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Journal of Periodontal Research Dec 2001Black-pigmented bacteria which produce cytotoxic metabolic end-products and cell membrane-associated proteases have been reported to play an important role in the...
Black-pigmented bacteria which produce cytotoxic metabolic end-products and cell membrane-associated proteases have been reported to play an important role in the pathogenesis of periodontal diseases. These bacterial virulence factors can be modified by the environmental conditions including nutrients supplied variously into the oral cavity. Although glucose is one of the most essential nutrients for oral bacteria, the exogenous supply of glucose may be discontinuous and the glucose concentration in a periodontal pocket may be influenced by the depth of the periodontal pocket. Therefore, effects of glucose as an environmental factor on the virulence factors of Prevotella intermedia, Prevotella nigrescens and Porphyromonas in the presence o glucose.,bo t P. intermedia and P. nigrescens markedly decreased the production of cytotoxic end-products including succinate.,isobutyrate,isovalerate and ammonia, although their growth was increased. Furthermore, the proteolytic activities such as immunoglobulin- albumin- and casein-degrading activities of these bacteria were decreased in the presence of glucose. On the other hand, no effect of glucose on the metabolic activity of P gingivalis was observed. These results suggest that pathogenicity of P. intermedia P. nigrescens may be decreased by the presence of glucose.
Topics: Ammonia; Bacterial Proteins; Endopeptidases; Fatty Acids, Volatile; Glucose; Hydrolysis; Porphyromonas gingivalis; Prevotella; Virulence
PubMed: 11762870
DOI: 10.1034/j.1600-0765.2001.360602.x -
Research in Microbiology 2004Hemolytic activity was evaluated in the putative periodontopathogens Prevotella intermedia and Prevotella nigrescens. Whole cells of both species present weak hemolytic...
Hemolytic activity was evaluated in the putative periodontopathogens Prevotella intermedia and Prevotella nigrescens. Whole cells of both species present weak hemolytic activity evidenced only by solid media assays after 48 h of bacterial growth or after 5 h of interaction with erythrocytes at 37 degrees C in liquid assays. In this work we show that the use of crude extract allowed the detection of a higher hemolytic activity for P. intermedia, but surprisingly not for P. nigrescens. Incubation at 37 degrees C for 9 h, or treatment with trypsin or proteinase K, increased or exposed the hemolytic activity of P. intermedia and P. nigrescens crude extract, respectively. The activation process was inhibited by TLCK and PMSF but not by EDTA, E-64 or pepstatin A, indicating the serino-protease nature of the factor involved in activation of P. intermedia and P. nigrescens hemolysins. Both the buffer and the pH employed for cell fractionation influenced the activation of hemolysin, and the best results were obtained with Universal buffer at pH 8.0. The activated hemolysins acted optimally at pH 6.5 at 37 degrees C and the maximum hemolytic activity was detected at the early log phase of growth. The results of this study show for the first time a strong hemolytic activity for P. nigrescens and evidence of proteolytic activation of hemolysins produced by periodontopathogens.
Topics: Animals; Edetic Acid; Endopeptidase K; Endopeptidases; Enzyme Activation; Enzyme Inhibitors; Enzyme Stability; Hemolysin Proteins; Hemolysis; Humans; Hydrogen-Ion Concentration; Kinetics; Leucine; Pepstatins; Phenylmethylsulfonyl Fluoride; Prevotella intermedia; Prevotella nigrescens; Rabbits; Temperature; Tosyllysine Chloromethyl Ketone; Trypsin
PubMed: 14759706
DOI: 10.1016/j.resmic.2003.09.010 -
Journal of Investigative and Clinical... Feb 2013To estimate differences in the prevalence of Prevotella intermedia and Prevotella nigrescens in the subgingival plaque of patients with periodontitis (including... (Comparative Study)
Comparative Study
AIM
To estimate differences in the prevalence of Prevotella intermedia and Prevotella nigrescens in the subgingival plaque of patients with periodontitis (including aggressive and advanced chronic periodontitis) compared to healthy controls, and to search for significant associations with clinical status.
