CLSM-Guided Imaging for Quantifying Endodontic Disinfection.

Elimination of microbes in the root canal system is crucial for achieving long-term success in endodontic treatment. Further efforts in study design and standardization are needed in order to improve the validity and comparability of in vitro results on endodontic disinfection procedures, in turn improving clinical outcomes. This study optimizes two models at all steps: tooth selection, pretreatment, inoculation method (by growth or centrifugation), and confocal laser scanning microscopy (CLSM)-guided imaging of LIVE/DEAD-stained specimens. Individual anatomical conditions lead to substantial differences in penetration depth. Sclerosis grading (SCG), a classification system introduced in this study, provides information about the sclerosis status of the dentine and is helpful for careful, specific, and comparable tooth selection in in vitro studies. Sonically activated EDTA for the pretreatment of roots, inoculation of Enterococcus faecalis in an overflow model, 3-4 weeks of incubation, as well as polishing of dentine slices before staining, led to advances in the visualization of bacterial penetration and irrigation depths. In contrast, NaOCl pretreatment negatively affected performance reproducibility and should be avoided in any pretreatment. Nonsclerotized teeth (SCG0) can be used for microbial semilunar-shaped inoculation by centrifugation as a "quick-and-dirty" model for initial orientation. In conclusion, CLSM-guided imaging for quantifying endodontic infection/disinfection is a very powerful method after the fine-tuning of materials and methods.

Isolation, identification, and significance of salivary Veillonella spp., Prevotella spp., and Prevotella salivae in patients with inflammatory bowel disease.

The global prevalence of inflammatory bowel disease (IBD) is on the rise, prompting significant attention from researchers worldwide. IBD entails chronic inflammatory disorders of the intestinal tract, characterized by alternating flares and remissions. Through high-throughput sequencing, numerous studies have unveiled a potential microbial signature for IBD patients showing intestinal enrichment of oral-associated bacteria. Simultaneously, the oral microbiome can be perturbed by intestinal inflammation. Our prior investigation, based on 16S rRNA amplicon sequencing, underscored elevated abundance of Veillonella spp. and Prevotella spp. in the salivary microbiomes of IBD patients. Noteworthy, Prevotella salivae emerged as a distinct species significantly associated with IBD. P. salivae is an under-recognized pathogen that was found to play a role in both oral and systemic diseases. In this study, we delve deeper into the salivary microbiomes of both IBD patients and healthy controls. Employing diverse cultivation techniques and real-time quantitative polymerase chain reactions (RT-qPCR), we gauged the prevalence and abundance of Veillonella spp., Prevotella spp., and P. salivae. Our isolation efforts yielded 407 and 168 strains of Veillonella spp., as well as 173 and 90 strains of Prevotella spp., from the saliva samples of IBD patients and healthy controls, respectively. Veillonella-vancomycin agar emerged as the discerning choice for optimal Veillonella spp. cultivation, while Schaedler kanamycin-vancomycin agar proved to be the most suitable medium for cultivating Prevotella spp. strains. Comparing our RT-qPCR findings to the previous 16S rRNA amplicon sequencing data, the results corroborated the higher abundance of Veillonella spp., Prevotella spp., and P. salivae in the saliva of IBD patients compared to healthy controls. However, it's worth noting that in contrast to RT-qPCR, the 16S rRNA amplicon sequencing data revealed greater absolute abundance of all three bacterial groups in both IBD patients and controls.

Bacteremia Prevention during Periodontal Treatment-An In Vivo Feasibility Study.

The link between periodontitis and systemic diseases has increasingly become a focus of research in recent years. In this context, it is reasonable-especially in vulnerable patient groups-to minimize bacteremia during periodontal treatment. The aim of the present in vivo feasibility study was to investigate the possibility of laser-based bacteremia prevention. Patients with stage III, grade B generalized periodontitis were therefore treated in a split-mouth design either with prior 445 nm laser irradiation before nonsurgical periodontal therapy or without. During the treatments, clinical (periodontal measures, pain sensation, and body temperature), microbiological (sulcus samples and blood cultures before, 25 min after the start, and 10 min after the end of treatment), and immunological parameters (CRP, IL-6, and TNF-α) were obtained. It was shown that periodontal treatment-related bacteremia was detectable in both patients with the study design used. The species isolated were Schaalia georgiae, Granulicatella adiacens, and Parvimonas micra. The immunological parameters increased only slightly and occasionally. In the laser-assisted treatments, all blood cultures remained negative, demonstrating treatment-related bacteremia prevention. Within the limitations of this feasibility study, it can be concluded that prior laser disinfection can reduce bacteremia risk during periodontal therapy. Follow-up studies with larger patient numbers are needed to further investigate this effect, using the study design presented here.

