Subgingival microbial profiles in pre- and postmenopausal women: Associations with serum estradiol levels.

BACKGROUND: Subgingival dental plaque is an ecosystem playing a key role in supporting both oral health and systemic health. Menopause-related changes have the potential to disrupt its balance, which is crucial to postmenopausal well-being. Our study explored how circulating estradiol levels correlate with subgingival microbial composition using checkerboard DNA-DNA hybridization in premenopausal and postmenopausal women. We also demonstrated that combining this method with 16S ribosomal RNA (rRNA) sequencing insights remains valuable for examining subgingival ecology. METHODS: We assessed 40 bacterial species in 77 premenopausal and 81 postmenopausal women using checkerboard DNA-DNA hybridization and measured serum estradiol with enzyme-linked immunosorbent assay (ELISA). Women were categorized by subgingival dysbiosis severity using a modified Subgingival Microbial Dysbiosis Index (mSMDI). Six women from each normobiotic and dysbiotic subgroup across premenopausal and postmenopausal women underwent 16S rRNA sequencing analysis. RESULTS: DNA checkerboard analysis revealed that most observed variability in individual bacterial proportions is associated with periodontitis. Two species, Leptotrichia buccalis and Streptococcus constellatus, exhibited differences related to estradiol levels within the premenopausal group (p = 0.055 and p = 0.009, respectively). 16S rRNA sequencing confirmed the mSMDI's validity in categorizing normobiotic and dysbiotic states. Menopausal status was not associated with a dysbiotic shift in the subgingival microbiome despite significantly more attachment loss in postmenopausal compared to premenopausal women. CONCLUSIONS: Our results indicate that decreased estradiol levels or increased attachment loss during menopause are not associated with changes in species abundance or dysbiotic shifts in women. The mSMDI may be a useful tool for classifying subgingival ecology based on its normobiotic or dysbiotic inclination.

Quantifying periodontitis-associated oral dysbiosis in tongue and saliva microbiomes-An integrated data analysis.

BACKGROUND: Periodontitis is primarily driven by subgingival biofilm dysbiosis. However, the quantification and impact of this periodontal dysbiosis on other oral microbial niches remain unclear. This study seeks to quantify the dysbiotic changes in tongue and salivary microbiomes resulting from periodontitis by applying a clinically relevant dysbiosis index to an integrated data analysis. METHODS: The National Center for Biotechnology Information (NCBI) database was searched to identify BioProjects with published studies on salivary and tongue microbiomes of healthy and periodontitis subjects. Raw sequence datasets were processed using a standardized bioinformatic pipeline and categorized by their ecological niche and periodontal status. The subgingival microbial dysbiosis index (SMDI), a dysbiosis index originally developed using the subgingival microbiome, was computed at species and genus levels and customized for each niche. Its diagnostic accuracy for periodontitis was evaluated using receiver operating characteristic curves. RESULTS: Four studies, contributing 328 microbiome samples, were included. At both species and genus levels, periodontitis samples had a higher SMDI, but the differences were only significant for subgingival biofilm and saliva (p < 0.001). However, SMDI showed good diagnostic accuracy for periodontitis status for all three niches (area under curve ranging from 0.76 to 0.90, p < 0.05). The dysbiosis index of subgingival biofilm was positively correlated with saliva consistently (p < 0.001) and with the tongue at the genus level (p = 0.036). CONCLUSIONS: While the impact on the tongue microbiome requires further investigation, periodontitis-associated dysbiosis affects the salivary microbiome and is quantifiable using the dysbiosis index. The diagnostic potential of salivary microbial dysbiosis as a convenient periodontal biomarker for assessing periodontal status has potential public health and clinical applications. PLAIN LANGUAGE SUMMARY: Periodontitis, a severe inflammation of the gums which causes bone loss, is a disease caused by an imbalance of good and bad bacteria under the gums. However, it is unclear how this bacterial imbalance in the gums affects the bacterial balance of other distinct parts of the mouth, such as the saliva and tongue. This study uses bacteria datasets of four previously published studies, contributing a total of 328 bacterial samples. The data were processed using a uniform data analysis workflow, and a bacterial score, the subgingival microbial dysbiosis index (SMDI), previously shown to capture periodontitis-associated bacteria imbalance, was calculated separately for samples from under the gums, the saliva, and the tongue. The SMDI was able to distinguish between health and periodontitis within each oral location, and in general, the scores were higher for periodontitis samples, though this difference was significant only for bacteria under the gums and in saliva. Saliva scores were also consistently correlated with bacteria under the gums. This study shows that periodontitis-associated bacterial imbalances are observed in oral locations beyond just under the gums, particularly the saliva. Thus, saliva bacteria may be used as a convenient biomarker for assessing gum disease, allowing for potential public health and clinical applications.

