Oral Biofilm Composition, Dissemination to Keratinocytes, and Inflammatory Attenuation Depend on Probiotic and Synbiotic Strain Specificity.

Several inflammatory diseases are characterized by a disruption in the equilibrium between the host and its microbiome. Due to the increase in resistance, the use of antibiotics for the widespread, nonspecific killing of microorganisms is at risk. Pro-microbial approaches focused on stimulating or introducing beneficial species antagonistic toward pathobionts may be a viable alternative for restoring the host-microbiome equilibrium. Unfortunately, not all potential probiotic or synbiotic species and even subspecies (to strain level) are equally effective for the designated pathology, leading to conflicting accounts of their efficacy. To assess the extent of these species- and strain-specific effects, 13 probiotic candidates were evaluated for their probiotic and synbiotic potential with glycerol on in vitro oral biofilms, dissemination from biofilms to keratinocytes, and anti-inflammatory activity. Species- and strain-specific effects and efficacies were observed in how they functioned as probiotics or synbiotics by influencing oral pathobionts and commensals within biofilms and affected the dissemination of pathobionts to keratinocytes, ranging from ineffective strains to strains that reduced pathobionts by 3 + log. In addition, a minority of the candidates exhibited the ability to mitigate the inflammatory response of LPS-stimulated monocytes. For a comprehensive assessment of probiotic therapy for oral health, a judicious selection of fully characterized probiotic strains that are specifically tailored to the designated pathology is required. This approach aims to challenge the prevailing perception of probiotics, shifting the focus away from "form over function." Rather than using unproven, hypothetical probiotic strains from known genera or species, one should choose strains that are actually functional in resolving the desired pathology before labelling them probiotics.

pH-Triggered Controlled Release of Chlorhexidine Using Chitosan-Coated Titanium Silica Composite for Dental Infection Prevention.

Peri-implantitis is a growing pathological concern for dental implants which aggravates the occurrence of revision surgeries. This increases the burden on both hospitals and the patients themselves. Research is now focused on the development of materials and accompanying implants designed to resist biofilm formation. To enhance this endeavor, a smart method of biofilm inhibition coupled with limiting toxicity to the host cells is crucial. Therefore, this research aims to establish a proof-of-concept for the pH-triggered release of chlorhexidine (CHX), an antiseptic commonly used in mouth rinses, from a titanium (Ti) substrate to inhibit biofilm formation on its surface. To this end, a macroporous Ti matrix is filled with mesoporous silica (together referred to as Ti/SiO2), which acts as a diffusion barrier for CHX from the CHX feed side to the release side. To limit release to acidic conditions, the release side of Ti/SiO2 is coated with crosslinked chitosan (CS), a pH-responsive and antimicrobial natural polymer. Scanning electron microscopy coupled with energy dispersive X-ray spectroscopy (SEM/EDX) and Fourier transform infrared (FTIR) spectroscopy confirmed successful CS film formation and crosslinking on the Ti/SiO2 disks. The presence of the CS coating reduced CHX release by 33% as compared to non-coated Ti/SiO2 disks, thus reducing the antiseptic exposure to the environment in normal conditions. Simultaneous differential scanning calorimetry and thermogravimetric analyzer (SDT) results highlighted the thermal stability of the crosslinked CS films. Quartz crystal microbalance with dissipation monitoring (QCM-D) indicated a clear pH response for crosslinked CS coatings in an acidic medium. This pH response also influenced CHX release through a Ti/SiO2/CS disk where the CHX release was higher than the average trend in the neutral medium. Finally, the antimicrobial study revealed a significant reduction in biofilm formation for the CS-coated samples compared to the control sample using viability quantitative polymerase chain reaction (v-qPCR) measurements, which were also corroborated using SEM imaging. Overall, this study investigates the smart triggered release of pharmaceutical agents aimed at inhibiting biofilm formation, with potential applicability to implant-like structures.

Unlocking the mechanism of action: a cost-effective flow cytometry approach for accelerating antimicrobial drug development.