Prevalence and Phylogenetic Analysis of Lipoprotein-Gene ragB-1 of Porphyromonas gingivalis-A Pilot Study.

Porphyromonas gingivalis (P.g.) is a key pathogen involved in periodontal diseases. The aim of this study was to investigate the prevalence and phylogenetic origin of the lipoprotein-gene ragB in its most virulent variant, ragB-1 (co-transcribed with ragA-1 as locus rag-1), in different P.g. strains collected worldwide. A total of 138 P.g. strains were analyzed for the presence of ragB-1 by pooled analysis and subsequently individual PCRs. Sequencing a core fragment of ragB-1 of the individual strains made it possible to carry out a phylogenetic classification using sequence alignment. In total, 22 of the 138 P.g. strains tested positive for ragB-1, corresponding to a prevalence of 16%. The fragment investigated was highly conserved, with variations in the base sequence detected in only three strains (OMI 1072, OMI 1081, and OMI 1074). In two strains, namely OMI 1072 (original name: I-433) and OMI 1081 (original name: I-372), which originate from monkeys, two amino-acid alterations were apparent. Since ragB-1 has also been found in animal strains, it may be concluded that rag-1 was transferred from animals to humans and that this originally virulent variant was weakened by mutations over time so that new, less virulent, adapted commensal versions of rag (rag-2, -3, and -4), with P.g. as the host, evolved.

Exploring the genetic and functional diversity of Porphyromonas gingivalis long fimbriae.

Porphyromonas gingivalis is a key pathobiont in periodontitis. Its long fimbriae consist of a single anchor (FimB), a varying number of stalk (FimA), and three accessory (tip-related) proteins (FimC, FimD, and FimE). Based on 133 strains/genomes available, it was our aim to investigate the diversity within FimA and FimB and explain the variety of long fimbriae (super-)structures. Combining the new forward primer fimAnewF with the established fimAunivR, we were able to amplify and sequence fimA including its leader region covering all genotypes and serotypes for phylogenetic analysis. We designed two primer pairs sensing the presence of an internal stop codon in fimB with an impact on fimbrial length. Finally, we examined fimbrial secondary structures by transmission electron microscopy (TEM) and scanning electron microscopy (SEM). The phylogeny of fimA/FimA revealed two new subtypes (IIa and IIb) with specific changes in functional domains and thus adding to the current classification scheme (I, Ib, and II-V). Regarding evolution, we confirm that Porphyromonas gulae fimA-type A is closely related to human P. gingivalis strains of cluster Ib and might be its ancestor genotype. A fimB internal stop codon is rare and was found in ATCC 33277 only. Comparing P. gingivalis TEM/SEM pictures of type I ATCC 33277 with type V OMI622 revealed a broad spectrum of fimbrial structures including bundling, cell-cell knotting, and brick-wall formation. In conclusion, FimA forms more distinct subtypes than previously known. The bundling of long fimbriae, a mechanism known from EPEC/EHEC and Salmonella, is proposed and supported by TEM/SEM pictures for the first time here. The role and variations of terminal accessory FimC-E in superstructure formation and/or (co-) adhesion should be investigated more closely next.

Effect of Er,Cr:YSGG laser with a side-firing tip on decontamination of titanium disc surface: an in vitro and in vivo study.

