Yuntao Jiang

Prevalence and molecular diversity of Archaea in subgingival pockets of periodontitis patients

INTRODUCTION The aim of this study was to investigate the prevalence and molecular diversity of Archaea in the subgingival crevices of patients with chronic periodontitis. METHODS Subgingival plaque was collected from 41 patients with chronic periodontitis and 15 healthy subjects. The prevalence of Archaea in those plaque samples was tested by polymerase chain reaction with two broad-range archaeal primer sets. Amplicons from eight Archaea-positive plaque samples were cloned and sequenced for molecular diversity analysis using one of these two primer sets and a novel third primer set. RESULTS Archaea were detected in the subgingival plaque of patients with chronic periodontitis at a prevalence of 70.7-73.2%, but were not detected in healthy subjects. Using one primer set, all sequences of the archaeal amplicons were identified as Methanobrevibacter oralis-like species. With another primer set, the amplicons were also found to be identical to the uncultured M. oralis-like species except one phylotype was found to belong to the class Thermoplasmata. CONCLUSION Archaea might be correlated with periodontal diseases. The diversity of Archaea associated with periodontitis was limited. Almost all sequenced amplicons fell into the genus Methanobrevibacter of the Euryarcheota phylum. M. oralis-like species was the predominant but non-exclusive archaeon in the subgingival dental plaque of patients with periodontitis.

Use of the quorum sensing inhibitor furanone C-30 to interfere with biofilm formation by Streptococcus mutans and its luxS mutant strain.

Streptococcus mutans is recognised as a major aetiological agent of dental caries. One of its important virulence factors is its ability to form biofilms on tooth surfaces. The aim of this study was to evaluate the effects of the quorum sensing inhibitor furanone C-30 on biofilm formation by S. mutans and its luxS mutant strain. The effects of furanone C-30 on biofilms of both strains formed on 96-well microtitre plates at 37 °C were determined by a colorimetric technique (MTT assay). Different concentrations of furanone C-30 (0.0, 2.0 and 4.0 μg/mL) and different time points of biofilm formation (4, 14 and 24 h) were investigated. The structures and thickness of the biofilms were observed by confocal laser scanning microscopy (CLSM). Quorum sensing-related gene expression (ftf, smu630, brpA, gbpB, gtfB, vicR, comDE and relA) was investigated by real-time polymerase chain reaction (RT-PCR). The results showed that synthetic furanone C-30 can inhibit biofilm formation by S. mutans and its luxS mutant strain, although it does not affect the bacterial growth rate itself. The quantities of biofilm formed by both strains significantly decreased (P<0.05) and the biofilms became thinner and looser as revealed by CLSM with increasing concentrations of furanone C-30. Expression of the genes tested was downregulated in the biofilms by the addition of furanone C-30. These results revealed that synthetic furanone C-30 can effectively inhibit biofilm formation by S. mutans and its luxS mutant strain.

Exploring the Dynamic Core Microbiome of Plaque Microbiota during Head-and-Neck Radiotherapy Using Pyrosequencing

Radiotherapy is the primary treatment modality used for patients with head-and-neck cancers, but inevitably causes microorganism-related oral complications. This study aims to explore the dynamic core microbiome of oral microbiota in supragingival plaque during the course of head-and-neck radiotherapy. Eight subjects aged 26 to 70 were recruited. Dental plaque samples were collected (over seven sampling time points for each patient) before and during radiotherapy. The V1–V3 hypervariable regions of bacterial 16S rRNA genes were amplified, and the high-throughput pyrosequencing was performed. A total of 140 genera belonging to 13 phyla were found. Four phyla (Actinobacteria, Bacteroidetes, Firmicutes, and Proteobacteria) and 11 genera (Streptococcus, Actinomyces, Veillonella, Capnocytophaga, Derxia, Neisseria, Rothia, Prevotella, Granulicatella, Luteococcus, and Gemella) were found in all subjects, supporting the concept of a core microbiome. Temporal variation of these major cores in relative abundance were observed, as well as a negative correlation between the number of OTUs and radiation dose. Moreover, an optimized conceptual framework was proposed for defining a dynamic core microbiome in extreme conditions such as radiotherapy. This study presents a theoretical foundation for exploring a core microbiome of communities from time series data, and may help predict community responses to perturbation as caused by exposure to ionizing radiation.

Preliminary study of the presence and association of bacteria and archaea in teeth with apical periodontitis

AIM To investigate, by reverse transcription polymerase chain reaction (RT-PCR), the presence and association of bacteria and archaea in primary and secondary root canal infections. METHODOLOGY A total of 77 root canal samples from 77 Chinese patients, 42 with necrotic pulp tissues (primary infection) and 35 with failed prior conventional root canal treatment (secondary infection), aseptically exposed at the first patient visit, were studied. Total RNA was isolated directly from each sample, and 16S rRNA gene-based RT-PCR assays were used to determine the presence of bacteria and archaea, respectively. RESULTS Bacteria were detected in 39/42 (93%) of root canal samples from teeth with primary infections, and archaea in 16/42 (38%). In the cases diagnosed as secondary root-infected canals, bacteria were detected in 30/35 (86%), whilst archaea were detected in 6/35 (17%) of cases. Amongst the canals, which were positive for bacteria, archaea were always found in combination with bacteria. The incidence of symptomatic cases positive for both bacteria and archaea (16/22, 73%) were significantly higher than those positive for bacteria alone (21/47, 45%) (P < 0.05). CONCLUSIONS This study confirms the presence of archaea in root canal infections and further implicates them in an association with clinical symptoms. The nature of this association requires further study.

