Endodontic Pathogens Research Articles

Comparative evaluation of antibacterial efficacy of nitrofurantoin, chitosan, and calcium hydroxide in combination with propylene glycol as an intracanal medicament against endodontic pathogen - An in vitro study.

The objective is to evaluate and compare the antibacterial efficacy of nitrofurantoin, chitosan, and calcium hydroxide (Ca(OH)2) in combination with propylene glycol (PG) as an intracanal medicament against endodontic pathogens. Fifty-two extracted single-rooted maxillary and mandibular anterior teeth and premolars were selected. The root canals were enlarged using Protaper universal rotary files. Clinical isolates of microorganisms collected from retreatment cases were used. Bacterial isolates obtained from infected root canals were introduced into brain-heart infusion (BHI) broth. Incubation of samples for 14 days was carried out to facilitate the development of mature biofilms. Intracanal medicaments were divided into four groups: Group 1 - Nirofurantoin+20% PG, Group 2 - Chitosan+20% PG, Group 3 - Ca(OH)2 + 20% PG, and Group 4 - 20% PG. The prepared root samples were incubated for 7 days. After collecting dentin samples, they were placed in a phosphate-buffered saline solution. Serial dilutions were then performed, and each dilution was plated on BHI agar. The plates were incubated for 24 h at 37°C. The antibacterial efficacy was assessed by calculating the percentage of remaining colony-forming units. Antibacterial efficacy of chitosan paste was significantly higher followed by nitrofurantoin as compared to other groups when used as an intracanal medicament. In the root-canal biofilm model, the combination of chitosan and PG demonstrated a significant reduction in the viability of endodontic pathogens when employed as intracanal medication for 7 days. This suggests its potential as an effective intracanal medicament for endodontic retreatment.

Antimicrobial Silk Fibroin Methacrylated Scaffolds for Regenerative Endodontics

IntroductionRecognizing the necessity of novel disinfection strategies for improved bacterial control to ultimately favor tissue regeneration, this study developed and characterized antibiotics-laden silk fibroin methacrylated (SilkMA) scaffolds for regenerative endodontics. MethodsSilkMA-based solutions (10% w/v) containing Clindamycin (CLI) or Tinidazole (TIN) (0 – control; 5, 10, or 15% w/w) or the combination of both drugs (BiMix CLI/TIN 10%) were electrospun and photocrosslinked. Morphology and composition were assessed using scanning electron microscopy and Fourier-transform infrared spectroscopy. Additionally, swelling and degradation profiles were also determined. Cytotoxicity was evaluated in stem cells from apical papilla. Antibacterial efficacy was tested using direct and indirect contact assays against Aggregatibacter actinomycetemcomitans/Aa, Actinomyces naeslundii/An, Enterococcus faecalis/Ef, and Fusobacterium nucleatum/Fn. E. faecalis biofilm inhibition on dentin discs was specifically evaluated for BiMix-laden scaffolds. Data were statistically analyzed with a significance level of 5%. ResultsScanning electron microscopy revealed that all scaffolds had similar characteristics, including fiber morphology and bead absence. Fourier-transform infrared spectroscopy showed the incorporation of CLI and TIN into the fibers and in BiMix scaffolds. Antibiotic-laden scaffolds exhibited lower swelling capacity than the control and were degraded entirely after 45 days. Scaffolds laden with CLI, TIN, or BiMix throughout all time points did not reduce stem cells from apical papilla's viability. CLI-laden scaffolds inhibited the growth of Aa, An, and Ef, while TIN-laden scaffolds inhibited Fn growth. BiMix-laden scaffolds significantly inhibited Aa, An, Ef, and Fn in direct contact, and their aliquots inhibited An and Fn through indirect contact, with additional biofilm inhibition against Ef. ConclusionsBiMix-laden SilkMA scaffolds are cytocompatible and exhibit antimicrobial effects against endodontic pathogens, indicating their therapeutic potential as a drug delivery system for regenerative endodontics.

A Novel Combination of Zinc Oxide with Two Essential Oils Exerts Antimicrobial Effect against Endodontic Pathogens In Vitro.

