No abstract available.
Cells, Cultured ; Prevotella ; Prevotella nigrescens

PURPOSE: Interleukin-8 (IL-8) is an important mediator of immune and inflammatory reactions and is produced by a variety of different cell types. This study was undertaken to investigate the effects of lipopolysaccharides (LPSs) from Prevotella intermedia and Prevotella nigrescens, the major causes of inflammatory periodontal disease, on the production of IL-8 and the expression of IL-8 mRNA in differentiated THP-1 cells, a human monocytic cell line. METHODS:LPSs from P. intermedia ATCC 25611 and P. nigrescens ATCC 33563 were prepared by the standard hot phenol-water method. THP-1 cells were incubated in the medium supplemented with phorbol myristate acetate to induce differentiation into macrophage-like cells. RESULTS: We found that LPS preparations from P. intermedia and P. nigrescens can induce IL-8 mRNA expression and stimulate the release of IL-8 in differentiated THP-1 cells without additional stimuli. CONCLUSIONS: There are no previous reports of the ability of P. intermedia and P. nigrescens LPS to stimulate the release of IL-8, and the present study clearly shows, for the first time, that LPSs from P. intermedia and P. nigrescens fully induced IL-8 mRNA expression and IL-8 production in differentiated human monocytic cell line THP-1. The ability of P. intermedia and P. nigrescens LPS to promote the production of IL-8 may be important in the pathogenesis of inflammatory periodontal disease.
Cell Line ; Humans ; Interleukin-8 ; Lipopolysaccharides ; Macrophages ; Periodontal Diseases ; Phorbols ; Prevotella ; Prevotella intermedia ; Prevotella nigrescens ; RNA, Messenger ; Tetradecanoylphorbol Acetate

No abstract available.
Prevotella intermedia* ; Prevotella*

The purpose of this study was to assess some biological activities of lipopolysaccharides (LPSs) from P. intermedia and P. nigrescens. LPS was prepared by the standard hot phenol-water method. NO production was assayed by measuring the accumulation of nitrite in culture supernatants. TNF-alpha production was determined by enzyme-linked immunosorbent assay. Western blot analysis of iNOS and analysis of reverse transcription (RT)-PCR products were carried out. LPS from P. intermedia demonstrated higher KDO content than those from two stains of P. nigrescens. LPSs from P. intermedia and P. nigrescens were mitogenic for spleen cells of BALB/C mouse. The present study clearly shows that LPSs from P. intermedia and P. nigrescens fully induced iNOS expression and NO production in RAW264.7 cells in the absence of other stimuli. Moreover, LPSs from P. intermedia and P. nigrescens clearly induced TNF-alpha production in RAW264.7 cells. The biological activities of LPS from P. intermedia was found to be comparable to those of P. nigrescens LPS. The ability of LPSs from P. intermedia and P. nigrescens to promote the production of NO and TNF-alpha may be important in the pathogenesis of inflammatory periodontal disease.
Animals ; Blotting, Western ; Coloring Agents ; Enzyme-Linked Immunosorbent Assay ; Lipopolysaccharides* ; Mice ; Nitric Oxide ; Periodontal Diseases ; Prevotella intermedia* ; Prevotella nigrescens* ; Prevotella* ; Reverse Transcription ; Spleen ; Tumor Necrosis Factor-alpha

Prevotella intermedia has been implicated as a potent pathogen in many kinds of periodontal, pulpal and periapical diseases. However, it has been isolated from periodontally healthy adults and from edentulous children as well. The intraspecies heterogeneity of Prevotella intermedia has been demonstrated in early studies and finally Shah & Gharbia confirmed the existence of 2 DNA homology groups and proposed dividing Prevotella intermedia into 2 species, Prevotella intermedia and Prevotella nigrescens. This study was designed to examine the frequency of Prevotella intermedia and Prevotella nigrescens in diseased periodontal pockets and healthy gingival sulcus of Korean people by PCR based on 16s ribosomal DNA sequence. One hundred adults who had adult periodontitis but not taken any periodontal treatment or antibiotics during previous 6 months and 50 adults who had healthy periodontal tissue were selected for this study. The sulcular fluid was collected into VMGA by sterilized paper point and diluted to 1,000 times in anaerobic chamber. 100ml of sample was cultured in 37degrees C for 10 days. Among the bacterial colonies, BPB were selected and cultured in BHI broth and then Prevotella intermedia was identified through Gram staining and biochemical test. Identified Prevotella intermedia was cultured again and centrifuged. DNA was extracted from the pellet using several reagents. PCR was performed by previously designed primer. The results were followed. 1.BPB were isolated from 39 of 100 samples of diseased periodontal pockets(39%). 2.Prevotella intermedia was identified from 24 of 39 BPB samples. 3.Among 24 Prevotella intermedia, 21 were confirmed as Prevotella intermedia(87.5) and 2 were confirmed as Prevotella nigrescens(8.33%). 4.BPB were isolated from 9 of 50 samples of periodontally healthy patients. Among them only two were identified as Prevotella intermedia, that is, one was confirmed as Prevotella intermedia and the other was Prevotella nigrescens.
Adult* ; Anti-Bacterial Agents ; Child ; Chronic Periodontitis* ; DNA ; DNA, Ribosomal ; Humans ; Indicators and Reagents ; Periapical Diseases ; Periodontal Pocket ; Polymerase Chain Reaction ; Population Characteristics ; Prevotella intermedia* ; Prevotella nigrescens* ; Prevotella*