PURPOSE: To evaluate the effectiveness of an erbium, chromium:yttrium-scandium-gallium-garnet (Er,Cr:YSGG) laser with side-firing tip in decontamination of titanium (Ti) disc. METHODS: In the first test series, 29 Ti-discs were contaminated with Staphylococcus aureus and treated as follows: positive control (no treatment); Perioflow; Laser A (0.75 W, 100 Hz), Laser B (1.5 W, 30 Hz); Laser C (no radiation, 60% water); and Laser D (no radiation, 50% water). For bacterial quantification, colony forming units (CFU, vital cells only) and quantitative PCR (qPCR, vital and devital cells) were performed. In a second test series, 92 Ti-discs were used, contaminated with in vivo-grown biofilm and treated as follows: positive control (no treatment); Perioflow; Laser E (1.5 W, 30 Hz), and Laser F (no radiation, 50% water). Considering the different and unknown culture conditions, quantification of bacteria was performed by broad-spectrum bacterial qPCR only. Based on the assumption that all cells of an organism contain an equivalent complement of genetic information, genome equivalent (GE) determination ensured the detection of the different intact and semi-intact genomes, regardless of type of bacterial species and vitality, circumvent the inherent bias of cultures. RESULTS: The GE values were significantly reduced by all interventions in both test series, compared to the positive control group (p < 0.001). In the first test series with S. aureus as model organism, Perioflow yielded a lower GE than the Laser groups A-D (all p < 0.025). The number of CFUs was significantly reduced in the intervention groups compared to the positive control (p < 0.001), except for Laser A (p = 0.157) and Laser D (p = 0.393). In the second test series, none of the pairwise comparisons of the intervention conditions showed a significant difference (Perioflow vs. Laser E: p = 0.732; Perioflow vs. Laser F: p = 0.590; Laser E vs. Laser F: p = 0.379). CONCLUSION: The Er,Cr:YSGG laser with side-firing tip and Perioflow were equally capable of effectively decontaminating a Ti-disc surface. It is assumed that the bacterial reduction was largely due to the mechanical effect of the air and water stream.

Evaluation of hydrocortisone as a strain-dependent growth-regulator of Porphyromonasgingivalis.

OBJECTIVE: Porphyromonas gingivalis is an oral key pathogen and known to be very diverse in geno- and phenotypes. It is a fastidious bacterium with low O2-tolerance and 3-7 days of incubation are necessary. With growing interest in the field of microbial endocrinology we explored the potential growth-stimulating effect of hydrocortisone (HC, synonym cortisol) on P. gingivalis cultures. MATERIAL AND METHODS: Six different P. gingivalis strains were pre-incubated in supplemented Brain-Heart-Infusion broth under appropriate conditions for 24 h, diluted and transferred into microplates. A newly developed and semi-automated spectrophotometric measurement in triplicate, applying a SpectraMax i3x microplate reader at an optical density of 600 nm, was conducted to test growth differences between test group (exposed to a supplement of either 1.25, 2.5, 5, 10, or 20 µg/ml of hydrocortisone) and control group over 48 h of anaerobic incubation (O2 ≤ 1%). Furthermore, strains were also incubated on HC-supplemented blood agar to test for a possible growth-stimulating effect on solid media. RESULTS: HC significantly stimulated the lag-phase growth of four out of six P. gingivalis strains. Our data suggest a concentration-dependent growth stimulatory effect of HC between 2.5 and 5 µg/ml, while below 1.25 µg/ml and above 10 µg/ml HC either did not stimulate or inhibited growth. CONCLUSIONS: HC could reduce the incubation time when isolating P. gingivalis from clinical samples and could boost low biomass cultivations especially during their lag-phase. The growth-modulating effect might be via modulation of virulence factors/quorum sensing gene expression or by reactive oxygen species(ROS)-capturing during early stages of bacterial growth. Further experiments are necessary to explain the mechanism behind our observations.

Antimicrobial efficiency and cytocompatibility of different decontamination methods on titanium and zirconium surfaces.

OBJECTIVES: The purpose of this study was to investigate the efficiency of different implant-decontamination methods regarding biofilm modification and potential cytotoxic effects. Therefore, the amount of biofilm reduction, cytocompatibility, and elementary surface alterations were evaluated after decontamination of titanium and zirconium surfaces. MATERIAL AND METHODS: Titanium and zirconium disks were contaminated with a newly developed high-adherence biofilm consisting of six microbial species. Decontaminations were performed using titanium curette, stainless steel ultrasonic scaler (US), glycine (GPAP) and erythritol (EPAP) powder air-polishing, Er:YAG laser, 1% chlorhexidine (CHX), 10% povidone-iodine (PVI), 14% doxycycline (doxy), and 0.95% NaOCl solution. Microbiologic analysis was done using real-time qPCR. For assessment of cytocompatibility, a multiplex assay for the detection of cytotoxicity, viability, and apoptosis on human gingival fibroblasts was performed. X-ray photoelectron spectroscopy (XPS) was used to evaluate chemical alterations on implant surfaces. RESULTS: Compared with untreated control disks, only GPAP, EPAP, US, and Er:YAG laser significantly reduced rRNA counts (activity) on titanium and zirconium (p < .01), whereas NaOCl decreased rRNA count on titanium (p < .01). Genome count (bacterial presence) was significantly reduced by GPAP, EPAP, and US on zirconium only (p < .05). X-ray photoelectron spectroscopy analyses revealed relevant re-exposure of implant surface elements after GPAP, EPAP, and US treatment on both materials, however, not after Er:YAG laser application. Cytocompatibility was impaired by CHX, PVI, doxy, and NaOCl. CHX and PVI resulted in the lowest viability and doxy in the highest apoptosis. CONCLUSIONS: Within the limits of this in vitro study, air-polishing methods and ultrasonic device resulted in effective biofilm inactivation with surface re-exposure and favorable cytocompatibility on titanium and zirconium. Chemical agents, when applied on implant surfaces, may cause potential cytotoxic effects.