Antimicrobial resistance is one of the greatest challenges to global health. While the development of new antimicrobials can combat resistance, low profitability reduces the number of new compounds brought to market. Elucidating the mechanism of action is crucial for developing new antimicrobials. This can become expensive as there are no universally applicable pipelines. Phenotypic heterogeneity of microbial populations resulting from antimicrobial treatment can be captured through flow cytometric fingerprinting. Since antimicrobials are classified into limited groups, the mechanism of action of known compounds can be used for predictive modeling. We demonstrate a cost-effective flow cytometry approach for determining the mechanism of action of new compounds. Cultures of Actinomyces viscosus and Fusobacterium nucleatum were treated with different antimicrobials and measured by flow cytometry. A Gaussian mixture mask was applied over the data to construct phenotypic fingerprints. Fingerprints were used to assess statistical differences between mechanism of action groups and to train random forest classifiers. Classifiers were then used to predict the mechanism of action of cephalothin. Statistical differences were found among the different mechanisms of action groups. Pairwise comparison showed statistical differences for 35 out of 45 pairs for A. viscosus and for 32 out of 45 pairs for F. nucleatum after 3.5 h of treatment. The best-performing random forest classifier yielded a Matthews correlation coefficient of 0.92 and the mechanism of action of cephalothin could be successfully predicted. These findings suggest that flow cytometry can be a cheap and fast alternative for determining the mechanism of action of new antimicrobials.IMPORTANCEIn the context of the emerging threat of antimicrobial resistance, the development of novel antimicrobials is a commonly employed strategy to combat resistance. Elucidating the mechanism of action of novel compounds is crucial in this development but can become expensive, as no universally applicable pipelines currently exist. We present a novel flow cytometry-based approach capable of determining the mechanism of action swiftly and cost-effectively. The workflow aims to accelerate drug discovery and could help facilitate a more targeted approach for antimicrobial treatment of patients.

Morphological aspects and distribution of granules composed of deproteinized bovine bone or human dentin into a putty mixture: an in vitro study.

OBJECTIVE: The main aim of this study was to evaluate the morphological aspects and distribution of granules composed of deproteinized bovine bone mineral (DBBM) and human dentin-derived bone graft (HDBG) into a putty consistency mixture. MATERIALS AND METHODS: DBBM or HDBG were mixed with an alginate-based hydrogel at two different granule/hydrogel ratio (1:1 and 1:3) and divided into four test groups while two control groups were composed of DBBM or HDBG free of hydrogel. Groups of specimens were cross-sectioned for morphological evaluation by scanning electron microscopy (SEM) at backscattered electrons mode. Details on the dimensions and pores' size of DBBM and HDBG were evaluated after mixing different amounts of particles and alginate-based hydrogels. RESULTS: Microscopic analyses revealed a size of DBBM granules ranging from 750 up to 1600 µm while HDBG particles showed particle size ranging from 375 up to 1500 µm. No statistical differences were identified regarding the size of granules (p > 0.5). The mean values of pores' size of DBBM particles were noticed at around 400 µm while HDBG particles revealed micro-scale pores of around 1-3 µm promoted by the dentin tubules (p < 0.05). The lowest distance between particles was at 125 µm for HDBG and 250 µm for DBBM when the particle content was increased. On decreasing the particles' content, the distance between particles was larger for DBBM (~ 1000 µm) and HDBG (~ 1100 µm). In fact, statistically significant differences were found when the content of granules increased (p < 0.05). CONCLUSIONS: The increased content of bioactive ceramic granules in a putty consistency mixture with hydrogel decreased the space among granules that can promote a high ceramic density and stimulate the bone growth over the healing process. Macro-scale pores on bovine bone mineral granules stimulate the formation of blood vessels and cell migration while the micro-scale pores of dentin-derived granules are proper for the adsorption of proteins and growth of osteogenic cells on the bone healing process. CLINICAL SIGNIFICANCE: A high amount of bioactive ceramic granules should be considered when mixing with hydrogels as a putty material since that result in small spaces among granules maintaining the bone volume over the bone healing process. Deproteinized bovine bone mineral granules have macro-scale pores providing an enhanced angiogenesis while dentin-derived granules possess only micro-scale pores for the adsorption of proteins and proliferation of osteogenic cells on the bone healing process. Further studies should evaluate the combination of different bioactive ceramic materials for enhanced bone healing.

Do autologous platelet concentrates (APCs) have a role in intra-oral bone regeneration? A critical review of clinical guidelines on decision-making process.