Evaluating the antimicrobial efficacy of the novel combinations of zinc oxide mixed with ajwain oil (ZNOA) and combination of ajwain and eugenol (ZNOAE) vs conventionally used zinc oxide eugenol (ZNOE) against endodontic pathogens like Escherichia coli and Enterococcus faecalis. The pure cultures of E. coli (MTCC 443) and E. faecalis (MTCC 439) were revived and grown on selective cultural media. The minimum inhibitory concentration (MIC) and minimum bactericidal concentration (MBC) of the test materials were determined correspondingly through sequential dilution and agar well diffusion methods, as per Clinical and Laboratory Standard Institute (CLSI) guidelines. The data values were presented as mean ± standard deviation (SD). The comparisons among groups were completed through the one-way analysis of variance (ANOVA) test, whereas intragroup pairwise comparisons were completed using the unpaired t-test (p < 0.05). Minimum inhibitory concentration values against E. coli and E. faecalis of ZNOE were 250 and 500 µg/mL, ZNOA was 250 µg/mL, and ZNOAE were 125 and 250 µg/mL, correspondingly. MBC values in the form of inhibition zone against E. coli by ZNOE were 21.33 ± 1.53 mm, ZNOA 18.67 ± 1.53 mm, and ZNOAE 20.33 ± 1.53 mm. The E. faecalis inhibition zone for ZNOE was 14.33 ± 2.08 mm, ZNOA 18.67 ± 2.08 mm, and ZNOAE 24.33 ± 1.53 mm. All test materials demonstrated good antibacterial efficacy. However, between the novel combinations of test materials, ZNOA showed better antimicrobial efficacy against resistant endodontic pathogens than ZNOE. Dahake PT, Joshi SS, Kale YJ, et al. A Novel Combination of Zinc Oxide with Two Essential Oils Exerts Antimicrobial Effect against Endodontic Pathogens In Vitro. Int J Clin Pediatr Dent 2024;17(S-1):S11-S16.

Pectin as a Biomaterial in Regenerative Endodontics-Assessing Biocompatibility and Antibacterial Efficacy against Common Endodontic Pathogens: An In Vitro Study.

Regenerative endodontics (REP) is a new clinical modality aiming to regenerate damaged soft and hard dental tissues, allowing for root completion in young adults' teeth. Effective disinfection is crucial for REP success, but commonly used antimicrobials often harm the niche dental pulp stem cells (DPSCs). To our knowledge, this is the first study to explore the biocompatibility and antimicrobial potential of pectin as a potential natural intracanal medicament for REPs. Low methoxyl commercial citrus pectin (LM) (pectin CU701, Herbstreith&Fox.de) was used in all experiments. The pectin's antibacterial activity against single species biofilms (E. faecalis and F. nucleatum) was assessed using growth curves. The pectin's antimicrobial effect against mature dual-species biofilm was also evaluated using confocal laser scanning microscopy (CLSM) after 30 min and 7 days of treatment. The DPSC biocompatibility with 2% and 4% w/v of the pectin coatings was evaluated using live/dead staining, LDH, and WST-1 assays. Pectin showed a concentration-dependent inhibitory effect against single-species biofilms (E. faecalis and F. nucleatum) but failed to disrupt dual-species biofilm. Pectin at 2% w/v concentration proved to be biocompatible with the HDPSCs. However, 4% w/v pectin reduced both the viability and proliferation of the DPSCs. Low concentration (2% w/v) pectin was biocompatible with the DPSCs and showed an antimicrobial effect against single-species biofilms. This suggests the potential for using pectin as an injectable hydrogel for clinical applications in regenerative endodontics.

Improving the Antimicrobial Potency of Berberine for Endodontic Canal Irrigation Using Polymeric Nanoparticles.

To address the challenges posed by biofilm presence and achieve a substantial reduction in bacterial load within root canals during endodontic treatment, various irrigants, including nanoparticle suspensions, have been recommended. Berberine (BBR), a natural alkaloid derived from various plants, has demonstrated potential applications in dentistry treatments due to its prominent antimicrobial, anti-inflammatory, and antioxidant properties. This study aimed to produce and characterize a novel polymeric nanoparticle of poly (lactic-co-glycolic acid) (PLGA) loaded with berberine and evaluate its antimicrobial activity against relevant endodontic pathogens, Enterococcus faecalis, and Candida albicans. Additionally, its cytocompatibility using gingival fibroblasts was assessed. The polymeric nanoparticle was prepared by the nanoprecipitation method. Physicochemical characterization revealed spheric nanoparticles around 140 nm with ca, -6 mV of surface charge, which was unaffected by the presence of BBR. The alkaloid was successfully incorporated at an encapsulation efficiency of 77% and the designed nanoparticles were stable upon 20 weeks of storage at 4 °C and 25 °C. Free BBR reduced planktonic growth at ≥125 μg/mL. Upon incorporation into PLGA nanoparticles, 20 μg/mL of [BBR]-loaded nanoparticles lead to a significant reduction, after 1 h of contact, of both planktonic bacteria and yeast. Sessile cells within biofilms were also considered. At 30 and 40 μg/mL, [BBR]-loaded PLGA nanoparticles reduced the viability of the sessile endodontic bacteria, upon 24 h of exposure. The cytotoxicity of BBR-loaded nanoparticles to oral fibroblasts was negligible. The novel berberine-loaded polymeric nanoparticles hold potential as a promising supplementary approach in the treatment of endodontic infections.