The effect of dental material type and masticatory forces on periodontitis-derived subgingival microbiomes.

Restorative dental materials can frequently extend below the gingival margin, serving as a potential haven for microbial colonization, and altering the local oral microbiome to ignite infection. However, the contribution of dental materials on driving changes of the composition of the subgingival microbiome is under-investigated. This study evaluated the microbiome-modulating properties of three biomaterials, namely resin dental composites (COM), antimicrobial piezoelectric composites (BTO), and hydroxyapatite (HA), using an optimized in vitro subgingival microbiome model derived from patients with periodontal disease. Dental materials were subjected to static or cyclic loading (mastication forces) during biofilm growth. Microbiome composition was assessed by 16S rRNA gene sequencing. Dysbiosis was measured in terms of subgingival microbial dysbiosis index (SMDI). Biomaterials subjected to cyclic masticatory loads were associated with enhanced biofilm viability except on the antibacterial composite. Biomaterials held static were associated with increased biofilm biomass, especially on HA surfaces. Overall, the microbiome richness (Chao index) was similar for all the biomaterials and loading conditions. However, the microbiome diversity (Shannon index) for the HA beams was significantly different than both composites. In addition, beta diversity analysis revealed significant differences between composites and HA biomaterials, and between both loading conditions (static and cyclic). Under static conditions, microbiomes formed over HA surfaces resulted in increased dysbiosis compared to composites through the enrichment of periopathogens, including Porphyromonas gingivalis, Porphyromonas endodontalis, and Fretibacterium spp., and depletion of commensals such as Granulicatella and Streptococcus spp. Interestingly, cyclic loading reversed the dysbiosis of microbiomes formed over HA (depletion of periopathogenes) but increased the dysbiosis of microbiomes formed over composites (enrichment of Porphyromonas gingivalis and Fusobacterim nucleatum). Comparison of species formed on both composites (control and antibacterial) showed some differences. Commercial composites enriched Selenomonas spp. and depleted Campylobacter concisus. Piezoelectric composites effectively controlled the microbiome viability without significantly impacting the species abundance. Findings of this work open new understandings of the effects of different biomaterials on the modulation of oral biofilms and the relationship with oral subgingival infections.

The short-term impact of comprehensive caries treatment on the supragingival microbiome of severe early childhood caries.

BACKGROUND: Children affected by severe early childhood caries (S-ECC) usually need comprehensive caries treatment due to the extensive of caries. How the oral microbiome changes after caries therapy within the short-term warrant further study. AIM: This study aimed to investigate the short-term impact of comprehensive caries treatment on the supragingival plaque microbiome of S-ECC children. DESIGN: Thirty-three children aged 2-4 years with severe caries (dt > 7) were recruited. Comprehensive caries treatment was performed under general anesthesia in one session and included restoration, pulp treatment, extraction, and fluoride application. Supragingival plaque was sampled pre- and 1-month posttreatment. The genomic DNA of the supragingival plaque was extracted, and bacterial 16S ribosomal RNA gene sequencing was performed. RESULTS: Our data showed that the microbial community evenness significantly decreased posttreatment. Furthermore, comprehensive caries treatment led to more diverse microbial structures among the subjects. The interbacterial interactions reflected by the microbial community's co-occurrence network tended to be less complex posttreatment. Caries treatment increased the relative abundance of Corynebacterium matruchotii, Corynebacterium durum, Actinomyces naeslundii, and Saccharibacteria HMT-347, as well as Aggregatibacter HMT-458 and Haemophilus influenzae. Meanwhile, the relative abundance of Streptococcus mutans, three species from Leptotrichia, Neisseria bacilliformis, and Provotella pallens significantly decreased posttreatment. CONCLUSION: Our results suggested that comprehensive caries treatment may contribute to the reconstruction of a healthier supragingival microbiome.

Bacteriome analysis of Aggregatibacter actinomycetemcomitans-JP2 genotype-associated Grade C periodontitis in Moroccan adolescents.

