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Journal of Endodontics Aug 2016The purpose of this study was to combine multiple displacement amplification and checkerboard DNA-DNA hybridization to qualitatively and quantitatively evaluate the...
INTRODUCTION
The purpose of this study was to combine multiple displacement amplification and checkerboard DNA-DNA hybridization to qualitatively and quantitatively evaluate the microbiota present in infections refractory to endodontic treatment.
METHODS
The subjects of this study were 40 patients presenting with periapical lesions refractory to endodontic treatment. Samples were taken by scraping or filing root canal walls with a #10 K-type hand file. Sample DNA was amplified by multiple displacement amplification, and the levels of 107 bacterial taxa were analyzed by checkerboard DNA-DNA hybridization. The taxa were divided into 3 distinct microbial populations depending on their mean proportion in samples (% DNA probe counts ± standard error of the mean) as follows: dominant (≥3.0%), subdominant (>1.6%-3.0%), and residual (≤1.6%) populations. The significance of differences was determined using the Mann-Whitney test.
RESULTS
The taxa present with the highest mean proportions (constituting the dominant population) were Corynebacterium diphtheriae (8.03 ± 0.98), Porphyromonas gingivalis (5.42 ± 2.09), Streptococcus sobrinus (5.33 ± 0.69), and Stenotrophomonas maltophilia (4.72 ± 1.73). Among the subdominant population were Eubacterium saphenum (3.85 ± 1.06), Helicobacter pylori (3.16 ± 0.62), Dialister pneumosintes (3.12 ± 1.1), Clostridium difficile (2.74 ± 0.41), Enterobacter agglomerans (2.64 ± 0.54), Salmonella enterica (2.51 ± 0.52), Mobiluncus mulieris (2.44 ± 0.6), and Klebsiella oxytoca (2.32 ± 0.66). In the population of bacteria present at the lowest mean proportions (the residual population), Bacteroides ureolyticus (0.04 ± 0.01), Haemophilus influenzae (0.04 ± 0.02), and Prevotella oris (0.01 ± 0.01) were found at the lowest mean proportions. Enterococcus faecalis was detected in the residual population (0.52 ± 0.26).
CONCLUSIONS
The microbial climax community in teeth refractory to endodontic treatment not only harbors medically important species but also contains distinct microbial consortia present with different population levels.
Topics: Bacteria; DNA Probes; DNA, Bacterial; Dental Pulp Cavity; Dental Pulp Necrosis; Humans; Microbiota; Nucleic Acid Hybridization; Periapical Diseases; Root Canal Therapy
PubMed: 27377440
DOI: 10.1016/j.joen.2016.05.014 -
Journal of Applied Oral Science :... Apr 2016Objective The aim of this study was to evaluate the association of Porphyromonas endodontalis, Filifactor alocis and Dialister pneumosintes with the occurrence of...
Objective The aim of this study was to evaluate the association of Porphyromonas endodontalis, Filifactor alocis and Dialister pneumosintes with the occurrence of periodontitis. Material and Methods Thirty subjects with chronic periodontitis (ChP) and 10 with periodontal health (PH) were included in the study. Nine subgingival biofilm samples were collected as follows: i) PH group - from the mesial/buccal aspect of each tooth in two randomly chosen contralateral quadrants; ii) ChP group - from three sites in each of the following probing depth (PD) categories: shallow (≤3 mm), moderate (4-6 mm) and deep (≥7 mm). Checkerboard DNA-DNA hybridization was used to analyze the samples. Results We found the three species evaluated in a higher percentage of sites and at higher levels in the group with ChP than in the PH group (p<0.05, Mann-Whitney test). We also observed these differences when the samples from sites with PD≤4 mm or ≥5 mm of subjects with ChP were compared with those from subjects with PH (p<0.05, Mann-Whitney test). In addition, the prevalence and levels of D. pneumosintes, and especially of F. alocis were very low in healthy subjects (0.12x105 and 0.01x105, respectively). Conclusion F. alocis and D. pneumosintes might be associated with the etiology of ChP, and their role in the onset and progression of this infection should be further investigated. The role of P. endodontalis was less evident, since this species was found in relatively high levels and prevalence in the PH group.