Metatranscriptome and Resistome of the Endodontic Microbiome

IntroductionIn this study, we used metatranscriptomics for the first time to investigate microbial composition, functional signatures, and antimicrobial resistance gene expression in endodontic infections. MethodsRoot canal samples were collected from ten teeth, including five primary and five persistent/secondary endodontic infections. RNA from endodontic samples was extracted, and RNA sequencing was performed on a NovaSeq6000 system (Illumina). Taxonomic analysis was performed using the Kraken2 bacterial database. Then, sequences with a taxonomic classification were annotated against the Universal Protein Knowledgebase for functional annotation and the Comprehensive Antibiotic Resistance Database for AR-like gene identification. ResultsProteobacteria, Bacteroidetes, Firmicutes, and Actinobacteria represented the dominant phyla, whereas Fusobacteria, Spirochetes, and Synergistetes were among the nondominant phyla. The top ten species were mainly represented by obligate (or quasiobligate) anaerobes, including Gram-negative (eg, Capnocytophaga sp. oral taxon 323, Fusobacterium nucleatum, Prevotella intermedia, Prevotella oris, Tannerella forsythia, and Tannerella sp. oral taxon HOT−286) and Gram-positive species (eg, Olsenella uli and Parvimonas micra). Transcripts encoding moonlighting proteins (eg, glycolytic proteins, translational elongation factors, chaperonin, and heat shock proteins) were highly expressed, potentially affecting bacterial adhesion, biofilm formation, host defense evasion, and inflammation induction. Endodontic bacteria expressed genes conferring resistance to antibiotic classes commonly used in dentistry, with a high prevalence and expression of tetracycline and lincosamide resistance genes. Antibiotic efflux and antibiotic target alteration/protection were the main resistance mechanisms. ConclusionsMetatranscriptomics revealed the activity of potential endodontic pathogens, which expressed putative virulence factors and a wide diversity of genes potentially involved in AR.

Estimation of MBC: MIC Ratio of Herbal Extracts against Common Endodontic Pathogens

ABSTRACT Herbal extracts have evoked interest owing to the small number of terpenoids and phenolic compounds, which impart antimicrobial, antioxidant, and anti-inflammatory properties. The minimum inhibitory concentration (MIC), minimum bactericidal concentration (MBC), and minimum fungicidal concentration (MFC) of four herbal extracts (lemon grass oil, basil oil, peppermint oil, and Obicure tea extract) against endodontic pathogens along with the MIC: MBC/MFC ratio were evaluated. The antimicrobial activity by detecting the MIC of three essential oils and tea extract was evaluated against eight common endodontic pathogens by the broth dilution method, while MBC was detected by subculturing onto blood agar from the first –three to five tubes from the MIC dilution tubes (showing no turbidity), which were plated on blood agar. All herbal extracts proved to be effective antimicrobials against tested endodontic pathogens. Basil oil had a bacteriostatic effect on all the organisms (P &lt; 0.05). Mint oil showed bacteriostatic activity on Enterococcus (E.) faecalis and Peptostreptococcus (P &gt; 0.05). Tea extract had a bacteriostatic effect (P &gt; 0.05) against all tested microbes except Actinomyces, Lactobacilli, Staphylococcus (S.) aureus, and Fusobacterium (F.) nucleatum. Lemon grass oil had a bactericidal effect against all the organisms and a bacteriostatic effect against Peptostreptococcus (P &gt; 0.05). It can be concluded that basil oil showed a strong bactericidal effect on the test organisms. The MIC for the organisms ranged from 0.2 to 50 μg/ml.