High concentrations of Porphyromonas gingivalis-LPS downregulate Tlr4 and modulate phosphorylation of ERK and AKT in murine cementoblasts.

Porphyromonas gingivalis lipopolysaccharide (PG-LPS) is an important virulence factor potentially contributing to periodontal tissue destruction. Toll-like receptor 4 (Tlr4) is a key mediator of NF-kB activation during pathogen recognition. Previous work using Tlr4-specific antibodies demonstrated a partial neutralization of PG-LPS effects on murine cementoblasts, which can affect cell function and regulate gene expression of osteoclastic markers. PG-LPS also potentially influence the inflammation process and the resorption of mineralized tissues. Yet, such inflammatory responses and cell signaling events remain to be characterized at the protein level. We thus investigated the effect of 1 and 10 µg/ml of PG-LPS, respectively, on cell morphology, cell viability, and selected key downstream molecules of the Tlr4 signaling cascade in cementoblasts. High concentrations of PG-LPS (10 µg/ml) significantly reduced cell viability after 48 h. Upon PG-LPS-stimulation, Tlr4 was significantly downregulated. Equally, IκBα, a downstream molecule, was downregulated in terms of phosphorylation and protein production. Furthermore, downstream signaling kinases, like serine/threonine kinase phospho-AKT and the mitogen-activated protein kinase (MAPK)-family, specifically phospho-ERK1/2, were significantly upregulated under high PG-LPS-concentrations. We provide new insights into PG-LPS-triggered intracellular signaling pathways in cementoblasts and thus deliver a basis for further research in PG-mediated periodontal inflammation.

The oral-gut axis: Salivary and fecal microbiome dysbiosis in patients with inflammatory bowel disease.

Inflammatory bowel disease (IBD) is a group of chronic inflammatory disorders that fall into two main categories: Crohn's disease (CD) and ulcerative colitis (UC). The gastrointestinal tract extends from the mouth to the anus and harbors diverse bacterial communities. Several sequencing-based studies have identified an intestinal enrichment of oral-associated bacteria and demonstrated their ability to induce intestinal inflammation in mice, suggesting that intestinal pathobionts originate from the oral cavity, particularly members of the genus Streptococcus. This study aimed to investigate the composition of the salivary and fecal microbiome of IBD patients (n = 14) compared to healthy controls (n = 12) and to determine the abundance of common bacterial taxa in both niches. Metagenomic DNA was extracted from saliva and fecal samples, and the 16S rRNA gene was targeted for sequencing. Our results revealed that the overall microbial composition of saliva was significantly altered in the IBD patients compared to the control subjects (p = 0.038). At the genus level, Veillonella and Prevotella were highly abundant in IBD (median: 25.4% and 22.2%, respectively) compared to the control group (17.9% and 13.4%, respectively). In contrast, Neisseria, Streptococcus, Haemophilus, and Fusobacterium were associated with a healthy gut state. Regarding the fecal microbiome, the IBD group had a significantly higher abundance of Clostridium sensu stricto 1 and Escherichia-Shigella (both comprising pathogenic bacteria) compared with the control group. Members of both bacterial groups have previously been shown to positively correlate with intestinal inflammation and high expression of pro-inflammatory cytokines that disrupt intestinal barrier integrity. In addition, we demonstrate that the increased abundance of Clostridium sensu stricto 1 and Escherichia-Shigella has also been associated with significant upregulation of certain metabolic pathways in the feces of the IBD group, including bacterial invasion of epithelial cells. Streptococcus was the only common genus detected in both the salivary and fecal microbiome and represented the oral-gut axis in our study. Using culture-based methods, we isolated 57 and 91 Streptococcus strains from saliva as well as 40 and 31 strains from fecal samples of the controls and IBD patients, respectively. The phylogenetic tree of streptococci based on sodA sequences revealed several patient-specific clusters comprising salivary and fecal streptococcal isolates from the same patient and belonging to the same species, suggesting that the oral cavity is an endogenous reservoir for intestinal strains.