Background: Grade C (previously aggressive) periodontitis (GCP) in adolescents is prevalent in certain parts of Africa where it is associated with JP2 genotype, a highly virulent strain of Aggregatibacter actinomycetemcomitans. The aim of this study was to characterize the subgingival bacteriome in Moroccan subjects with GCP positive to A. actinomycetemcomitans JP2 genotype. Methods: Subgingival plaque samples were collected from shallow and deep pockets of 8 subjects with GCP (17.2 ± 1.5 years) and from gingival sulci of 13 controls with no periodontitis (14.6 ± 1.1 years). Identification and genotyping of A. actinomycetemcomitans was performed using PCR analysis of the ltx operon, while bacteriome profiling was done by 16S rRNA gene sequencing (V1-V3 region). Groups were compared in terms of microbial diversity, abundances, and dysbiosis. Results: The shallow and deep pocket sites from GCP cases had a significantly altered microbial composition compared to controls. Species associated with health included Haemophilus parainfluenzae, Lautropia mirabilis, Streptococcus spp., Gemella spp., and Rothia spp. While known periodontal pathogens, including Porphyromonas gingivalis, Tannerella forsythia, Treponema spp. and Fretibacterium spp., were significantly enriched in GCP, non-conventional taxa, including Pseudomonas oral taxon C61 and Enterobacter cloacae were more abundant and showed stronger association with the disease. Less significant differences in abundances of individual taxa were observed between shallow and deep pockets. Overall dysbiosis measured in terms of Subgingival Microbial Dysbiosis Index (SMDI) differentiated between GCP and no-periodontitis with 95% accuracy. Conclusions: The results suggest that several periodontal pathogens involved in the adult-type periodontitis also play a role in JP2 genotype-associated GCP. The potential role of non-conventional taxa in the pathogenesis of GCP warrants further investigation.

PepO and CppA modulate Streptococcus sanguinis susceptibility to complement immunity and virulence.

Streptococcus sanguinis is a ubiquitous commensal species of the oral cavity commonly involved as an opportunistic pathogen in cardiovascular infections. In this study, we investigated the functions of endopeptidase O (PepO) and a C3-degrading protease (CppA) in the systemic virulence of S. sanguinis. Isogenic mutants of pepO and cppA obtained in strain SK36 showed increased susceptibility to C3b deposition and to opsonophagocytosis by human polymorphonuclear neutrophils (PMN). These mutants differ, however, in their profiles of binding to serum amyloid P component (SAP) and C1q, whereas both showed reduced interaction with C4b-binding protein (C4BP) and/or factor H (FH) regulators as compared to SK36. The two mutants showed defects in ex vivo persistence in human blood, serum-mediated invasion of HCAEC endothelial cells, and virulence in a Galleria mellonella infection model. The transcriptional activities of pepO and cppA, assessed by RT-qPCR in nine wild-type strains, further indicated strain-specific profiles of pepO/cppA expression. Moreover, non-conserved amino acid substitutions were detected among the strains, mostly in CppA. Phylogenetic comparisons with homologues of streptococcal species of the oral and oropharyngeal sites suggested that S. sanguinis PepO and CppA have independent ancestralities. Thus, this study showed that PepO and CppA are complement evasion proteins expressed by S. sanguinis in a strain-specific manner, which are required for multiple functions associated with cardiovascular virulence.

Phages are unrecognized players in the ecology of the oral pathogen Porphyromonas gingivalis.

BACKGROUND: Porphyromonas gingivalis (hereafter "Pg") is an oral pathogen that has been hypothesized to act as a keystone driver of inflammation and periodontal disease. Although Pg is most readily recovered from individuals with actively progressing periodontal disease, healthy individuals and those with stable non-progressing disease are also colonized by Pg. Insights into the factors shaping the striking strain-level variation in Pg, and its variable associations with disease, are needed to achieve a more mechanistic understanding of periodontal disease and its progression. One of the key forces often shaping strain-level diversity in microbial communities is infection of bacteria by their viral (phage) predators and symbionts. Surprisingly, although Pg has been the subject of study for over 40 years, essentially nothing is known of its phages, and the prevailing paradigm is that phages are not important in the ecology of Pg. RESULTS: Here we systematically addressed the question of whether Pg are infected by phages-and we found that they are. We found that prophages are common in Pg, they are genomically diverse, and they encode genes that have the potential to alter Pg physiology and interactions. We found that phages represent unrecognized targets of the prevalent CRISPR-Cas defense systems in Pg, and that Pg strains encode numerous additional mechanistically diverse candidate anti-phage defense systems. We also found that phages and candidate anti-phage defense system elements together are major contributors to strain-level diversity and the species pangenome of this oral pathogen. Finally, we demonstrate that prophages harbored by a model Pg strain are active in culture, producing extracellular viral particles in broth cultures. CONCLUSION: This work definitively establishes that phages are a major unrecognized force shaping the ecology and intra-species strain-level diversity of the well-studied oral pathogen Pg. The foundational phage sequence datasets and model systems that we establish here add to the rich context of all that is already known about Pg, and point to numerous avenues of future inquiry that promise to shed new light on fundamental features of phage impacts on human health and disease broadly. Video Abstract.