Topics: Adult; Biofilms; Brazil; Case-Control Studies; Chronic Periodontitis; Colony Count, Microbial; DNA Probes; DNA, Bacterial; Dental Plaque; Female; Gingiva; Humans; Male; Middle Aged; Peptostreptococcus; Porphyromonas endodontalis; Statistics, Nonparametric; Veillonellaceae
PubMed: 27119767
DOI: 10.1590/1678-775720150445 -
Frontiers in Cellular and Infection... 2015Patients with acquired immunodeficiency syndrome (AIDS) are at high risk of opportunistic infections. Oral manifestations have been associated with the level of...
Patients with acquired immunodeficiency syndrome (AIDS) are at high risk of opportunistic infections. Oral manifestations have been associated with the level of immunosuppression, these include periodontal diseases, and understanding the microbial populations in the oral cavity is crucial for clinical management. The aim of this study was to examine the salivary bacterial diversity in patients newly admitted to the AIDS ward of the Public Health Clinical Center (China). Saliva samples were collected from 15 patients with AIDS who were randomly recruited between December 2013 and March 2014. Extracted DNA was used as template to amplify bacterial 16S rRNA. Sequencing of the amplicon library was performed using a 454 GS-FLX Titanium sequencing platform. Reads were optimized and clustered into operational taxonomic units for further analysis. A total of 10 bacterial phyla (106 genera) were detected. Firmicutes, Bacteroidetes, and Proteobacteria were preponderant in the salivary microbiota in AIDS patients. The pathogen, Capnocytophaga sp., and others not considered pathogenic such as Neisseria elongata, Streptococcus mitis, and Mycoplasma salivarium but which may be opportunistic infective agents were detected. Dialister pneumosintes, Eubacterium infirmum, Rothia mucilaginosa, and Treponema parvum were preponderant in AIDS patients with periodontitis. Patients with necrotic periodontitis had a distinct salivary bacterial profile from those with chronic periodontitis. This is the first study using advanced sequencing techniques focused on hospitalized AIDS patients showing the diversity of their salivary microbiota.
Topics: Acquired Immunodeficiency Syndrome; Adult; Bacteria; Biota; China; Cluster Analysis; DNA, Bacterial; DNA, Ribosomal; Female; Gingivitis; Humans; Male; Middle Aged; Periodontal Diseases; Phylogeny; RNA, Ribosomal, 16S; Saliva; Sequence Analysis, DNA
PubMed: 26191508
DOI: 10.3389/fcimb.2015.00055 -
Clinical Oral Investigations Mar 2016The objective of this study was to compare the periodontopathogen prevalence and tetracycline resistance genes in Dominican patients with different periodontal...
OBJECTIVE
The objective of this study was to compare the periodontopathogen prevalence and tetracycline resistance genes in Dominican patients with different periodontal conditions.
METHODS
Seventy-seven samples were collected from healthy, gingivitis, chronic (CP) and aggressive (AgP) periodontitis patients. Porphyromonas gingivalis, Treponema denticola, Tannerella forsythia, Aggregatibacter actinomycetemcomitans, Fusobacterium nucleatum, Prevotella intermedia, Parvimonas micra, Eikenella corrodens and Dialister pneumosintes and 11 resistance genes were studied by PCR. P. gingivalis fimA genotype was determined.
RESULTS
In healthy patients, P. micra and P. intermedia were the most and least frequently detected, respectively. T. forsythia and E. corrodens appeared in 100% of gingivitis patients. Red complex, D. pneumosintes and E. corrodens were significantly more prevalent in CP compared to healthy patients. F. nucleatum and T. denticola were detected more frequently in AgP. A. actinomycetemcomitans was the most rarely observed in all groups. The fimA II genotype was the most prevalent in periodontitis patients. Seven tetracycline-resistant genes were detected. tet(Q), tet(32) and tet(W) showed the greatest prevalence. tet(32) was significantly more prevalent in CP than in healthy patients.
CONCLUSIONS
Red complex bacteria and D. pneumosintes were significantly the most prevalent species among periodontitis patients. T. forsythia was the most frequently detected in this population. To our knowledge, this is the first study describing the tet(32) gene in subgingival biofilm from healthy and periodontally diseased subjects.