Novel bioactive nanospheres show effective antibacterial effect against multiple endodontic pathogens

AimThe current study evaluated the antibacterial activity of a newly developed quaternary ammonium polymethacrylate (QAPM)-containing bioactive glasses (BGs) via a two-step method by our group, namely BGs-HAEMB, and explored its cytotoxicity and biocompatibility. MethodsThe antibacterial effects of the BGs-HAEMB against planktonic bacteria, bacterial biofilm formation, and experimental root canal biofilms of persistent pathogens (Enterococcus faecalis, Streptococcus sanguis and Porphyromonas endodontalis) associated with endodontic infection were evaluated in vitro by agar diffusion tests, direct contact tests and live/dead staining. The cytotoxicity and biocompatibility of BGs-HAEMB were evaluated by CCK-8 assays in vitro and a skin implantation model in vivo. ResultsCompared to three clinically used endodontic sealers (Endofill, AH Plus, and iRoot SP), BGs-HAEMB exhibited the relatively strongest antibacterial effect against E. faecalis, S. sanguis and P. endodontalis after sitting for 14 and 28 days (P < 0.01). SEM images and CLSM images also showed that for each tested bacteria, BGs-HAEMB killed the most microorganism among all the experimental groups, regardless of treatment for 7 days or 28 days (P < 0.05). Besides, the BGs-HAEMB-treated groups showed a relatively low cytotoxicity (RGRs ranging from 88.6% to 102.9%) after 1, 3, and 7 days of exposure. Meanwhile, after 28 days of implantation, the inflammatory grade in BGs-HAEMB treated group was assessed as Grade I, in which the average numbers of inflammatory cells (6.7 ± 2.1) were less than 25. ConclusionsBGs-HAEMB exerted a long-term and stable antibacterial effect. The remarkable biocompatibility of BGs-HAEMB in vitro and in vivo confirmed its possible clinical application as a potential alternative in the development of the next generation of endodontic sealers.

Leaf Extracts of Moringa oleifera Cultivated in Baghdad: Characterization and Antimicrobial Potential against Endodontic Pathogens.

The use of medicinal plant preparations to clean and disinfect root canal infection is gaining popularity. The aim of this study was to evaluate the bioactive composition of leaf extracts of Moringa oleifera plants cultivated in Iraq (specifically Baghdad) and their antimicrobial activity against selected root canal pathogens for potential application in endodontic treatment. Materials and Methods. Moringa leaf extracts were prepared either through cold maceration or warm digestion techniques to perform an ethanolic or aqueous extraction, respectively. Phytochemical detection was performed before thin layer chromatography (TLC) and high-performance liquid chromatography (HPLC) to measure flavonoids and phenolic compounds within both extracts. Then, their antimicrobial activities were investigated against Streptococcus mutans, Enterococcus faecalis, and Candida albicans through minimal inhibitory concentration (MIC), minimal bactericidal concentration (MBC), and agar well diffusion assay in comparison to NaOCl and Ca(OH)2. Results. Phytochemical screening showed several active ingredients but with higher expression of flavonoids and phenolic compounds. Also, different types of these compounds were detected through TLC and quantified by HPLC. MIC values for ethanolic extract against Streptococcus mutans, Enterococcus faecalis, and Candida albicans were 60, 65, and 55, respectively, while for aqueous extract, MIC values were 70, 80, and 50, respectively. Aqueous extract showed a higher inhibition zone than ethanolic extract for both Streptococcus mutans and Enterococcus faecalis with a statistically significant difference (p ≤ 0.001) for all tested materials except with NaOCl and Ca(OH)2 in Streptococcus mutans and Enterococcus faecalis, respectively. The ethanolic extract showed a higher inhibition zone against Candida albicans, with a statistically significant difference (p ≤ 0.001) for all tested materials. Conclusion. Ethanolic and aqueous extracts of Moringa oleifera leaves cultivated in Baghdad contain considerable quantities of phytochemicals, especially flavonoid and phenolic compounds, and demonstrated antimicrobial activities against selected endodontic pathogens. Therefore, Moringa leaf extracts could be suggested as an alternative antimicrobial material in endodontic treatment.

Comparison of the Antibacterial Effect of AH26, Adseal and Beta RCS Root Canal Sealers against Enterococcus Faecalis, an in Vitro Study.