In the past decades, personalized regenerative medicine has gained increased attention. Autologous platelet concentrates (APCs) such as PRP, PRGF, and L-PRF, all serving as a source of a large variety of cells and growth factors that participate in hard and soft tissue healing and regeneration, could play a significant role in regenerative periodontal procedures. This narrative review evaluated the relative impact of APCs in alveolar ridge preservation, sinus floor augmentation, and the regeneration of bony craters around teeth, both as a single substitute or in combination with a xenograft. L-PRF has a significant beneficial effect on alveolar ridge preservation (bone quality). The data for PRGF are less convincing, and PRP is controversial. L-PRF can successfully be used as a single substitute during transcrestal (≥3.5 mm bone gain) as well as 1-stage lateral window sinus floor elevation (>5 mm bone gain). For PRGF and especially PRP the data are very scarce. In the treatment of bony craters around teeth, during open flap debridement, L-PRF as a single substitute showed significant adjunctive benefits (e.g., >PPD reduction, >CAL gain, >crater depth reduction). The data for PRP and PRGF were non-conclusive. Adding PRP or L-PRF to a xenograft during OFD resulted in additional improvements (>PPD reduction, >CAL gain, >bone fill), for PRGF no data were found. Autologous platelet concentrates demonstrated to enhance bone and soft tissue healing in periodontal regenerative procedures. The data for L-PRF were most convincing. L-PRF also has the advantage of a greater simplicity of production, and its 100% autologous character.

Spatiotemporal monitoring of a periodontal multispecies biofilm model: demonstration of prebiotic treatment responses.

Biofilms are complex polymicrobial communities which are often associated with human infections such as the oral disease periodontitis. Studying these complex communities under controlled conditions requires in vitro biofilm model systems that mimic the natural environment as close as possible. This study established a multispecies periodontal model in the drip flow biofilm reactor in order to mimic the continuous flow of nutrients at the air-liquid interface in the oral cavity. The design is engineered to enable real-time characterization. A community of five bacteria, Streptococcus gordonii-GFPmut3*, Streptococcus oralis-GFPmut3*, Streptococcus sanguinis-pVMCherry, Fusobacterium nucleatum, and Porphyromonas gingivalis-SNAP26 is visualized using two distinct fluorescent proteins and the SNAP-tag. The biofilm in the reactor develops into a heterogeneous, spatially uniform, dense, and metabolically active biofilm with relative cell abundances similar to those in a healthy individual. Metabolic activity, structural features, and bacterial composition of the biofilm remain stable from 3 to 6 days. As a proof of concept for our periodontal model, the 3 days developed biofilm is exposed to a prebiotic treatment with L-arginine. Multifaceted effects of L-arginine on the oral biofilm were validated by this model setup. L-arginine showed to inhibit growth and incorporation of the pathogenic species and to reduce biofilm thickness and volume. Additionally, L-arginine is metabolized by Streptococcus gordonii-GFPmut3* and Streptococcus sanguinis-pVMCherry, producing high levels of ornithine and ammonium in the biofilm. In conclusion, our drip flow reactor setup is promising in studying spatiotemporal behavior of a multispecies periodontal community.ImportancePeriodontitis is a multifactorial chronic inflammatory disease in the oral cavity associated with the accumulation of microorganisms in a biofilm. Not the presence of the biofilm as such, but changes in the microbiota (i.e., dysbiosis) drive the development of periodontitis, resulting in the destruction of tooth-supporting tissues. In this respect, novel treatment approaches focus on maintaining the health-associated homeostasis of the resident oral microbiota. To get insight in dynamic biofilm responses, our research presents the establishment of a periodontal biofilm model including Streptococcus gordonii, Streptococcus oralis, Streptococcus sanguinis, Fusobacterium nucleatum, and Porphyromonas gingivalis. The added value of the model setup is the combination of simulating continuously changing natural mouth conditions with spatiotemporal biofilm profiling using non-destructive characterization tools. These applications are limited for periodontal biofilm research and would contribute in understanding treatment mechanisms, short- or long-term exposure effects, the adaptation potential of the biofilm and thus treatment strategies.

Probiotics for oral health: do they deliver what they promise?

Probiotics have demonstrated oral health benefits by influencing the microbiome and the host. Although promising, their current use is potentially constrained by several restrictions. One such limiting factor lies in the prevailing preparation of a probiotic product. To commercialize the probiotic, a shelf stable product is achieved by temporarily inactivating the live probiotic through drying or freeze drying. Even though a lyophilized probiotic can be kept dormant for an extended period of time, their viability can be severely compromised, making their designation as probiotics questionable. Additionally, does the application of an inactive probiotic directly into the oral cavity make sense? While the dormancy may allow for survival on its way towards the gut, does it affect their capacity for oral colonisation? To evaluate this, 21 probiotic product for oral health were analysed for the number of viable (probiotic), culturable (CFU) and dead (postbiotic) cells, to verify whether the commercial products indeed contain what they proclaim. After isolating and uniformly lyophilizing three common probiotic species in a simple yet effective lyoprotective medium, the adhesion to saliva covered hydroxyapatite discs of lyophilized probiotics was compared to fresh or reactivated lyophilized probiotics. Unfortunately, many of the examined products failed to contain the claimed amounts of viable cells, but also the strains used were inadequately characterized and lacked clinical evidence for that unknown strain, questioning their label of a 'probiotic'. Additionally, lyophilized probiotics demonstrated low adhesive capacity compared to their counterparts, prompting the question of why fresh or reactivated probiotics are not currently used.