The DNA probes Pn17 and Pn34 were evaluated for their ability to specifically detect clinical strains of P. intermedia and P. nigrescens from a Korean population by dot blot hybridization. These probes were sequenced by extension termination and their specificity was determined by Southern blot analysis. The results revealed that the Pn17 sequence (2,517 bp) partially encodes an RNA polymerase beta subunit (rpoB) and that Pn34 (1,918 bp) partially encodes both rpoB (1-169 nts) and the RNA polymerase beta subunit (rpoB'; 695-1918 nts). These probes hybridized with both HindIII- and PstI-digested genomic DNAs from the strains of P. intermedia and P. nigrescens used in this study. Interestingly, each of the hybrid bands generated from the HindIII-digested genomic DNAs of the two bacterial species could be used to distinguish between them via restriction fragment length polymorphism. These results thus indicate that Pn17 and Pn34 can simultaneously detect P. intermedia and P. nigrescens.
Blotting, Southern ; Chimera ; DNA ; DNA Probes ; DNA-Directed RNA Polymerases ; Polymorphism, Restriction Fragment Length ; Prevotella ; Prevotella intermedia ; Prevotella nigrescens ; Sensitivity and Specificity

The results of this study confirm that the availability of hemin influences the expression of selected membrane proteins of Porphyromonas gingivalis, Prevotella intermedia, and Prevotella nigrescens. A 30 kDa (heated 24 kDa) hemin-binding protein whose expression is hemin regulated was identified and purified in P. gingivalis. A strong hemin-binding function was found by LDS-PAGE and TMBZ staining when P. gingivalis cells were grown under hemin-limited conditions. A 50 kDa cell envelope associated protein, whose expression is hemin regulated, is considered to be a putative hemin binding protein from P. intermedia and P. nigrescens, respectively. N-terminal amino acid sequence analysis of CNBr-digested 24 kDa hemin binding protein from P. gingivalis revealed that this protein belongs to a new, so far undescribed hemin-binding class of proteins. N-terminal amino acid sequence of a 50 kDa putative hemin binding protein from P. intermedia was identical with Enolase from Streptococcus intermedia. Work is in progress to further characterize the molecular structure of these proteins.
Amino Acid Sequence ; Carrier Proteins ; Hemin ; Membrane Proteins ; Molecular Structure ; Phosphopyruvate Hydratase ; Porphyromonas gingivalis* ; Porphyromonas* ; Prevotella intermedia* ; Prevotella nigrescens* ; Prevotella* ; Sequence Analysis, Protein ; Streptococcus

It has been established that berberine has strong antimicrobial effects. Little is known however regarding the antimicrobial activity of berberine against endodontic pathogenic bacteria or its cytotoxicity in human oral tissue cells. The antibacterial properties of berberine were tested against 5 strains of Enterococcus faecalis and type strains of Aggregatibacter actinomycetemcomitans, Prevotella nigrescens, Prevotella intermedia, and Tannerella forsythia, which are involved in endodontic infections. Antimicrobial activity was evaluated through minimum inhibitory concentration (MIC) and minimum bactericidal concentration (MBC) measurements. The viability of normal human gingival fibroblast (NHGF) cells after exposure to berberine was measured using a methyl thiazolyl tetrazolium (MTT) assay. The data showed that berberine has antimicrobial effects against A. actinomycetemcomitans with an MIC and MBC of 12.5 microg/ml and 25 microg/ml, respectively. In the cytotoxicity studies, cell viability was maintained at 66.1% following exposure to 31.3 microg/ml berberine. Overall, these findings suggest that berberine has antimicrobial activity against the tested bacteria. Nevertheless, lower concentrations in combination with other reagents will need to be tested before these in vitro results can be translated to clinical use.
Aggregatibacter ; Bacteria* ; Berberine* ; Cell Survival ; Enterococcus faecalis ; Fibroblasts ; Forsythia ; Humans ; Indicators and Reagents ; Microbial Sensitivity Tests ; Prevotella intermedia ; Prevotella nigrescens

No abstract available.
Prevotella*

The purpose of this study was to investigate the frequency of 7 putative pathogens in endodontic infections. The specimens were collected from infected pulpal tissue of patients who were referred for root canal treatment to the department of conservative dentistry, Chosun University. Samples were collected aseptically using a barbed broach and a paper point. The cut barbed broaches and paper points were transferred to an eppendorf tube containing 500 ml of 1 X PBS. DNAs were extracted from the samples by direct DNA extraction method using lysis buffer (0.5% EDTA, 1% Triton X-100). Identification of 7 putative pathogens was performed by PCR based on 16S rDNA. The target species were as follows: Porphyromonas endodontalis, Porphyromonas gingivalis, Prevotella intermedia, Prevotella nigrescens, Bacteroides forsythus, Actinobacillus actinomycetemcomitans, and Treponema denticola. Our data revealed that the prevalence of P. endodontalis was found in 88.6% (39/54), P. gingivalis 52.3% (23/44), P. nigrescens 18.2% (8/44), P. intermedia 15.9% (7/44), B. forsythus 18.2% (8/44), A. actinomycetemcomitans 2.3% (1/44), T. denticola 25% (11/44) of the samples. The high prevalence of P. endodontalis and P. gingivalis suggests that they may play an important role in the etiology of endodontic infections.
Aggregatibacter actinomycetemcomitans ; Bacteroides ; Dental Pulp Cavity ; Dentistry ; DNA ; DNA, Ribosomal* ; Edetic Acid ; Humans ; Neptune ; Polymerase Chain Reaction* ; Porphyromonas endodontalis ; Porphyromonas gingivalis ; Prevalence ; Prevotella intermedia ; Prevotella nigrescens ; Treponema denticola