CLINICAL RELEVANCE
This study contributes to the knowledge on the subgingival microbiota and its resistance genes of a scarcely studied world region. Knowing the prevalence of resistance genes could impact on their clinical prescription and could raise awareness to the appropriate use of antibiotics.
Topics: Adult; Aged; Bacterial Infections; Biofilms; Case-Control Studies; Dominican Republic; Female; Genotype; Humans; Male; Microbiota; Middle Aged; Periodontitis; Polymerase Chain Reaction; Prevalence; Tetracycline Resistance
PubMed: 26121972
DOI: 10.1007/s00784-015-1516-2 -
Internal Medicine (Tokyo, Japan) 2015A 62-year-old Japanese woman was hospitalized at the Department of Senology for positive signals on two sets of blood cultures obtained in the Emergency Department. The...
A 62-year-old Japanese woman was hospitalized at the Department of Senology for positive signals on two sets of blood cultures obtained in the Emergency Department. The initial physical examination with enhanced computed tomography of the chest and abdomen did not identify the infectious source. Dialister pneumosintes was identified on 16S rRNA sequencing, and dental caries with sinusitis were subsequently diagnosed based on a dental examination and magnetic resonance imaging. History taking with respect to dental hygiene and oral examinations should be performed in daily clinical practice, especially in immunosuppressed patients.
Topics: Anti-Bacterial Agents; Bacteremia; Dental Caries; Female; Humans; Middle Aged; RNA, Ribosomal, 16S; Sinusitis; Treatment Outcome; Veillonellaceae
PubMed: 25786460
DOI: 10.2169/internalmedicine.54.2904 -
Journal of Oral Microbiology 2014We recently reported that large amounts of oral bacterial DNA can be found in thrombus aspirates of myocardial infarction patients. Some case reports describe bacterial...
BACKGROUND
We recently reported that large amounts of oral bacterial DNA can be found in thrombus aspirates of myocardial infarction patients. Some case reports describe bacterial findings in pericardial fluid, mostly done with conventional culturing and a few with PCR; in purulent pericarditis, nevertheless, bacterial PCR has not been used as a diagnostic method before.
OBJECTIVE
To find out whether bacterial DNA can be measured in the pericardial fluid and if it correlates with pathologic-anatomic findings linked to cardiovascular diseases.
METHODS
Twenty-two pericardial aspirates were collected aseptically prior to forensic autopsy at Tampere University Hospital during 2009-2010. Of the autopsies, 10 (45.5%) were free of coronary artery disease (CAD), 7 (31.8%) had mild and 5 (22.7%) had severe CAD. Bacterial DNA amounts were determined using real-time quantitative PCR with specific primers and probes for all bacterial strains associated with endodontic disease (Streptococcus mitis group, Streptococcus anginosus group, Staphylococcus aureus/Staphylococcus epidermidis, Prevotella intermedia, Parvimonas micra) and periodontal disease (Aggregatibacter actinomycetemcomitans, Porphyromonas gingivalis, Treponema denticola, Fusobacterium nucleatus, and Dialister pneumosintes).
RESULTS
Of 22 cases, 14 (63.6%) were positive for endodontic and 8 (36.4%) for periodontal-disease-associated bacteria. Only one case was positive for bacterial culturing. There was a statistically significant association between the relative amount of bacterial DNA in the pericardial fluid and the severity of CAD (p=0.035).
CONCLUSIONS
Oral bacterial DNA was detectable in pericardial fluid and an association between the severity of CAD and the total amount of bacterial DNA in pericardial fluid was found, suggesting that this kind of measurement might be useful for clinical purposes.
PubMed: 25412607
DOI: 10.3402/jom.v6.25835 -
Oral Diseases Sep 2012To compare the microbiota of endodontic infections in necrotic pulp from HIV-negative and HIV-positive subjects. (Comparative Study)
Comparative Study
OBJECTIVE
To compare the microbiota of endodontic infections in necrotic pulp from HIV-negative and HIV-positive subjects.
MATERIALS AND METHODS
Root canal samples from necrotic pulp were collected from 40 HIV- and 20 HIV+ subjects. Pulps were amplified using multiple displacement amplification (MDA). Then, checkerboard DNA-DNA hybridization was employed to assess the levels of 107 microbial taxa. The percentage of DNA probe count and the percentage of teeth colonized by each test species were investigated. Significant differences between groups regarding proportions of taxa and prevalence of the test species were sought using the Mann-Whitney test and the Chi-square analysis, respectively.