Characterization of oral bacterial diversity of irradiated patients by high-throughput sequencing

The objective of this study was to investigate the compositional profiles and microbial shifts of oral microbiota during head-and-neck radiotherapy. Bioinformatic analysis based on 16S rRNA gene pyrosequencing was performed to assess the diversity and variation of oral microbiota of irradiated patients. Eight patients with head and neck cancers were involved in this study. For each patient, supragingival plaque samples were collected at seven time points before and during radiotherapy. A total of 147 232 qualified sequences were obtained through pyrosequencing and bioinformatic analysis, representing 3 460 species level operational taxonomic units (OTUs) and 140 genus level taxa. Temporal variations were observed across different time points and supported by cluster analysis based on weighted UniFrac metrics. Moreover, the low evenness of oral microbial communities in relative abundance was revealed by Lorenz curves. This study contributed to a better understanding of the detailed characterization of oral bacterial diversity of irradiated patients.

luxS Mutant Regulation: Quorum Sensing Impairment or Methylation Disorder?

AI-2–mediated quorum sensing has been identified in various bacteria, including both Gram-negative and Gram-positive species, and numerous phenotypes have been reported to be regulated by this mechanism, using the luxS-mutant strain. But the AI-2 production process confused this regulatory function; some considered this regulation as the result of a metabolic change, which refers to an important metabolic cycle named activated methyl cycle (AMC), caused by luxS-mutant simultaneously with the defect of AI-2. Herein we hypothesized that the quorum sensing system—not the metabolic aspect—is responsible for such a regulatory function. In this study, we constructed plasmids infused with sahH and induced protein expression in the luxS-mutant strain to make the quorum-sensing system and metabolic system independent. The biofilm-related genes were investigated by real-time polymerase chain reaction (PCR), and the results demonstrated that the quorum-sensing completed strain restored the gene expression of the defective strain, but the metabolically completed one did not. This evidence supported our hypothesis that the autoinducer-2-mediated, quorum-sensing system, not the AMC, was responsible for luxS mutant regulation.

Modeling of diffusion transport through oral biofilms with the inverse problem method.

AimThe purpose of this study was to develop a mathematical model to quantitatively describe the passive transport of macromolecules within dental biofilms.MethodologyFluorescently labeled dextrans with different molecular mass (3 kD, 10 kD, 40 kD, 70 kD, 2 000 kD) were used as a series of diffusion probes. Streptococcus mutans, Streptococcus sanguinis, Actinomyces naeslundii and Fusobacterium nucleatum were used as inocula for biofilm formation. The diffusion processes of different probes through the in vitro biofilm were recorded with a confocal laser microscope.ResultsMathematical function of biofilm penetration was constructed on the basis of the inverse problem method. Based on this function, not only the relationship between average concentration of steady‐state and molecule weights can be analyzed, but also that between penetrative time and molecule weights.ConclusionThis can be used to predict the effective concentration and the penetrative time of anti‐biofilm medicines that can diffuse through oral biofilm. Furthermore, an improved model for large molecule is proposed by considering the exchange time at the upper boundary of the dental biofilm.

Stimulation of bone formation in the expanding premaxillary suture with a GSK‐3β inhibitor

OBJECTIVES Glycogen synthase kinase-3β (GSK-3β)/β-catenin signaling mediates osteogenesis in response to mechanical loading. We tested the hypothesis that local administration of a GSK-3β inhibitor could stimulate new bone formation in the expanding premaxillary suture. MATERIALS AND METHODS Thirty-five Sprague-Dawley rats were subjected to premaxillary suture expansion using a helix spring. The experimental rats were given one or two local injections of SB-415286, a small-molecule GSK-3β inhibitor. Animals were administered calcein and sacrificed on day 7 to quantify new bone formation. To evaluate the proliferation and differentiation of osteoblasts, rats were labeled with bromodeoxyuridine on day 1 and sacrificed on day 2 or 4. β-catenin expression was evaluated by immunohistochemical staining. RESULTS Two injections of SB-415286 led to an elevation of β-catenin expression and an increase in the number of proliferating osteoblasts in expanding sutures on day 2 and day 4. Consequently, new bone formation in the suture increased significantly on day 7. CONCLUSIONS These results suggest that local delivery of a GSK-3β inhibitor could stimulate bone formation in the expanding premaxillary suture by eliciting β-catenin signaling. GSK-3β could be a pharmaceutical target for improving the effect of orthodontic treatments such as rapid palatal expansion.

Denaturing gradient gel electrophoresis analysis with different primers of subgingival bacterial communities under mechanical debridement

DGGE of 16S rDNA is one of the most frequently used methods to study microbial communities. In this study, the DGGE profiles of different 16S rDNA regions of the periodontal pathogens Porphyromonas gingivalis, Fusobacterium nucleatum, and Prevotella nigrescens were investigated. The results suggested that V3‐V5 and V6‐V8 fragments may be suitable for community analysis of subgingival bacteria. Further analysis of subgingival samples with V3‐V5 and V6‐V8 regions as target fragments suggested that, in chronic periodontitis, re‐colonization by periodontal bacteria with a population very similar to the baseline may occur by 6 weeks after mechanical debridement.