Chlorin-e6 conjugated to the antimicrobial peptide LL-37 loaded nanoemulsion enhances photodynamic therapy against multi-species biofilms related to periodontitis.

In our previous studies, Chlorin-e6 (Ce6) demonstrated a significant reduction of microorganisms' viability against multi-species biofilm related to periodontitis while irradiated with blue light. However, the conjugation of Ce6 and antimicrobial peptides, and the incorporation of this photosensitizer in a nanocarrier, is still poorly explored. We hypothesized that chlorin-e6 conjugated to the antimicrobial peptide LL-37 loaded nanoemulsion could inhibit a multi-species biofilm related to periodontitis during photodynamic therapy (PDT), the pre-treatment with hydrogen peroxide was also tested. The nanoemulsion (NE) incorporated with Ce6 was characterized regarding the physiochemical parameters. Images were obtained by transmission electron microscopy (TEM) and scanning electron microscopy (SEM). Later, the Ce6 and LL-37 incorporated in NE was submitted to UV-Vis analysis and Reactive Oxygen Species (ROS) assay. Finally, the combined formulation (Ce6+LL-37 in nanoemulsion) was tested against multi-species biofilm related to periodontitis. The formed nanoformulation was kinetically stable, optically transparent with a relatively small droplet diameter (134.2 unloaded and 146.9 loaded), and weak light scattering. The NE system did not impact the standard UV-VIS spectra of Ce6, and the ROS production was improved while Ce6 was incorporated in the NE. The combination of Ce6 and LL-37 in NE was effective to reduce the viability of all bacteria tested. The treatment with hydrogen peroxide previous to PDT significantly impacted bacterial viability. The current aPDT regimen was the best already tested against periodontal biofilm by our research team. Our results suggest that this combined protocol must be exploited for clinical applications in localized infections such as periodontal disease. - Nanoemulsion demonstrated to be an excellent nanocarrier for photodynamic application. - Chlorin-e6 incorporated in nanoemulsion showed great physicochemical and biophotonic parameters. - The combination of chlorin-e6 and LL-37 peptide in nanoemulsion is effective to eliminate periodontal pathogenic bacteria. - The treatment with hydrogen peroxide previous to PDT significantly impacted bacterial viability.

Risk factors for gingival recessions after orthodontic treatment: a systematic review.

BACKGROUND/OBJECTIVES: To systematically review the available evidence concerning the risk factors for gingival recessions (GR) after orthodontic treatment (OT). DATA COLLECTION AND ANALYSIS: Data was obtained and collected by systematically searching 3 data bases: Pubmed, EMBASE, and Web of Science until 20 April 2023. Controlled trials, cohort, case-control or cross-sectional studies describing GR or clinical crown height (CCH) after OT were included. The risk of bias in the selected studies was evaluated with the methodological index for non-randomized studies. RESULTS: Forty-eight articles were included, investigating the following six risk factors for GR: 1. OT (n = 21), 2. Type of orthodontic intervention (n = 32), 3. Patient's baseline occlusal and skeletal characteristics (n = 14), 4. Mucogingival characteristics (n = 10), 5. Oral hygiene (n = 9), and 6. Others (n = 12). Significantly higher prevalence, severity and extent of GR were found in orthodontic patients by 10/15, 4/10, and 2/2 articles respectively. 10/16 articles reported significantly more GR and increased CCH in patients where orthodontic incisor proclination was performed. The evidence surrounding maxillary expansion and orthodontic retention was too heterogeneous to allow for? definitive conclusions. Pre-treatment angle classification, ANB, overjet, overbite, arch width and mandibular divergence were found not to be associated with GR (9/14), while pre-treatment crossbite, symphysis height and width were (5/7 studies). A thin gingival biotype, presence of previous GR, baseline width of keratinized gingiva and facial gingival margin thickness were correlated with increased risk of GR after OT by nine articles, while pocket depth was not. Oral hygiene, sex, treatment duration, and oral piercings were found not to be linked with GR in orthodontic patients, while GR was reported to increase with age in orthodontic patients by 50 per cent of the articles investigating this factor. The mean risk of bias for comparative and not comparative studies was 14.17/24 and 9.12/16. LIMITATIONS: The selected studies were quite heterogeneous regarding study settings, variables reported and included very limited sample sizes. CONCLUSION: Although studies regarding the risk factors for GR are relatively abundant, they are very heterogeneous concerning design, studied factors, methodology and reporting, which often leads to contradictory results. Uniform reporting guidelines are urgently needed for future research. PROSPERO REGISTRATION: CRD42020181661. FUNDING: This research received no funding.