Objectives: Antibacterial activity against endodontic pathogens is a desirable feature for root canal sealers. The objective of this study was to compare the antibacterial effect of three resin-based endodontic sealers (AH26, Adseal, and Beta RCS) against Enterococcus faecalis in vitro. Materials and Methods: The antibacterial properties of the sealers were assessed against E. faecalis using agar diffusion test (ADT) for fresh state (N=10) and direct contact test (DCT) for freshly-mixed and set states of the materials (N=10). In ADT, the diameter of the zones of inhibition was measured after 24h of contact. In DCT, the colony-forming units of the bacteria were counted after 30 minutes and 180 minutes of exposure. The results were analyzed with two-way ANOVA and independent sample t-test. P<0.05 was considered significant. Results: Regarding DCT results, all test materials indicated an antibacterial effect, both in freshly-mixed and set states. The highest antibacterial effect was related to Adseal, whereas the lowest was observed in Beta RCS. There was a significant difference between all study groups (different sealers, setting states, and contact times; P<0.001), except for freshly-mixed AH26 and Adseal at 180 minutes (P>0.05). According to ADT, AH26 and Adseal represented the widest and the smallest inhibition zones, respectively (P<0.001). Conclusion: Within the limitations of this in vitro study, AH26, Adseal, and Beta RCS showed antibacterial effects against E. faecalis in both freshly-mixed and set states. The antibacterial effect increased over time in all of the studied sealers.

Targeting the Enterococcal Fibronectin Binding Protein- A of Enterococcus faecalis with Bioactive Compounds from Aegle Marmelos: An In-vitro and In-silico Pilot Study

Introduction: Enterococcus faecalis (E. faecalis) is a significant endodontic pathogen, and its virulence has been shown to be significantly influenced by Enterococcal fibronectin binding protein-A (EfbA). Targeting this virulent protein using alternative strategies would be a novel idea to combat the complications of E. faecalis in dental healthcare settings. Thus, the rationale of this investigation is to identify potent bioactive compounds from Aegle marmelos for their antimicrobial properties against EfbA of E. faecalis. Aim: To assess the frequency of EfbA among clinical isolates of E. faecalis and evaluate the antibacterial activity of essential bioactive compounds from Aegle marmelos. Materials and Methods: An in-vitro and in-silico pilot study was conducted from April 2022 to June 2022 in the Department of Microbiology, Saveetha Dental College and Hospitals, Chennai, Tamil Nadu, India to detect the frequency of EfbA in E. faecalis. Patients with typical root caries were included. Microbiological processing of 20 root caries samples was performed to characterise E. faecalis and detect the EfbA gene using Polymerase Chain Reaction (PCR) amplification. Crude methanol extract was obtained from A. marmelos and assessed for its antimicrobial effect against the isolated strains of E. faecalis. Furthermore, several biomolecules from A. marmelos were evaluated for their inhibitory properties through in-silico docking analysis. Results: E. faecalis was identified in 12 (60%) out of 20 root caries samples, and three isolates (25%) were identified as Multidrug-Resistant (MDR) strains based on the antibiogram susceptibility profile, showing resistance to more than three different tested antibiotic groups. EfbA was detected in two of the three MDR strains (66.7%). The crude extract of A. marmelos exhibited promising antibacterial activity. In-silico analysis of the essential oil compounds from A. marmelos revealed that Aegeline had a high interaction with low docking energy and a high number of hydrogen bonds. Conclusion: The current study highlights the potential of aegeline from A. marmelos as an antibacterial agent against resistant strains of E. faecalis. However, additional in-vivo research must be conducted to experimentally validate these findings.

Bifidobacterium animalis subsp. lactis HN019 has antimicrobial activity against endodontic pathogens in vitro.

The aim of the present study was to evaluate, in vitro, the antimicrobial activity of the probiotic Bifidobacterium animalis subsp. lactis HN019, through the well technique, against 10 microorganisms can be found involved in endodontic infections. The antimicrobial activity of the probiotic was performed on Streptococcus mutans, Streptococcus sobrinus, Lacticaseibacillus casei, Enterococcus faecalis, Staphylococcus aureus, Candida albicans, Porphyromonas gingivalis, Porphyromonas endodontalis,Fusobacterium nucleatumandPrevotella intermedia. For the control group, it was used non-pathogenic bacteria Escherichia coli, Saccharomyces cerevisiae, and Kocuria rizhopilla. After 48 to 72 h of incubation of the petri dishes containing the culture medium, the microorganism strains, and the probiotic, the plates were examined to assess the uniformity of microbial growth, presence of contaminants, and the halo of inhibition. After visual inspection, the reading of the halo of inhibition was performed with the aid of a digital caliper using a reflected light source to illuminate the inverted plate on a black, opaque background after removing the cap. Thus, 3 values were obtained from each bacterial inoculum, which were added and divided by three to obtain the average of the values. The results of the in vitro study demonstrated that the probiotic B. animalis subsp. lactis HN019 promoted the inhibition of all strains of the pathogens evaluated, with the exception of Candida albicans, demonstrating antimicrobial activity on these microorganisms.