Microbiota present in combined endodontic-periodontal diseases and its risks for endocarditis.

INTRODUCTION: Infective endocarditis (IE) is an inflammatory disease usually caused by bacteria that enter the bloodstream and establish infections in the inner linings or valves of the heart, including blood vessels. Despite the availability of modern antimicrobial and surgical treatments, IE continues to cause substantial morbidity and mortality. Oral microbiota is considered one of the most significant risk factors for IE. The objective of this study was to evaluate the microbiota present in root canal (RC) and periodontal pocket (PP) clinical samples in cases with combined endo-periodontal lesions (EPL) to detect species related to IE using NGS. METHODS: Microbial samples were collected from 15 RCs and their associated PPs, also from 05 RCs with vital pulp tissues (negative control, NC). Genomic studies associated with bioinformatics, combined with structuring of a database (genetic sequences of bacteria reported for infective endocarditis), allowed for the assessment of the microbial community at both sites. Functional prediction was conducted using PICRUSt2. RESULTS: Parvimonas, Streptococcus, and Enterococcus were the major genera detected in the RCs and PPs. A total of 79, 96, and 11 species were identified in the RCs, PPs, and NCs, respectively. From them, a total of 34 species from RCs, 53 from PPs, and 2 from NCs were related to IE. Functional inference demonstrated that CR and PP microbiological profiles may not be the only risk factors for IE but may also be associated with systemic diseases, including myocarditis, human cytomegalovirus infection, bacterial invasion of epithelial cells, Huntington's disease, amyotrophic lateral sclerosis, and hypertrophic cardiomyopathy. Additionally, it was possible to predict antimicrobial resistance variants for broad-spectrum drugs, including ampicillin, tetracycline, and macrolides. CONCLUSION: Microorganisms present in the combined EPL may not be the only risk factor for IE but also for systemic diseases. Antimicrobial resistance variants for broad-spectrum drugs were inferred based on PICRUSt-2. State-of-the-art sequencing combined with bioinformatics has proven to be a powerful tool for conducting studies on microbial communities and could considerably assist in the diagnosis of serious infections. CLINICAL RELEVANCE: Few studies have investigated the microbiota in teeth compromised by combined endo-periodontal lesions (EPL), but none have correlated the microbiological findings to any systemic condition, particularly IE, using NGS techniques. In such cases, the presence of apical periodontitis and periodontal disease can increase IE risk in susceptible patients.

Integrative Metatranscriptomic Analysis Reveals Disease-specific Microbiome-host Interactions in Oral Squamous Cell Carcinoma.