Pre-operative, chair-side Zn-containing surgical stents affect morbidity and wound healing after free gingival graft harvesting: a randomized clinical trial.

OBJECTIVE: To compare a pre-operatively, chair-side made, zinc-containing surgical stent (ZN) and suturing of a gelatin-based hemostatic agent (HA) on palatal wound healing and patient morbidity after free gingival graft surgery (FGG). MATERIALS AND METHODS: Sixty patients requiring FGG were randomly divided into two groups to receive either a ZN or a sterile HA sutured on the surgical area. Patients were evaluated at 1st, 3rd, 7th, 14th, 28th, and 56th days following surgery. Overall surgical time, donor site surgical time, postoperative pain (PP), delayed bleeding (DB), changes in dietary habits (DH), burning sensation (BS), completion of re-epithelialization (CE), and patients' discomfort (PD) were evaluated. RESULTS: Donor site surgical time, PP, DB, DH, BS were statistically significantly lower in the ZN group together with faster completion of re-epithelialization compared to the HA group. CONCLUSION: Pre-operatively, chair-side made, zinc-containing surgical stents provided significant benefits for wound healing parameters and patients' postoperative morbidity after FGG harvesting. CLINICAL RELEVANCE: The results show that using Zn-containing palatal stent after free gingival graft surgery significantly reduces pain and patient morbidity during the postoperative period.

Europe's contribution to the evaluation of the use of systemic antimicrobials in the treatment of periodontitis.

This narrative review celebrates Europe's contribution to the current knowledge on systemically administered antimicrobials in periodontal treatment. Periodontitis is the most frequent chronic noncommunicable human disease. It is caused by dysbiotic bacterial biofilms and is commonly treated with subgingival instrumentation. However, some sites/patients do not respond adequately, and its limitations and shortcomings have been recognized. This has led to the development of alternative or adjunctive therapies. One is the use of antimicrobials to target bacteria in subgingival biofilms in the periodontal pocket, which can be targeted directly through the pocket entrance with a locally delivered antibiotic or systemically by oral, intravenous, or intramuscular methods. Since the early 20th century, several studies on systemic antibiotics have been undertaken and published, especially between 1990 and 2010. Europe's latest contribution to this topic is the first European Federation of Periodontology, S3-level Clinical Practice Guideline, which incorporates recommendations related to the use of adjuncts to treat stage I-III periodontitis. Understanding the etiopathogenesis of periodontal diseases, specifically periodontitis, has influenced the use of systemic periodontal antibiotic therapy. Randomized clinical trials and systematic reviews with meta-analyses have demonstrated the clinical advantages of adjunctive systemic antimicrobials. However, current recommendations are restrictive due to concerns about antibiotic misuse and the increase in microbial antibiotic resistance. European researchers have contributed to the use of systemic antimicrobials in the treatment of periodontitis through clinical trials and by providing rational guidelines. Nowadays, European researchers are exploring alternatives and directing clinical practice by providing evidence-based guidelines to limit the use of systemic antimicrobials.

Mode of killing determines the necrotrophic response of oral bacteria.

Background: Bacteria respond to changes in their environment, such as nutrient depletion and antimicrobials exposure. Antimicrobials result not only in bacterial death, but also have a hand in determining species abundances and ecology of the oral biofilms. Proximity of dead bacterial cells to living ones is an important environmental change or stress factor. Dead bacteria represent high concentrations of nutrients, such as proteins, lipids, sugars, and nucleic acids. Living bacteria can use these biomasses as a nutrients source, which is termed necrotrophy. Aim: This study investigates the effect of exposing living oral bacteria (planktonic and biofilms) to their dead siblings after being killed by heat or hydrogen peroxide. Results: Tested bacterial species showed different responses towards the dead cells, depending on the mode of killing, the nutritional value of the culture media, and the the dead cells density. The multispecies oral biofilms showed different responses towards the supplementation of dead cells during biofilm development, while matured biofilms were more resilient. Conclusion: This study indicates that dead bacteria resulting from antiseptics use may imbalance the nutrient availability in the oral cavity, resulting in overgrowth of opportunistic species, and hence ecological changes in oral communities, or introducing new bacterial phenotypes.