Comparative Evaluation of the Physical and Antimicrobial Properties of Mineral Trioxide Aggregate, Biodentine, and a Modified Fast-Setting Mineral Trioxide Aggregate Without Tricalcium Aluminate: An In Vitro Study.

Background Tricalcium aluminate, one of the major constituents of mineral trioxide aggregate (MTA), has been shown to have cytotoxic properties. Mineral trioxide aggregate has moderate to low antimicrobial activity against the most common endodontic pathogen, Enterococcus faecalis. Aim To assess the physical and antimicrobial properties of a newly modified formulation of mineral trioxide aggregate. Materials & methods The final setting time, compressive strength, and antimicrobial properties were tested for three groups of materials. The material used for Group 1 was mineral trioxide aggregate (white MTA, Angelus, Londrina, Brazil); the material for Group 2 was Biodentine (Septodont, Saint Maur des Fossés, France); and for Group 3, a modified MTA formulation was used. Results Group 1 had the longest setting time, and Group 2 had the shortest setting time. Group 3's material was set at 83.65 ± 0.28 minutes. This difference among the groups was statistically significant (p < 0.05). The highest mean compressive strength during all the time periods was seen in Group 2, followed by Group 3, and the least in Group 1. This difference in compressive strength was statistically significant (p=0.001). The largest zone of inhibition against Enterococcus faecalis, Candida albicans, and Streptococcus mutans was seen in Group 3, followed by Group 2 and Group 1. Conclusion Under the limitations of the present study, the newly modified MTA could serve as an alternative to the conventional MTA in terms of faster setting, higher strength, and better antimicrobial properties.

Intermembrane rhomboid protease activity of Rothia mucilaginosa

Rothia mucilaginosa is a part of the normal oral microflora, which has been recognized as an endodontic pathogen and is also an emerging opportunistic pathogen in immune compromised hosts. In this study, we identified the rhomboid protease in R. mucilaginosa as a candidate virulence factor. A database search identified two rhomboid genes on the genome with open reading frames of 828 bp and 822 bp, respectively. The two genes are separated by 57 bp, suggesting that they constitute a bicistronic operon. Motifs conserved among rhomboid family proteins (HxxxN, GxSG, and GxxxG) were found in their deduced amino acid sequences. The two recombinant rhomboids and the rhomboid/liposome complexes (proteoliposomes) were prepared using a wheatgerm cell-free translation system, and their serine protease activities were assessed. The two rhomboids showed serine protease activity in the form of rhomboid/liposome complex, while the recombinant rhomboids alone showed little or no activity. On the supposition of host invasion, R. mucilaginosa was cultivated in medium with type I collagen, and the rhomboid mRNA levels were examined. Real-time reverse transcription polymerase chain reaction analysis revealed that the levels of the two rhomboid mRNAs increased slightly in culture with type I collagen, although the changes were not statistically significant. These results suggested that R. mucilaginosa possesses two active rhomboids with protease activity on the membrane, and the two rhomboids may be virulence factors involved in the response to host environmental conditions.

The potential of reuterin derived from Indonesian strain of Lactobacillus reuteri against endodontic pathogen biofilms in vitro and ex vivo