Impact of sucroferric oxyhydroxide on the oral and intestinal microbiome in hemodialysis patients.

Hyperphosphatemia is a consequence of chronic kidney disease associated with mineral/bone impairment, increased cardiovascular events and mortality. Therapeutically, most dialysis patients have to take phosphate binders. Here, we investigated effects of the Fe(3+)-based phosphate binder sucroferric oxyhydroxide (SFOH) on the oral and gastrointestinal microbiome of 11 hemodialysis patients. Saliva, dental plaque and stool were collected at baseline, one and four weeks of SFOH intake and subjected to 16S rRNA gene (V3-V4 region) directed Illumina MiSeq-based analysis. Total Fe, Fe(2+) and Fe(3+) were determined in stool and saliva. Overall, the microbiome did not change significantly. However, some patient-, sample- and taxon-specific differences were noted, which allowed patients to be divided into those with a shift in their microbiome (6/11) and those without a shift (5/11). Total Fe and Fe(2+) were highest after one week of SFOH, particularly in patients who exhibited a shift in microbiome composition. Eight bacterial taxa showed significant unidirectional changes during treatment. In-depth microbiome analysis revealed that taxa that significantly benefited from iron plethora had no iron-binding siderophores or alternatives, which was in contrast to taxa that significantly declined under iron plethora. Patients with microbiome-shift were significantly younger and had higher serum phosphate concentrations. In conclusion, this study sheds light on the impact of iron on the microbiome of hemodialysis patients.

Impact of Three Nonsurgical, Full-Mouth Periodontal Treatments on Total Bacterial Load and Selected Pathobionts.

For the treatment of periodontitis stage III/IV, a quadrant/week-wise debridement (Q-SRP) was compared with three full-mouth approaches: full-mouth scaling (FMS, accelerated Q-SRP within 24 h), full-mouth scaling with chlorhexidine-based disinfection (FMD), and FMD with adjuvant erythritol air polishing (FMDAP). The objective of this prospective, randomized study (a substudy of ClinicalTrials.gov, identifier: NCT03509233) was to compare the clinical and microbiological effects of the treatments. In total, 105 patients were randomized to one of the four aforementioned treatment groups, with n = 25, 28, 27, and 25 patients allocated to each group, respectively. At baseline and 3 and 6 months after treatment, the clinical parameters, including the pocket probing depths, clinical attachment level, and bleeding on probing, were recorded, and the prevalence of the total bacteria and four periodontal pathobionts (Aggregatibacter actinomycetemcomitans, Porphyromonas gingivalis, Prevotella intermedia, and Tannerella forsythia) was determined using real-time quantitative PCR. Concerning the clinical outcomes, all the treatment modalities were effective, but the full-mouth approaches, especially FMDAP, were slightly superior to Q-SRP. Using the FMD approach, the reduction in the bacterial load and the number of pathobionts was significantly greater than for FMS, followed by Q-SRP. FMDAP was the least effective protocol for microbial reduction. However, after a temporary increase 3 months after therapy using FMDAP, a significant decrease in the key pathogen, P. gingivalis, was observed. These findings were not consistent with the clinical results from the FMDAP group. In conclusion, the dynamics of bacterial colonization do not necessarily correlate with clinical outcomes after full-mouth treatments for periodontitis stage III/IV.

Isoeugenol-functionalized nanogels inhibit peri-implantitis associated bacteria in vitro.