Association of nine pathobionts with periodontitis in four South American and European countries.

Aim: Our aim was to compare the prevalence and load of nine pathobionts in subgingival samples of healthy individuals and periodontitis patients from four different countries. Methods: Five hundred and seven subgingival biofilm samples were collected from healthy subjects and periodontitis patients in Belgium, Chile, Peru and Spain. The prevalence and load of Eubacterium brachy, Filifactor alocis, Fretibacterium fastidiosum, Porphyromonas endodontalis, Porphyromonas gingivalis, Selenomonas sputigena, Treponema denticola, Tannerella forsythia and Treponema socranskii were measured by quantitative PCR. Results: The association with periodontitis of all species, except for T. socranskii, was confirmed in all countries but Peru, where only P. endodontalis, P. gingivalis and T. denticola were found to be significantly associated. Moreover, most species showed higher loads at greater CAL and PPD, but not where there was BOP. Through Principal Component Analysis, samples showed clearly different distributions by diagnosis, despite observing a smaller separation in Peruvian samples. Conclusions: Unlike prevalence, relative load was found to be a reliable variable to discriminate the association of the species with periodontitis. Based on this, F. alocis, P. endodontalis, P. gingivalis, T. denticola and T. forsythia may be biomarkers of disease in Belgium, Chile and Spain, due to their significantly higher abundance in periodontitis patients.

Adjunctive locally and systemically delivered antimicrobials during surgical treatment of peri-implantitis: A systematic review.

AIM: To answer the following PICOS question: "In patients with peri-implantitis, what is the efficacy of surgical therapy with adjunctive systemic or local antimicrobials, in comparison with surgical therapy alone, in terms of pocket probing depth reduction, as assessed in randomized controlled trials (RCTs) with at least 6 months of follow-up?" MATERIALS AND METHODS: A systematic literature search was conducted. Reduction in mean probing pocket depth (PPD) was the primary outcome. Secondary clinical outcomes were changes in suppuration (%), changes in bleeding on probing (BOP) (%), marginal bone level changes (mm), disease resolution (%), and implant/prosthesis loss (%). Patient-reported outcome measures, possible adverse effects, and oral-health-related quality of life were also extracted if such data were available. RESULTS: Four RCTs assessing the use of locally (two RCTs) and systemically (two RCTs) administered antimicrobial adjuncts to surgical treatment of peri-implantitis, with 6-36-month follow-up, were included. Because of the substantial heterogeneity of interventions between the studies, meta-analysis could not be performed. A reduction in the mean PPD was observed following all the involved surgical treatments, irrespective of the addition of antimicrobials. Except for the effect of systemic antimicrobials on marginal bone level changes and local antimicrobials on BOP, the effect of systemic and local antimicrobials was equivocal for all secondary outcome measures. CONCLUSIONS: Based on the limited available evidence, the adjunctive use of the currently tested systemic or local antimicrobials during surgical therapy, in comparison with surgical therapy alone, in patients with peri-implantitis does not seem to improve the clinical efficacy. With regard the use of systemic antimicrobials, only 50% of the cases showed disease resolution after 1 year. There is a lack of studies that consider the sole use of local antimicrobials. Therefore, their true effect remains unclear.

Antimicrobial potential of known and novel probiotics on in vitro periodontitis biofilms.

Several oral diseases are characterized by a shift within the oral microbiome towards a pathogenic, dysbiotic composition. Broad-spectrum antimicrobials are often part of patient care. However, because of the rising antibiotic resistance, alternatives are increasingly desirable. Alternatively, supplying beneficial species through probiotics is increasingly showing favorable results. Unfortunately, these probiotics are rarely evaluated comparatively. In this study, the in vitro effects of three known and three novel Lactobacillus strains, together with four novel Streptococcus salivarius strains were comparatively evaluated for antagonistic effects on proximal agar growth, antimicrobial properties of probiotic supernatant and the probiotic's effects on in vitro periodontal biofilms. Strain-specific effects were observed as differences in efficacy between genera and differences within genera. While some of the Lactobacillus candidates were able to reduce the periodontal pathobiont A. actinomycetemcomitans, the S. salivarius strains were not. However, the S. salivarius strains were more effective against periodontal pathobionts P. intermedia, P. gingivalis, and F. nucleatum. Vexingly, most of the Lactobacillus strains also negatively affected the prevalence of commensal species within the biofilms, while this was lower for S. salivarius strains. Both within lactobacilli and streptococci, some strains showed significantly more inhibition of the pathobionts, indicating the importance of proper strain selection. Additionally, some species showed reductions in non-target species, which can result in unexpected and unexplored effects on the whole microbiome.