ObjectivesDespite the use of common irrigating solution with antimicrobial properties, failed root canal treatment remains a significant problem in endodontics. In the present study, we examined the efficacy of reuterin derived from probiotic bacteria, Lactobacillus reuteri on the biofilms of major endodontic pathogens using ex vivo model of root canal infections. MethodsBiofilms of major endodontic pathogens namely Enteoroccus faecalis, Fusobacterim nucleatum, Porphyromonas gingivalis, and Candida albicans were formed on root canals of 60 human premolar tooth samples accordingly a standard protocol. Thereafter, teeth were treated with either 2.5 % NaOCl (positive control), various concentrations of reuterin (test-group) or sterilized-distilled water (negative control) in a time-dependent assay. The efficacy of irrigation was evaluated by a time-dependent assay at 5 min and 30 min after irrigation by colony-forming units assay. The findings were further confirmed by species-specific real-time PCR. Data were statistically analysed using one way ANOVA with a significance level of P < 0.05. ResultsReuterin isolated from L. reuteri was effective against E. faecalis, C. albicans, F. nucleatum, and P. gingivalis biofilms, with a concentration of 100 µg/mL being the most effective compared to the negative control (P < 0.05) and also showed similar efficacy when compared with NaOCl. ConclusionReuterin isolated from L. reuteri has ability to inhibit in vitro and ex-vivo biofilms of endodontic pathogens, namely E. faecalis, F. nucleatum, P. gingivalis, and C. albicans. Reuterin has potential as a root canal irrigating solution due to its antibiofilm activity. Further research is warranted to determine the potential of probiotic bacteriotherapy in root canal systems.

Teeth withacute apical abscess vs. teeth withchronic apical periodontitis: a quantitative and qualitative proteomic analysis.

To quantitatively and qualitatively analyze the proteomic profile ofteeth with acute apical abscesses (AAA) compared with teeth withchronic apical periodontitis (CAP) and to correlate the expression of detected human proteins with their main biological functions. Samples were obtained from root canals of9 patients diagnosed with AAA and 9 with CAP. Samples were analyzed by reversed-phase liquid chromatography coupled to mass spectrometry. Label-free quantitative proteomic analysis was performed by Protein Lynx Global Service software. Differences in protein expression were calculated using the t-test (p < 0.05). In total, 246 human proteins were identified from all samples. Proteins exclusively found in the AAA group were mainly associated with the immunoinflammatory response and oxidative stress response. In the quantitative analysis, 17 proteins were upregulated (p < 0.05) in the AAA group, including alpha-1-acid glycoprotein, hemopexin, fibrinogen gamma chain, and immunoglobulin. Additionally, 61 proteins were downregulated (p < 0.05), comprising cathepsin G, moesin, gelsolin, and transketolase. Most of the proteins were from the extracellular matrix, cytoplasm, and nucleus. The common proteins between the groups were mainly associated with the immune response at both expression levels. Upregulated proteins mostly belonged to the acute-phase proteins, while the downregulated proteins were associated with DNA/RNA regulation and repair, and structural function. The host response is directly related to the development of apical abscesses. Thus, understanding the behavior of human proteins against the endodontic pathogens involved in this condition might contribute to the study of new approaches related to the treatment of this disease.

Antagonistic interaction between two key endodontic pathogens Enterococcus faecalis and Fusobacterium nucleatum

ABSTRACT Background Endodontic infections are known to be caused by pathogenic bacteria. Numerous previous studies found that both Fusobacterium nucleatum and Enterococcus faecalis are associated with endodontic infections, with Fusobacterium nucleatum more abundant in primary infection while Enterococcus faecalis more abundant in secondary infection. Little is known about the potential interactions between different endodontic pathogens. Objective This study aims to investigate the potential interaction between F. nucleatum and E. faecalis via phenotypical and genetic approaches. Methods Physical and physiological interactions of F. nucleatum and E. faecalis under both planktonic and biofilm conditions were measured with co-aggregation and competition assays. The mechanisms behind these interactions were revealed with genetic screening and biochemical measurements. Results E. faecalis was found to physically bind to F. nucleatum under both in vitro planktonic and biofilm conditions, and this interaction requires F. nucleatum fap2, a galactose-inhibitable adhesin-encoding gene. Under our experimental conditions, E. faecalis exhibits a strong killing ability against F. nucleatum by generating an acidic micro-environment and producing hydrogen peroxide (H2O2). Finally, the binding and killing capacities of E. faecalis were found to be necessary to invade and dominate a pre-established in vitro F. nucleatum biofilm. Conclusions This study reveals multifaceted mechanisms underlying the physical binding and antagonistic interaction between F. nucleatum and E. faecalis, which could play a potential role in the shift of microbial composition in primary and secondary endodontic infections.

The Potential of Phenothiazines against Endodontic Pathogens: A Focus on Enterococcus-Candida Dual-Species Biofilm.