OBJECTIVES: Obligate and facultative anaerobic bacteria adhering to dental implants are a major cause for peri-implant inflammation, which, if left untreated, can lead to implant loss. Previously, our group developed a new route for the synthesis of isoeugenol-functionalized aqueous nanogels for implant coatings. METHODS: Here, the antimicrobial activity of several new nanogels differing in spacer length (n = 6, 9, 44), radius (60-200 nm), and amount of isoeugenol functional substance (1-20 mol%) was tested against the following peri-implantitis-associated species: Fusobacterium nucleatum, Porphyromonas gingivalis, Prevotella intermedia, Aggregatibacter actinomycetemcomitans, Escherichia coli, Actinomyces viscosus, Enterococcus faecalis, Staphylococcus aureus, Streptococcus oralis, S. parasanguinis, and the yeast Candida albicans. The minimal bactericidal concentration (MBC) and fungicidal concentration (MFC) were determined for each combination. In addition, transmission electron microscopy (TEM) and fluorescence microscopy after live-dead-staining (LD-S) were performed to visualize nanogel-microbe interactions. RESULTS: Two nanogels, NG9-3 and NG9-4 (colloids of 80-150 nm, with a spacer length of n = 9 and feeding between 5 and 10 mol% isoeugenol), had an inhibitory effect on all Gram-positive species and on P. gingivalis and P. intermedia with MBC ≥31.25 µg/ml. TEM and LD-S images showed that cellular adhesion and uptake of nanogels resulted in swelling, shedding, or even complete detachment of the cell wall and then to bursting (see graphical abstract). CONCLUSIONS: Functional nanogels can be used as building blocks in the design of bioactive coatings on implants to prevent infection and accelerate tissue regeneration, but the concentrations required are higher than for antibiotics.

Nonsurgical Periodontal Treatment Options and Their Impact on Subgingival Microbiota.

BACKGROUND: Different periodontal treatment methods (quadrant-wise debridement, scaling and root planing (Q-SRP), full-mouth scaling (FMS), full-mouth disinfection (FMD), and FMD with adjuvant erythritol air-polishing (FMDAP)) were applied in periodontitis patients (stage III/IV). The study objective (substudy of ClinicalTrials.gov Identifier: NCT03509233) was to compare the impact of treatments on subgingival colonization. METHODS: Forty patients were randomized to the treatment groups. Periodontal parameters and subgingival colonization were evaluated at baseline and 3 and 6 months after treatment. RESULTS: Positive changes in clinical parameters were recorded in every treatment group during the 3-month follow-up period, but did not always continue. In three groups, specific bacteria decreased after 3 months; however, this was associated with a renewed increase after 6 months (FMS: Porphyromonas gingivalis; FMD: Eubacterium nodatum, Prevotella dentalis; and FMDAP: uncultured Prevotella sp.). CONCLUSIONS: The benefit of all clinical treatments measured after 3 months was associated with a decrease in pathogenic bacteria in the FMS, FMD, and FMDAP groups. However, after 6 months, we observed further improvement or some stagnation in clinical outcomes accompanied by deterioration of the microbiological profile. Investigating the subgingival microbiota might help appraise successful periodontal treatment and implement individualized therapy.

The Effect of Toothpastes Containing Natural Extracts on Bacterial Species of a Microcosm Biofilm and on Enamel Caries Development.

This study investigated the effects of herbal toothpaste on bacterial counts and enamel demineralization. Thirty-six bovine enamel samples were exposed to a microcosm biofilm using human saliva and McBain saliva (0.2% sucrose) for 5 days at 37 °C and first incubated anaerobically, then aerobically-capnophilically. The following experimental toothpaste slurries (2 × 2 min/day) were applied: (1) Vochysia tucanorum (10 mg/g); (2) Myrcia bella (5 mg/g); (3) Matricaria chamomilla (80 mg/g); (4) Myrrha and propolis toothpaste (commercial); (5) fluoride (F) and triclosan (1450 ppm F), 0.3% triclosan and sorbitol (Colgate®, positive control); (6) placebo (negative control). The pH of the medium was measured, bacteria were analyzed using quantitative polymerase chain reaction, and enamel demineralization was quantified using transverse microradiography. The total bacterial count was reduced by toothpaste containing Myrcia bella, Matricaria chamomilla, fluoride, and triclosan (commercial) compared to the placebo. As far as assessable, Myrcia bella, Matricaria chamomilla, and Myrrha and propolis (commercial) inhibited the outgrowth of S. mutans, while Lactobacillus spp. were reduced/eliminated by all toothpastes except Vochysia tucanorum. Mineral loss and lesion depth were significantly reduced by all toothpastes (total: 1423.6 ± 115.2 vol% × µm; 57.3 ± 9.8 µm) compared to the placebo (2420.0 ± 626.0 vol% × µm; 108.9 ± 21.17 µm). Herbal toothpastes were able to reduce enamel demineralization.