Individual "alveolar phenotype" limits dimensions of lateral bone augmentation.

AIM: Alveolar ridge resorption following tooth extraction often renders a lateral bone augmentation inevitable. Some patients, however, suffer from severe early (during graft healing, Eres ) and/or late (during follow-up, Lres ) graft resorption. We explored the hypothesis that the "individual phenotypic dimensions" may partially explain the degree of such resorptions. MATERIALS AND METHODS: Patients who underwent a guided bone regeneration (GBR) procedure were screened for inclusion according to the following criteria: (1) a relatively symmetrical maxillary arch; (2) an intact contra-lateral alveolar bone dimension; (3) the availability of a pre-operative cone-beam CT (CBCT); (4) a CBCT taken immediately after GBR, and (5) at least one CBCT scan ≥6 months after surgery. CBCT scans from different timepoints were registered and imported into the Mimics software (Materialise, Leuven, Belgium). Bone dimensions of the contra-lateral site of the augmentation, representing the "individual phenotypical dimension (IPD) of the alveolar crest", were superimposed on the augmented site and registered accordingly. As such, Eres and Lres could be measured over time, in relation to the IPD (in two dimensions; per millimetre apically from the alveolar crest, in the centre of the GBR), as well as in three dimensions (the entire GBR, 2 mm away from the mesial, distal, and apical border for standardization). RESULTS: A total of 17 patients (23 augmented sites) were included. After Eres , the outline of the augmentation was in general located ±1 mm outside the IPD, but ≥1.5 years after GBR, it further moved towards the IPD (85% within 0.5 mm distance). CONCLUSIONS: Within the limitations of this study, the results indicate that the dimensions of a lateral bone augmentation are defined by the "individual phenotypic bone boundaries" of the patient.

Root canal disinfection and maintenance of the remnant tooth tissues by using grape seed and cranberry extracts.

The purpose of this study was to perform an integrative review on the effects of cranberry and grape seed extracts concerning the disinfection of root canals maintaining the strength of the remnant tooth tissues' structure. A bibliographical search was carried out on the PubMed electronic platform using the following key terms: cranberry, grape seed, vaccinium macrocarpon, proanthocyanidin, antibacterial, antimicrobial, decontamination, disinfection, bacteria removal, bacteria eradication, bacteria elimination, endodontic, root canal, faecalis, and strength. The inclusion criteria involved articles published in the English language, until March, 2022, reporting the antibacterial effect of grape seed and cranberry extracts. Of 185 studies identified, 13 studies were selected for the present review. The grape seed extract (GSE), composed of proanthocyanidins, showed an antioxidant activity against the main bacteria found in endodontic secondary infection. The percentage of bacteria removal was recorded at around 96.97% by using GSE. Studies on cranberry extracts, which are composed of proanthocyanidins, revealed antimicrobial effects against bacteria related to periodontitis and dental caries. Additionally, GSE or cranberry allowed the dentin collagen cross-linking that preserved the 3D collagen network leading to the maintenance of the strength of the remnant tooth structure. However, the contaminated smear layer could not be removed by using only GSE or cranberry. Cranberry extracts and GSE revealed a significant antimicrobial activity in endodontic disinfection without changing the mechanical properties of the remnant dentin tissues. Furthermore, those components can be associated with traditional compounds to enhance their antimicrobial effects and eliminate the smear layer.

Quantitative PCR Effectively Quantifies Triazole-Susceptible and Triazole-Resistant Aspergillus fumigatus in Mixed Infections.

Increasing resistance to triazole antifungals in Aspergillus fumigatus is worrisome because of the associated high mortality of triazole-resistant A. fumigatus (TRAF) infections. While most studies have focused on single triazole-susceptible (WT) or TRAF infections, reports of TRAF cases developing mixed WT and TRAF infections have been described in several studies. However, the prevalence of mixed infections and their responses to current recommended therapies are unknown and could be inappropriate, leading to poor clinical outcomes. To address the urgent need for tools to diagnose, monitor disease development and therapy efficacies in mixed infection settings where quantification of WT versus TRAF is key, this study developed a novel qPCR assay to differentiate WT and TRAF harboring the cyp51A-TR34/L98H mutation. The proposed assay successfully quantified A. fumigatus and discriminated TRAF-TR34 in vitro and in vivo, which was achieved by increasing the yield of extracted DNA through improved homogenization and specific primers targeting the WT-sequence or TR34-insertion and a TaqMan-probe directed to A. fumigatus. The here-developed qPCR assay overcomes sensitivity issues of methodologies such as CFU counts, providing specific, reproducible, and reliable quantitative information to study and follow up the (interplay and individual) effects of mixed A. fumigatus infections on disease development and treatment responses.