RESULTS
The most prevalent taxa detected were Dialister pneumosintes, Stenotrophomonas maltophilia, Streptococcus sobrinus, Corynebacterium diphteriae, and Helicobacter pylori among HIV- subjects and D. pneumosintes, Prevotella tannerae, Porphyromonas gingivalis, Parvimonas micra, Prevotella nigrescens, and Corynebacterium diphtheriae among HIV+ individuals. D. pneumosintes, C. diphtheria, and C. albicans were the most abundant species in the HIV- group, whereas the predominant taxa in HIV+ samples were P. tannerae, D. pneumosintes and Olsenella uli. P. tannerae, O. uli, Veilonella dispar, Bacteroides fragilis, and Actinomyces meyeri were significantly more abundant in HIV+ samples.
CONCLUSIONS
There were significant differences in the prevalence and proportions of specific microbial taxa between HIV- and HIV+ individuals. The root canal microbiota may represent a reservoir of important oral and medical pathogens, mainly in HIV+ individuals.
Topics: Actinomyces; Adolescent; Adult; Bacteria; Bacteroides fragilis; Candida albicans; Child; Corynebacterium diphtheriae; DNA Probes; Dental Pulp Cavity; Dental Pulp Necrosis; Female; Gram-Negative Anaerobic Straight, Curved, and Helical Rods; HIV Seronegativity; HIV Seropositivity; Helicobacter pylori; Humans; Male; Middle Aged; Nucleic Acid Amplification Techniques; Nucleic Acid Hybridization; Peptostreptococcus; Porphyromonas gingivalis; Prevotella; Prevotella nigrescens; Stenotrophomonas maltophilia; Streptococcus sobrinus; Veillonella; Young Adult
PubMed: 22335194
DOI: 10.1111/j.1601-0825.2012.01908.x -
Oral Surgery, Oral Medicine, Oral... Aug 2011Viral-bacterial and bacterial synergism have been suggested to contribute to the pathogenesis of several human diseases. This study sought to investigate the possible...
OBJECTIVE
Viral-bacterial and bacterial synergism have been suggested to contribute to the pathogenesis of several human diseases. This study sought to investigate the possible associations between 9 candidate endodontic bacterial pathogens and 9 human viruses in samples from acute apical abscesses.
STUDY DESIGN
DNA extracts from purulent exudate aspirates of 33 cases of acute apical abscess were surveyed for the presence of 9 selected bacterial species using a 16S ribosomal RNA gene-based nested polymerase chain reaction (PCR) approach. Single or nested PCR assays were used for detection of the human papillomavirus (HPV) and herpesviruses types 1 to 8.
RESULTS
Two-thirds of the abscess samples were positive for at least one of the target viruses. Specifically, the most frequently detected viruses were HHV-8 (54.5%); HPV (9%); and varicella zoster virus (VZV), Epstein-Barr virus (EBV), and HHV-6 (6%). Bacterial DNA was present in all cases and the most prevalent bacterial species were Treponema denticola (70%), Tannerella forsythia (67%), Porphyromonas endodontalis (67%), Dialister invisus (61%), and Dialister pneumosintes (57.5%). HHV-8 was positively associated with 7 of the target bacterial species and HPV with 4, but all these associations were weak. Several bacterial pairs showed a moderate positive association. Viral coinfection was found in 6 abscess cases, but no significant viral association could be determined.
CONCLUSIONS
Findings demonstrated that bacterial and viral DNA occurred concomitantly in two-thirds of the samples from endodontic abscesses. Although this may suggest a role for viruses in the etiology of apical abscesses, the possibility also exists that the presence of viruses in abscess samples is merely a consequence of the bacterially induced disease process. Further studies are necessary to clarify the role of these viral-bacterial interactions, if any, in the pathogenesis of acute apical abscesses.