Studies on the microbiome of oral squamous cell carcinoma (OSCC) have been limited to 16S rRNA gene sequencing. Here, laser microdissection coupled with brute-force, deep metatranscriptome sequencing was employed to simultaneously characterize the microbiome and host transcriptomes and predict their interaction in OSCC. The analysis involved 20 HPV16/18-negative OSCC tumor/adjacent normal tissue pairs (TT and ANT) along with deep tongue scrapings from 20 matched healthy controls (HC). Standard bioinformatic tools coupled with in-house algorithms were used to map, analyze, and integrate microbial and host data. Host transcriptome analysis identified enrichment of known cancer-related gene sets, not only in TT versus ANT and HC, but also in the ANT versus HC contrast, consistent with field cancerization. Microbial analysis identified a low abundance yet transcriptionally active, unique multi-kingdom microbiome in OSCC tissues predominated by bacteria and bacteriophages. HC showed a different taxonomic profile yet shared major microbial enzyme classes and pathways with TT/ANT, consistent with functional redundancy. Key taxa enriched in TT/ANT compared with HC were Cutibacterium acnes, Malassezia restricta, Human Herpes Virus 6B, and bacteriophage Yuavirus. Functionally, hyaluronate lyase was overexpressed by C. acnes in TT/ANT. Microbiome-host data integration revealed that OSCC-enriched taxa were associated with upregulation of proliferation-related pathways. In a preliminary in vitro validation experiment, infection of SCC25 oral cancer cells with C. acnes resulted in upregulation of MYC expression. The study provides a new insight into potential mechanisms by which the microbiome can contribute to oral carcinogenesis, which can be validated in future experimental studies. Significance: Studies have shown that a distinct microbiome is associated with OSCC, but how the microbiome functions within the tumor interacts with the host cells remains unclear. By simultaneously characterizing the microbial and host transcriptomes in OSCC and control tissues, the study provides novel insights into microbiome-host interactions in OSCC which can be validated in future mechanistic studies.

Specific host metabolite and gut microbiome alterations are associated with bone loss during spaceflight.

Understanding the axis of the human microbiome and physiological homeostasis is an essential task in managing deep-space-travel-associated health risks. The NASA-led Rodent Research 5 mission enabled an ancillary investigation of the gut microbiome, varying exposure to microgravity (flight) relative to ground controls in the context of previously shown bone mineral density (BMD) loss that was observed in these flight groups. We demonstrate elevated abundance of Lactobacillus murinus and Dorea sp. during microgravity exposure relative to ground control through whole-genome sequencing and 16S rRNA analyses. Specific functionally assigned gene clusters of L. murinus and Dorea sp. capable of producing metabolites, lactic acid, leucine/isoleucine, and glutathione are enriched. These metabolites are elevated in the microgravity-exposed host serum as shown by liquid chromatography-tandem mass spectrometry (LC-MS/MS) metabolomic analysis. Along with BMD loss, ELISA reveals increases in osteocalcin and reductions in tartrate-resistant acid phosphatase 5b signifying additional loss of bone homeostasis in flight.

Longitudinal changes in subgingival biofilm composition following periodontal treatment.

BACKGROUND: Current periodontal treatment involves instrumentation using hand and/or ultrasonic instruments, which are used either alone or in combination based on patient and clinician preference, with comparable clinical outcomes. This study sought to investigate early and later changes in the subgingival biofilm following periodontal treatment, to identify whether these changes were associated with treatment outcomes, and to investigate whether the biofilm responded differently to hand compared with ultrasonic instruments. METHODS: This was a secondary-outcome analysis of a randomized-controlled trial. Thirty-eight periodontitis patients received full-mouth subgingival instrumentation using hand (n = 20) or ultrasonic instrumentation (n = 18). Subgingival plaque was sampled at baseline and 1, 7, and 90 days following treatment. Bacterial DNA was analyzed using 16S rRNA sequencing. Periodontal clinical parameters were evaluated before and after treatment. RESULTS: Biofilm composition was comparable in both (hand and ultrasonics) treatment groups at all time points (all genera and species; p[adjusted] > 0.05). Large-scale changes were observed within groups across time points. At days 1 and 7, taxonomic diversity and dysbiosis were reduced, with an increase in health-associated genera including Streptococcus and Rothia equating to 30% to 40% of the relative abundance. When reassessed at day 90 a subset of samples reformed a microbiome more comparable with baseline, which was independent of instrumentation choice and residual disease. CONCLUSIONS: Hand and ultrasonic instruments induced comparable impacts on the subgingival plaque microbiome. There were marked early changes in the subgingival biofilm composition, although there was limited evidence that community shifts associated with treatment outcomes.

Optimization of conditions for in vitro modeling of subgingival normobiosis and dysbiosis.