Persistent apical periodontitis occurs when the endodontic treatment fails to eradicate the intraradicular infection, and is mainly caused by Gram-positive bacteria and yeasts, such as Enterococcus faecalis and Candida albicans, respectively. Phenothiazines have been described as potential antimicrobials against bacteria and fungi. This study aimed to investigate the antimicrobial potential of promethazine (PMZ) and chlorpromazine (CPZ) against E. faecalis and C. albicans dual-species biofilms. The susceptibility of planktonic cells to phenothiazines, chlorhexidine (CHX) and sodium hypochlorite (NaOCl) was initially analyzed by broth microdilution. Interaction between phenothiazines and CHX was examined by chequerboard assay. The effect of NaOCl, PMZ, CPZ, CHX, PMZ + CHX, and CPZ + CHX on biofilms was investigated by susceptibility assays, biochemical and morphological analyses. Results were evaluated through one-way ANOVA and Tukey's multiple comparison post-test. PMZ, alone or in combination with irrigants, was the most efficient phenothiazine, capable of reducing cell counts, biomass, biovolume, carbohydrate and protein contents of dual-species biofilms. Neither PMZ nor CPZ increased the antimicrobial activity of CHX. Further investigations of the properties of phenothiazines should be performed to encourage their use in endodontic clinical practice.

Determining the potential use of biosurfactants in preventing endodontic infections.

Microbial biofilms play a dominant role in the failure of endodontic therapies. Bacterial adhesion is the first step in the establishment of biofilms, activating the host immune response leading to tissue damage. Biosurfactants are microbe-derived tensioactive molecules with latent anti-adhesive and anti-microbial activity. This study reports the extraction and characterization of a biosurfactant from Lactobacillus (L.) plantarum (Lp-BS) and investigates its anti-microbial and anti-adhesive properties compared to rhamnolipid, a commercially available biosurfactant. Lp-BS, extracted from L. plantarum during the growth phase, was characterized as a glycoprotein, able to reduce surface tension and emulsify non-polar liquids. Proteomic analysis of Lp-BS identified three bacterial adhesin-like proteins, suggesting roles in hindering bacterial adhesion. Lp-BS did not show significant anti-microbial activity against endodontic pathogens from the Streptococcus (Strep.) anginosus group or Enterococcus (Ent.) faecalis at 50mg/ml. However, anti-adhesive activity on abiotic surfaces was observed against both Strep. anginosus and Strep. intermedius. Rhamnolipid exhibited strong anti-microbial activity, with minimum inhibitory concentrations of 0.097mg/ml against Strep. anginosus, and 0.048mg/ml against Strep. constellatus and Strep. intermedius, in addition to a marked anti-adhesive activity. These findings offer preliminary evidence for the potential application of biosurfactants as an anti-microbial and/or anti-adhesive pharmacotherapy in endodontics.

Evaluation of the antibacterial activity of Lactobacilli probiotics supernatants against Enterococcus faecalis (in-vitro study)

BackgroundThere is an increasing demand to find a potent antibacterial agent against endodontic pathogens with the least toxic effect. The study aimed to evaluate the antibacterial activity of Lactobacilli probiotics on Enterococcus faecalis (E. faecalis) in comparison to calcium hydroxide paste.MethodsThe study involved two stages; (stage one): determination of the antibacterial properties of three strains of Lactobacilli probiotics supernatants (PS); Lactobacillus plantarum, Lactobacillus rhamnosus, Lactobacillus acidophilus, and a cocktail mix of the three supernatants against E. faecalis using agar well diffusion method in both solution and gel phase. The formed zones of inhibition (ZOI) were measured in millimeters (mm) and compared to each other. PS solution and gel of the largest ZOI were further compared with calcium hydroxide paste (35% Ultra Cal XS Ca(OH)2) This was followed by (stage two): assessment of the minimum inhibitory concentration (MIC) of the PS that showed the largest ZOI against E. faecalis by agar well diffusion assay for both PS solution and gel.ResultsAll supernatants showed growth inhibition against E. faecalis, and the cocktail mix showed the largest ZOI. However, no significant difference was found between the supernatants in both the solution and gel phases (p > 0.05). Ca(OH)2 showed a significantly lower effect than both the cocktail mix solution and gel (p < 0.05). The MIC of the cocktail mix solution and gel against E. faecalis was 50 mg/ml. (p > 0.05).ConclusionPS has an antibacterial effect on E. faecalis and was more effective than Ca(OH)2.Lactobacilli probiotics could be a promising antibacterial agent used as an irrigant or an intracanal medication.