Topics: Adolescent; Adult; Bacteroidaceae Infections; Bacteroides Infections; DNA, Bacterial; DNA, Viral; Epstein-Barr Virus Infections; Gram-Negative Anaerobic Straight, Curved, and Helical Rods; Gram-Negative Bacterial Infections; Herpes Zoster; Herpesviridae; Herpesviridae Infections; Herpesvirus 6, Human; Herpesvirus 8, Human; Humans; Middle Aged; Papillomavirus Infections; Periapical Abscess; Polymerase Chain Reaction; Porphyromonas gingivalis; RNA, Bacterial; RNA, Ribosomal, 16S; Roseolovirus Infections; Treponema denticola; Treponemal Infections; Young Adult
PubMed: 21507684
DOI: 10.1016/j.tripleo.2011.01.029 -
Journal of Dental Research Nov 2010The subgingival microbiome is largely uncultivated, and therefore, cultivation-based and targeted molecular approaches have limited value in examining the effect of... (Comparative Study)
Comparative Study
The subgingival microbiome is largely uncultivated, and therefore, cultivation-based and targeted molecular approaches have limited value in examining the effect of smoking on this community. We tested the hypothesis that the subgingival biofilm is compositionally different in current and never-smokers by using an open-ended molecular approach for bacterial identification. Subgingival plaque from deep sites of current and never-smokers matched for disease was analyzed by 16S sequencing. Smokers demonstrated greater abundance of Parvimonas, Fusobacterium, Campylobacter, Bacteroides, and Treponema and lower levels of Veillonella, Neisseria, and Streptococcus. Several uncultivated Peptostreptococci, Parvimonas micra, Campylobacter gracilis, Treponema socranskii, Dialister pneumosintes, and Tannerella forsythia were elevated in this group, while Veillonella sp. oral clone B2, Neisseria sp. oral clone 2.24, Streptococcus sanguinis, and Capnocytophaga sp. clone AH015 were at lower levels. The microbial profile of smoking-associated periodontitis is distinct from that of non-smokers, with significant differences in the prevalence and abundance of disease-associated and health-compatible organisms.
Topics: Alveolar Bone Loss; Bacteria; Bacteroides; Biofilms; Campylobacter; Capnocytophaga; Case-Control Studies; Chronic Periodontitis; Dental Plaque; Female; Fusobacterium; Gingiva; Gram-Negative Anaerobic Straight, Curved, and Helical Rods; Humans; Male; Middle Aged; Neisseria; Peptostreptococcus; Periodontal Attachment Loss; Periodontal Pocket; Smoking; Streptococcus; Treponema; Veillonella
PubMed: 20739702
DOI: 10.1177/0022034510377203 -
Journal of Clinical Microbiology Jul 2010It has been demonstrated that smoking cessation alters the subgingival microbial profile; however, the response of individual bacteria within this ecosystem has not been...
It has been demonstrated that smoking cessation alters the subgingival microbial profile; however, the response of individual bacteria within this ecosystem has not been well studied. The aim of this investigation, therefore, was to longitudinally examine the effect of smoking cessation on the prevalence and levels of selected subgingival bacteria using molecular approaches for bacterial identification and enumeration. Subgingival plaque was collected from 22 smokers at the baseline and 12 months following periodontal nonsurgical management and smoking cessation counseling. The prevalence and abundance of selected organisms were examined using nested PCR and multiplexed bead-based flow cytometry. Eleven subjects successfully quit smoking over 12 months (quitters), while 11 continued to smoke throughout (smokers). Smoking cessation led to a decrease in the prevalence of Porphyromonas endodontalis and Dialister pneumosintes at 12 months and in the levels of Parvimonas micra, Filifactor alocis, and Treponema denticola. Smoking cessation also led to an increase in the levels of Veillonella parvula. Following nonsurgical periodontal therapy and smoking cessation, the subgingival microbiome is recolonized by a greater number of health-associated species and there are a significantly lower prevalence and abundance of putative periodontal pathogens. The results indicate a critical role for smoking cessation counseling in periodontal therapy for smokers in order to effectively alter the subgingival microbiome.
Topics: Analysis of Variance; Bacteria; Cross-Sectional Studies; DNA, Bacterial; Gingiva; Humans; Polymerase Chain Reaction; Smoking; Smoking Cessation; Statistics, Nonparametric
PubMed: 20410352
DOI: 10.1128/JCM.01821-09