Modeling subgingival microbiome in health and disease is key to identifying the drivers of dysbiosis and to studying microbiome modulation. Here, we optimize growth conditions of our previously described in vitro subgingival microbiome model. Subgingival plaque samples from healthy and periodontitis subjects were used as inocula to grow normobiotic and dysbiotic microbiomes in MBEC assay plates. Saliva supplemented with 1%, 2%, 3.5%, or 5% (v/v) heat-inactivated human serum was used as a growth medium under shaking or non-shaking conditions. The microbiomes were harvested at 4, 7, 10 or 13 days of growth (384 microbiomes in total) and analyzed by 16S rRNA gene sequencing. Biomass significantly increased as a function of serum concentration and incubation period. Independent of growth conditions, the health- and periodontitis-derived microbiomes clustered separately with their respective inocula. Species richness/diversity slightly increased with time but was adversely affected by higher serum concentrations especially in the periodontitis-derived microbiomes. Microbial dysbiosis increased with time and serum concentration. Porphyromonas and Alloprevotella were substantially enriched in higher serum concentrations at the expense of Streptococcus, Fusobacterium and Prevotella. An increase in Porphyromonas, Bacteroides and Mogibacterium accompanied by a decrease in Prevotella, Catonella, and Gemella were the most prominent changes over time. Shaking had only minor effects. Overall, the health-derived microbiomes grown for 4 days in 1% serum, and periodontitis-derived microbiomes grown for 7 days in 3.5%-5% serum were the most similar to the respective inocula. In conclusion, normobiotic and dysbiostic subgingival microbiomes can be grown reproducibly in saliva supplemented with serum, but time and serum concentration need to be adjusted differently for the health and periodontitis-derived microbiomes to maximize similarity to in vivo inocula. The optimized model could be used to identify drivers of dysbiosis, and to evaluate interventions such as microbiome modulators.

Prolonged mask wearing does not alter the oral microbiome, salivary flow rate or gingival health status - A pilot study.

The COVID-19 pandemic has resulted in the widespread use of N95 respirators and surgical masks, with anecdotal reports among healthcare providers and the public of xerostomia, halitosis, and gingivitis, a consortium of symptoms colloquially termed "mask mouth". However, this has not been scientifically verified. The aim of this study was to assess changes in salivary flow rate, gingival health status and oral microbiome associated with prolonged mask use. A total of 25 dental students (mean age = 26.36 ± 1.58) were included in the study and evaluated at three time points: T1, at the end of at least 2 months of full-day mask wear (7.26 ± 1.56 hours/day); T2, at the end of a period of minimal mask use (1.13 ± 1.13 hours/day); and T3, at the end of 2-3 weeks of resuming full-day mask wear (6.93 ± 1.80 hours/day). Unstimulated whole saliva (UWS) flow rate, xerostomia (on a quantitative scale of 10), gingival index (GI) and plaque index (PI) were assessed at each time point. The salivary microbiome was characterized using 16S rRNA gene sequencing. Overall, UWS flow rates were normal (mean of 0.679 ml/min) and xerostomia, PI and GI scores were low (Mean of 3.11, 0.33 and 0.69, respectively) with no significant differences as a result of prolonged mask wearing. Similarly, there were no significant microbial changes at a false discovery rate (FDR) ≤ 0.05. However, some trends were identified using a nominal p-value cut-off of ≤ 0.01, namely Gemella sanguinis, Streptococcus sp. Oral taxon 066 and Oral taxon 058 were associated with prolonged mask wear. Trends were also seen by gender, race and age, for example an increase in P. gingivalis and P. intermedia with age. In conclusion, we found no evidence that prolonged mask wear adversely affects oral health. The findings support that the oral microbiome of healthy individuals is resilient.

Multimodal Data Integration Reveals Mode of Delivery and Snack Consumption Outrank Salivary Microbiome in Association With Caries Outcome in Thai Children.

Early childhood caries (ECC) is not only the most common chronic childhood disease but also disproportionately affects underserved populations. Of those, children living in Thailand have been found to have high rates of ECC and severe ECC. Frequently, the cause of ECC is blamed on a handful of cariogenic organisms, such as Streptococcus mutans and Streptococcus sobrinus. However, ECC is a multifactorial disease that results from an ecological shift in the oral cavity from a neutral pH (~7.5) to an acidic pH (<5.5) environment influenced by the host individual's biological, socio-behavioral, and lifestyle factors. Currently, there is a lack of understanding of how risk factors at various levels influence the oral health of children at risk. We applied a statistical machine learning approach for multimodal data integration (parallel and hierarchical) to identify caries-related multiplatform factors in a large cohort of mother-child dyads living in Chiang Mai, Thailand (N=177). Whole saliva (1 mL) was collected from each individual for DNA extraction and 16S rRNA sequencing. A set of maternal and early childhood factors were included in the data analysis. Significantly, vaginal delivery, preterm birth, and frequent sugary snacking were found to increase the risk for ECC. The salivary microbial diversity was significantly different in children with ECC or without ECC. Results of linear discriminant analysis effect size (LEfSe) analysis of the microbial community demonstrated that S. mutans, Prevotella histicola, and Leptotrichia hongkongensis were significantly enriched in ECC children. Whereas Fusobacterium periodonticum was less abundant among caries-free children, suggesting its potential to be a candidate biomarker for good oral health. Based on the multimodal data integration and statistical machine learning models, the study revealed that the mode of delivery and snack consumption outrank salivary microbiome in predicting ECC in Thai children. The biological and behavioral factors may play significant roles in the microbial pathobiology of ECC and warrant further investigation.

