4 received no medicament (control). Kruskal–Wallis ANOVA was used to compare the four groups and Mann–Whitney U-test for pair-wise comparisons.Results:Within the limitations of the study, anti-inflammatory nonantibiotics (ibuprofen; diclofenac) were shown to have anti-bacterial effect against E. faecalis.Conclusion:Since nonsteroidal anti-inflammatory drugs (NSAIDs) have an anti-bacterial effect, it is possible to replace Ca(OH)2 with NSAIDs, or even combine them to form a cocktail of local disinfectants to optimize canal disinfection.

Comparison of paperpoint and curette sampling of subgingival microbiome composition as analyzed by 16S rRNA gene amplicon sequencing

ABSTRACTObjective: Our aim was to compare the subgingival microbiome composition in samples collected by curettes and paperpoints.Methods: Subgingival plaque of rheumatoid arthritis patients with periodontitis (N=66) or gingivitis (N=15) was collected at two timepoints by sterile curettes and paperpoints. Unused sterile paperpoints were included as controls. The microbial DNA was processed for 16S rRNA gene amplicon sequencing. The data was processed and clustered into Operational Taxonomic Units (OTUs) and assessed by multivariate analyses.Results: Unused paperpoints showed various levels of DNA contamination. The OTUs specific for unused paperpoints were classified as Exiguobacterium, Enterococcus, Methylobacterium, Aquabacterium and Pseudomonas and were removed from the dataset.Microbial profiles of curette samples differed significantly from the paperpoint samples (PERMANOVA, p=0.0009, F=3.167). The paperpoint samples had significantly higher proportion of OTUs classified as Streptococcus, Gemella, Parvimonas, Haemophilus, Aggregatibacter and Clostridiales family XIII incertae sedis (p<0.05). Curette samples harbored higher proportion of Corynebacterium, Prevotella, Selenomonas, Actinomyces and Treponema (p<0.05).Curette samples had significantly higher species richness (p=0.01) and Shannon Diversity Index (p=0.009) than the paperpoint samples.Conclusions: Different subgingival plaque sampling techniques result in different microbiome profiles. Samples by paperpoints introduce microbial DNA contaminants and show underestimated microbial diversity compared to samples obtained by curettes.

Rapid Bacterial Detection during Endodontic Treatment

Bacteria present in the root canal (RC) space following an RC treatment (RCT) can lead to persistent infections, resulting in treatment failure and the need for reintervention or extraction. Currently, there are no standardized methods in use to clinically detect bacterial presence within RC spaces. The use of paper point sampling and fluorescence staining was shown to be a rapid method, able to detect residual bacteria following treatment. The study demonstrated that Calcein acetoxymethyl (AM) proved to be a suitable dye for detecting vital bacteria within mature endodontic biofilms, with an improved sensitivity over colony-forming unit counting in a stressed biofilm model. Furthermore, in a clinical trial with primary RCTs, 53 infected teeth were sampled in vivo, and increased detection of vital cells was found when compared with colony-forming unit counting, highlighting the sensitivity of the technique in detecting low cell numbers. By combining fluorescent staining and microspectroscopy with software-based spectral analysis, successful detection of vital cells from RCs was possible after 5 min of Calcein AM incubation. Application of this technology during RCT has the potential to reduce persistent infections through vital cell detection and additional treatment. Furthermore, this technique could be applied to antimicrobial research and disinfection control in clinical settings (ClinicalTrials.gov NCT03055975).

In Situ Anabolic Activity of Periodontal Pathogens Porphyromonas gingivalis and Filifactor alocis in Chronic Periodontitis.

Porphyromonas gingivalis and Filifactor alocis are fastidious anaerobic bacteria strongly associated with chronic forms of periodontitis. Our understanding of the growth activities of these microorganisms in situ is very limited. Previous studies have shown that copy numbers of ribosomal-RNA precursor (pre-rRNA) of specific pathogen species relative to genomic-DNA (gDNA) of the same species (P:G ratios) are greater in actively growing bacterial cells than in resting cells. The method, so-called steady-state pre-rRNA-analysis, represents a novel culture-independent approach to study bacteria. This study employed this technique to examine the in situ growth activities of oral bacteria in periodontitis before and after non-surgical periodontal therapy. Sub-gingival paper-point samples were taken at initial and re-evaluation appointments. Pre-rRNA and gDNA levels of P. gingivalis and F. alocis were quantified and compared using reverse-transcriptase qPCR. The results indicate significantly reduced growth activity of P. gingivalis, but not F. alocis, after therapy. The P:G ratios of P. gingivalis and F. alocis were compared and a low-strength, but statistically significant inter-species correlation was detected. Our study demonstrates that steady-state pre-rRNA-analysis can be a valuable culture-independent approach to studying opportunistic bacteria in periodontitis.

Detection and Quantification of Periodontopathogenic Bacteria in Subgingival Plaque Samples on Patients Undergoing Orthodontic Treatment

Abstract Introduction: According to last years' research, periodontopathogens may have a negative impact on treatment options in patients with periodontal lesions. However, not all infected sites suffer periodontal destructions, which can be explained on the assumption that only a limited number of pathogens present in a sufficient amount, are capable of affecting the periodontal tissue. Thermal cycling polymerase chain reaction (PCR) is a new technique used for the identification and quantification of periodontopathogenic bacteria. The aim of our study was to confirm the presence of periodontal pathogens, and to evaluate the amount of microbacterial pathogens in the periodontal pockets of patients undergoing orthodontic treatment for a more predictable result. Material and methods: A total amount of 32 subgingival samples were collected from periodontal pockets ≥6 mm in 8 patients. Clinical examinations, periapical radiographs and periodontal screenings were performed. Only patients undergoing orthodontic treatment with fixed appliances were included in the study. PCR and DNA hybridization-based identification were performed by paper-point sampling using a micro-IDent plus, Hain Lifescience Germany kit. Results and Discussions: Results showed that bacterial load may be connected to disease progression. The prevalence of the periodontopathogenic bacteria Actinobacillus a. was established in 42.8% of cases, P. Gingivalis in 71.42%, P. Intermedia 57.14%, Bacteroides F. was found in 85.71% of cases, Treponema D. in 100% of cases. Extremely high bacterial loads were recorded for Actinobacillus a., Bacteroides F. and Prevotella I.