The Antimicrobial Susceptibility of Porphyromonas gingivalis: Genetic Repertoire, Global Phenotype, and Review of the Literature.

The in vitro antimicrobial susceptibility of 29 strains of the major periodontal pathogen Porphyromonas gingivalis and three P. gulae (as an ancestor) to nine antibiotics (amoxicillin, amoxicillin/clavulanate, clindamycin, metronidazole, moxifloxacin, doxycycline, azithromycin, imipenem, and cefoxitin) was evaluated by E-testing of minimal inhibitory concentration (MIC) according to international standards. The results were compared with 16 international studies reporting MICs from 1993 until recently. In addition, 77 currently available P. gingivalis genomes were screened for antimicrobial resistance genes. E-testing revealed a 100% sensitivity of P. gingivalis and P. gulae to all antibiotics. This was independent of the isolation year (1970 until 2021) or region, including rural areas in Indonesia and Africa. Regarding studies worldwide (675 strains), several method varieties regarding medium, McFarland inoculation standards (0.5-2) and incubation time (48-168 h) were used for MIC-testing. Overall, no resistances have been reported for amoxicillin + clavulanate, cefoxitin, and imipenem. Few strains showed intermediate susceptibility or resistance to amoxicillin and metronidazole, with the latter needing both confirmation and attention. The only antibiotics which might fail in the treatment of P. gingivalis-associated mixed anaerobic infections are clindamycin, macrolides, and tetracyclines, corresponding to the resistance genes erm(B), erm(F), and tet(Q) detected in our study here, as well as fluoroquinolones. Periodical antibiotic susceptibility testing is necessary to determine the efficacy of antimicrobial agents and to optimize antibiotic stewardship.

Antimicrobial Impact of Different Air-Polishing Powders in a Subgingival Biofilm Model.

Subgingival air-polishing devices (SAPD) can reduce bacterial biofilms and thus support periodontal healing. The authors of this study evaluated the effectiveness of the glycine-based and trehalose-based air-polishing powders in removing pathogenic bacteria in a subgingival biofilm model. We treated 56 subgingival pockets in porcine jaws with SAPD. Subgingival air polishing was performed in three groups of 13 pockets each: I, glycine-based powder; II, trehalose-based powder; and III, water alone. Another group (IV) served as untreated controls. Prior to air polishing, inoculated titanium bars were inserted into the pockets containing periopathogenic bacteria such as Porphyromonas gingivalis and Tannerella forsythia. Remaining bacteria were evaluated using real-time PCR. The numbers of remaining bacteria depended on the treatment procedure, with the lowest number of total bacteria in group I (median: 1.96 × 106 CFU; min: 1.46 × 105; max: 9.30 × 106). Both polishing powders in groups I and II (median: 1.36 × 107 CFU; min: 5.22 × 105; max: 7.50 × 107) showed a statistically significantly lower total bacterial load in comparison to both group IV (median: 2.02 × 108 CFU; min: 5.14 × 107; max: 4.51 × 108; p < 0.05) and group III (median: 4.58 × 107 CFU; min: 2.00 × 106; max: 3.06 × 108; p < 0.05). Both subgingival air-polishing powders investigated can reduce periopathogenic bacteria and thus support antimicrobial therapy approaches in periodontal treatment regimens.

A New Species-Specific Typing Method for Salivarius Group Streptococci Based on the Dephospho-Coenzyme A Kinase (coaE) Gene Sequencing.

Viridans streptococci are a group of α-hemolytic streptococcal species. They are mainly commensals, most abundant in the mouth supporting oral health. But they also include important human pathogens such as Streptococcus pneumoniae. Identification and molecular typing of viridans group streptococci are challenging, especially for members of the salivarius group. In this study, we developed a single-locus molecular typing method that is able to differentiate among the highly phylogenetically related members of the salivarius group (S. salivarius, S. vestibularis and S. thermophilus) and might support differentiation in other groups as well. This typing approach is based on the amplification and sequence analysis of the housekeeping gene dephospho-coenzyme A kinase (coaE), a gene with unrecognized taxonomic potential to date. Here, we analysed coaE gene sequences of 154 publicly available genomes and of 30 salivarius group isolates of our own collection that together belong to 20 different gram-positive bacterial (sub) species. Our results revealed that the coaE phylogeny distinguished between streptococcal and non-streptococcal genomes and that coaE gene sequences were species-specific. In contrast to MALDI-TOF MS performance, the coaE typing was able to precisely identify the phylogenetically very closely related members of the salivarius group.