Chemomechanical preparation influences the microbial community and the levels of LPS, LTA and cytokines in combined endodontic-periodontal lesions: A clinical study.

BACKGROUND: This study was conducted to compare the microbiomes, the levels of lipopolysaccharides (LPS), lipoteichoic acid (LTA), and cytokines (interleukin [IL]-1ß and tumor necrosis factor-alpha [TNF-α]), before and after chemomechanical preparation (CMP) of the root canals (RC) and their associated periodontal pockets (PP) in teeth with combined EPL. MATERIALS: Samples were taken from 10 RC and PP, before and after CMP. The microbiomes (next-generation sequencing, V3-V4 hypervariable region of the 16S rRNA gene), microbiome diversity (bioinformatics analyses), LPS (limulus amebocyte lysate), LTA, IL-1ß, and TNF-α (ELISA) were evaluated. A statistical analysis was performed with significance level set at 5%. RESULTS: The most abundant phyla in both sites were Firmicutes and Proteobacteria. Comparative studies of bacterial genera species revealed that some increased and others decreased after CMP at both sites. A 3% reduction in Gram-negative bacteria (RC) and a 4% increase in Gram-positive bacteria (PP) were detected. LPS levels were 4.4 times higher in PP than in the RC. LTA was detected in all samples investigated. Higher levels of IL-1ß and TNF-α were detected in both sites at baseline. After CMP, LPS, LTA, IL-1ß and TNF-α were reduced in both sites. CONCLUSION: The microbial community in the RC and PP in teeth with combined EPL indicated a similarity between both sites. CMP effectively reduced the microbial load and the LPS levels from teeth with EPL, and consequently diminished the cytokine levels. The reduction in LTA levels in the RC and PP proved challenging.

Complete Genome Sequence of Arachnia rubra Strain DSM 100122T, a Cultured Member of the Human Oral Microbiome.

We report the complete genome of Arachnia rubra strain DSM 100122T. The genome is 3.32 Mb, with a GC content of 64.2%. The genome contains 3,005 predicted genes, including 2,923 predicted protein-coding genes.

Pseudomonas aeruginosa transcriptome adaptations from colonization to biofilm infection of skin wounds.

In burn patients Pseudomonas aeruginosa infection is a major cause of morbidity. Analysis of the pathogen's gene expression as it transitions from colonization to acute and then biofilm wound infection may provide strategies for infection control. Toward this goal, we seeded log-phase P. aeruginosa (PAO1) into 3-day-old, full-thickness excision wounds (rabbit ear) and harvested the bacteria during colonization (Hrs 2 and 6), acute infection (Hr 24), and biofilm infection (Days 5 and 9) for transcriptome analysis (RNA-Seq). After 2-6 h in the wound, genes for metabolism and cell replication were down-regulated while wound-adaptation genes were up-regulated (vs. expression in log-phase culture). As the infection progressed from acute to biofilm infection, more genes became up-regulated than down-regulated, but the down-regulated genes enriched in more pathways, likely because the genes and pathways that bacteria already colonizing wounds up-regulate to establish biofilm infection are less known. Across the stages of infection, carbon-utilization pathways shifted. During acute infection, itaconate produced by myeloid cells appears to have been a carbon source because myeloid cell infiltration and the expression of the host gene, ACOD1, for itaconate production peaked coincidently with the expression of the PAO1 genes for itaconate transport and catabolism. Additionally, branched-chain amino acids are suggested to be a carbon source in acute infection and in biofilm infection. In biofilm infection, fatty acid degradation was also up-regulated. These carbon sources feed into the glyoxylate cycle that was coincidently up-regulated, suggesting it provided the precursors for P. aeruginosa to synthesize macromolecules in establishing wound infection.