Glycerol strengthens probiotic effect of Limosilactobacillus reuteri in oral biofilms: A synergistic synbiotic approach.

Both in vitro and in vivo studies have shown that the probiotic Limosilactobacillus reuteri can improve oral health. Limosilactobacillus reuteri species are known to produce the antimicrobial "reuterin" from glycerol. In order to further increase its antimicrobial activity, this study evaluated the effect of the combined use of glycerol and Limosilactobacillus reuteri (ATCC PTA 5289) in view of using a synergistic synbiotic over a probiotic. An antagonistic agar growth and a multispecies biofilm model showed that the antimicrobial potential of the probiotic was significantly enhanced against periodontal pathobionts and anaerobic commensals when supplemented with glycerol. Synbiotic biofilms also showed a significant reduction in inflammatory expression of human oral keratinocytes (HOK-18A), but only when the keratinocytes were preincubated with the probiotic. Probiotic preincubation of keratinocytes or probiotic and synbiotic treatment of biofilms alone was insufficient to significantly reduce inflammatory expression. Overall, this study shows that combining glycerol with the probiotic L. reuteri into a synergistic synbiotic can greatly improve the effectiveness of the latter.

The accuracy of probing, ultrasound and cone-beam CT scans for determining the buccal bone plate dimensions around oral implants - A systematic review.

OBJECTIVE: The objective of this review was to assess the accuracy of available means of determining the BBT (buccal bone thickness) and/or BBL (buccal bone level). This was translated into the following research question: What is the accuracy of the available means of visualizing the BBP (buccal bone plate) to establish the BBT and/or the BBL, when compared to control measurements? As control measurements histomorphometric measurements, direct measurements and cone-beam computed tomography (CBCT) measurements in the absence of metal are accepted. BACKGROUND DATA: METHODS: The literary search was performed by searching the databases of MEDLINE, Embase, and Web of Science, up to July 13, 2021. Types of studies included were clinical, in vitro and animal trials, specifically looking into the bone level and/or bone thickness of the buccal bone plate at oral implants. Reference lists were hand searched for relevant articles. Two reviewers performed the data extraction and analysis. Only studies using reliable control measurements to evaluate the accuracy of the tested means of visualizing BBT and/or BBL were included for analysis. The QUADAS-2 tool was used to perform bias analysis on the relevant studies. Extracted data was tabulated to show the differences between test and control measurements for BBT and BBL. For in vitro studies on CBCT measurements of BBT meta-analysis could be performed. RESULTS: A total of 1176 papers were identified in the search. Twenty-two articles were used for data extraction and qualitative analysis. Of these studies nine were animal studies, 9 were in vitro studies and four were human studies. Six animal studies and three human studies provided data on probing. CBCT and sonography as techniques for visualizing the buccal bone plate. Probing at implant sites seems to provide data that correlates with a consistent distance from the BBP. Meta-analysis for probing studies could not be performed due to heterogeneity in the setups of these studies. Eleven studies on CBCT were eligible for inclusion. Of these three were animal studies, the remaining 8 studies were all in vitro studies. Meta-analysis was performed on the accuracy of CBCT for in vitro studies, finding a significant underestimation of the BBT when compared to control measurements by a mean difference of -0.15 mm with 95%CI [-0.26,-0.03]. Three studies were identified on measurement of BBT and/or BBL by sonography. This included one human study and two in vitro studies. The identified studies show a low error when determining the buccal bone level or thickness using sonography. All included studies possess a high risk of bias according to risk of bias analysis, mostly due to selection of the patient. CONCLUSION: A strong limitation of this systematic review is the inclusion of different studies with heterogeneous designs. Within the limits of this analysis it cannot be concluded that probing is an accurate way of visualizing the BBP. CBCT cannot yet be recommended as a standard diagnostic tool for follow-up of the BBP at oral implants. The application of sonography as a diagnostic tool to visualize the BBP needs further scientific validation.