The Effect of Solutions Containing Extracts of Vochysia tucanorum Mart., Myrcia bellaCambess., Matricaria chamomilla L. and Malva sylvestris L. on Cariogenic Bacterial Species and Enamel Caries Development.

This study evaluated the effect of experimental solutions containing plant extracts on bacterial species and enamel caries prevention. Microcosm biofilm was produced from human saliva mixed with McBain saliva (0.2% sucrose) on bovine enamel for 5 days (3 days under anaerobiosis and 2 days under aerobiosis) at 37°C. From the 2nd day, the following treatments were applied (1 × 60 s/day): Vochysia tucanorum (10 mg/mL); Myrcia bella (5 mg/mL); Matricaria chamomilla (80 mg/mL); Malva sylvestris, fluoride, and xylitol (Malvatricin Plus®); 0.12% chlorhexidine (CHX, PerioGard®); and PBS (negative control). The medium pH was measured. Quantitative polymerase chain reaction was performed for the detection of Streptococcus mutans and Lactobacillus spp. Enamel demineralization was measured by spectral-domain optical coherence tomography. The data were compared by means of the Kruskal-Wallis/Dunn, two-way ANOVA/Bonferroni, and ANOVA/Tukey tests (p < 0.05). The pH decreased after sucrose exposure; only CHX reestablished pH >5.5 by the last day. CHX also eliminated Lactobacillusspp., but the other treatments did not differ significantly from PBS. Malvatricin Plus® and CHX eliminated S. mutans, but the other treatments did not differ from PBS. Similar results were seen concerning the reduction of lesion depth and reflectivity. The experimental natural-extract solutions were ineffective against cariogenic bacteria and in preventing the development of enamel caries.

A concerted probiotic activity to inhibit periodontitis-associated bacteria.

Periodontitis can result in tooth loss and the associated chronic inflammation can provoke several severe systemic health risks. Adjunctive to mechanical treatment of periodontitis and as alternatives to antibiotics, the use of probiotic bacteria was suggested. In this study, the inhibitory effect of the probiotic Streptococcus salivarius subsp. salivarius strains M18 and K12, Streptococcus oralis subsp. dentisani 7746, and Lactobacillus reuteri ATCC PTA 5289 on anaerobic periodontal bacteria and Aggregatibacter actinomycetemcomitans was tested. Rarely included in other studies, we also quantified the inverse effect of pathogens on probiotic growth. Probiotics and periodontal pathogens were co-incubated anaerobically in a mixture of autoclaved saliva and brain heart infusion broth. The resulting genome numbers of the pathogens and of the probiotics were measured by quantitative real-time PCR. Mixtures of the streptococcal probiotics were also used to determine their synergistic, additive, or antagonistic effects. The overall best inhibitor of the periodontal pathogens was L. reuteri ATCC PTA 5289, but the effect is coenzyme B12-, anaerobiosis-, as well as glycerol-dependent, and further modulated by L. reuteri strain DSM 17938. Notably, in absence of glycerol, the pathogen-inhibitory effect could even turn into a growth spurt. Among the streptococci tested, S. salivarius M18 had the most constant inhibitory potential against all pathogens, followed by K12 and S. dentisani 7746, with the latter still having significant inhibitory effects on P. intermedia and A. actinomycetemcomitans. Overall, mixtures of the streptococcal probiotics did inhibit the growth of the pathogens equally or-in the case of A. actinomycetemcomitans- better than the individual strains. P. gingivalis and F. nucleatum were best inhibited by pure cultures of S. salivarius K12 or S. salivarius M18, respectively. Testing inverse effects, the growth of S. salivarius M18 was enhanced when incubated with the periodontal pathogens minus/plus other probiotics. In contrast, S. oralis subsp. dentisani 7746 was not much influenced by the pathogens. Instead, it was significantly inhibited by the presence of other streptococcal probiotics. In conclusion, despite some natural limits such as persistence, the full potential for probiotic treatment is by far not utilized yet. Especially, further exploring concerted activity by combining synergistic strains, together with the application of oral prebiotics and essential supplements and conditions, is mandatory.