Interrelationship between the Microbial Communities of the Root Canals and Periodontal Pockets in Combined Endodontic-Periodontal Diseases.

Periodontal and Endodontic diseases are biofilm-related diseases. The presence of microorganisms in root canals (RCs) and the complex microbiota of periodontal pockets (PPs) contribute to the development of endodontic-periodontal diseases. This study performed a systemic analysis using state-of-the-art sequence data to assess the microbial composition of infected RCs and PPs to further assess the microbiota and verify the possibility of cross-infection between these sites. The microbiomes of these combined diseases were examined with a focus on the V3-V4 hypervariable region of the 16S rRNA gene. The number of species in PP was higher than in RC, and there was a predominance of obligate anaerobes and gram-negative bacteria. In the RCs, the genera Enterococcus, Parvimonas, Stomatobaculum predominated, in contrast, the PPs revealed a predominance of Enterococcus, Parvimonas, Stomatobaculum, Peptostreptococcus and Mogibacterium. The RC and PP microbiome was not similar with regards to the sharing of OTUs for phyla and genera (8 and 67, respectively). The evaluation of molecular markers revealed a large number of markers for resistance to antibiotics of the carbapenem and beta-lactam type (broad spectrum). Another relevant finding of this study was the markers related to systemic diseases related to cardiac muscle and rheumatology, among others. In conclusion, the RC microbiota was less complex and diverse than PP. Interactions between microbial communities were present. The shared genus can signal communication between the endodontic and periodontal microbiomes.

Glycemic fluctuation exacerbates inflammation and bone loss and alters microbiota profile around implants in diabetic mice with experimental peri-implantitis.

BACKGROUND: The impact of glycemic fluctuation under diabetic condition on peri-implantitis in diabetic patients remains unclear. We hypothesized that glycemic fluctuation has greater adverse effect on experimental peri-implantitis, compared with sustained high blood glucose in diabetes. RESULTS: Maxillary left first and second molars of diabetic db/db mice were extracted and were replaced with one dental implant in the healed edentulous space. Glycemic control or fluctuation were managed by constant or interrupted oral administration of rosiglitazone to these mice. Meanwhile, experimental peri-implantitis was induced by ligation around implants. After 14 weeks, inflammatory responses, and peri-implant bone loss, together with oral microbiota profile were analyzed. Diabetic mice with glycemic fluctuation showed greater peri-implant bone loss, inflammatory cell infiltration, and osteoclastogenesis, compared with mice with sustained hyperglycemia. Compared to sustained hyperglycemia, glycemic fluctuation led to further increase in IL-1ß, TNFα, RANKL, TLR2/4, IRAK1, and TRAF6 mRNA expression in peri-implant gingival tissues. Both rosiglitazone-induced glycemic control and glycemic fluctuation caused microbiota profile change in diabetic mice compared to that in uncontrolled hyperglycemic mice. CONCLUSIONS: This study suggests that glycemic fluctuation may aggravate peri-implantitis inflammation and bone loss, which may be associated with a shift in peri-implant microbial profile towards dysbiotic changes and the activation of TLR2/4-IRAK1-TRAF6 signaling.

Association between metabolic syndrome and periodontitis: The role of lipids, inflammatory cytokines, altered host response, and the microbiome.

Periodontitis has been associated with many systemic diseases and conditions, including metabolic syndrome. Metabolic syndrome is a cluster of conditions that occur concomitantly and together they increase the risk of cardiovascular disease and double the risk of type 2 diabetes. In this review, we focus on the association between metabolic syndrome and periodontitis; however, we also include information on diabetes mellitus and cardiovascular disease, since these two conditions are significantly intertwined with metabolic syndrome. With regard to periodontitis and metabolic syndrome, to date, the vast majority of studies point to an association between these two conditions and also demonstrate that periodontitis can contribute to the development of, or can worsen, metabolic syndrome. Evaluating the effect of metabolic syndrome on the salivary microbiome, data presented herein support the hypothesis that the salivary bacterial profile is altered in metabolic syndrome patients compared with healthy patients. Considering periodontitis and these three conditions, the vast majority of human and animal studies point to an association between periodontitis and metabolic syndrome, diabetes, and cardiovascular disease. Moreover, there is evidence to suggest that metabolic syndrome and diabetes can alter the oral microbiome. However, more studies are needed to fully understand the influence these conditions have on each other.