METHODS
Sixteen patients and 16 healthy controls were enrolled in this study. Interproximal plaque index, oral hygiene index, gingival index, bleeding on probing, probing depth, and clinical attachment level were recorded. Samples of subgingival plaque were taken with paper points from four teeth of each individual and immediately plated on appropriate supplemented Columbia agar. Black pigmented colonies were identified with the Rapid ID32 A system, and further differentiated using matrix-assisted laser desorption ionization-time of flight mass spectrometry. For the statistical analysis, chi-squared test and the Mann-Whitney U-test were used.
RESULTS
Prevotella nigrescens was isolated from 10 patients and three controls, while P. intermedia was isolated from only two patients. P. nigrescens was found more frequently in the subgingival plaque of patients (P = 0.029), and was significantly associated with high values of clinical indices (P ≤ 0.025). Significant differences for P. intermedia were not found.
CONCLUSIONS
Periodontitis seems to be associated with increased colonization with P. nigrescens. Whether or not it is a major pathogen needs to be determined.
Topics: Adult; Aged; Aggressive Periodontitis; Alveolar Bone Loss; Bacteriological Techniques; Bacteroidaceae Infections; Chronic Periodontitis; Dental Plaque; Dental Plaque Index; Female; Humans; Male; Middle Aged; Oral Hygiene Index; Periodontal Attachment Loss; Periodontal Index; Periodontal Pocket; Prevotella intermedia; Prevotella nigrescens; Spectrometry, Mass, Matrix-Assisted Laser Desorption-Ionization; Young Adult
PubMed: 22767485
DOI: 10.1111/j.2041-1626.2012.00129.x -
International Journal of Systematic... Jul 1995The elevation of the two genotypes of Prevotella intermedia to species rank as P. intermedia and Prevotella nigrescens has increased the need for reliable...
The elevation of the two genotypes of Prevotella intermedia to species rank as P. intermedia and Prevotella nigrescens has increased the need for reliable differentiation between the two taxa. In this study, 53 strains, including strains whose species affiliations were known as well as fresh dental plaque isolates, were subjected to a multilocus enzyme electrophoretic analysis, DNA analyses in which we used whole genomic DNA, rRNA sequences, and an oligonucleotide specific for the former P. intermedia genotype II (P. nigrescens) as probes, and a sodium dodecyl sulfate-polyacrylamide gel electrophoresis analysis of soluble cellular proteins. All of these tests consistently separated the strains into the same two distinct groups corresponding to P. intermedia and P. nigrescens, confirming that the two species constitute two distinct populations of bacteria. Each of the tests used independently provided reliable identification to the species level. A previously reported heterogeneity in the pattern of human immunoglobulin A1 (IgA1) degradation was not confirmed. No differences between species were observed. All of the strains induced total degradation of IgA1 within 48 h, a property that may be a virulence factor in periodontal disease development. The enzymes responsible for IgA1 degradation were not inactivated by the physiological proteinase inhibitors alpha 2-macroglobulin and alpha 1-proteinase inhibitor.
Topics: Alleles; Blotting, Southern; Blotting, Western; DNA, Bacterial; DNA, Ribosomal; Dental Plaque; Electrophoresis, Polyacrylamide Gel; Genetic Variation; Humans; Immunoglobulin A; Periodontal Diseases; Polymorphism, Restriction Fragment Length; Prevotella; Prevotella intermedia; Proteins; Time Factors
PubMed: 8590668
DOI: 10.1099/00207713-45-3-429 -
Letters in Applied Microbiology 2005To characterize a minimal bacteriocin operon of Prevotella nigrescens ATCC 25261.
AIMS
To characterize a minimal bacteriocin operon of Prevotella nigrescens ATCC 25261.
METHODS AND RESULTS
A genomic DNA library of Pr. nigrescens ATCC 25261 was constructed and screened for bacteriocin production by an agar overlay assay. Sequence analysis of the bacteriocin-producing recombinant plasmid, pGP2, has shown that the insert DNA consists of 4868 base pairs, termed nig locus. There is a cluster of four genes within the nig locus, respectively designated nigA, B, C and D. Deleting 160 nucleotides at the 3'-end of nigAB resulted in loss of bacteriocin production, indicating that nigAB may belong to a bacteriocin operon. nigA is thought to be the bacteriocin gene, while nigB may encode an immunity protein. Escherichia coli containing pGP2 expressed the bacteriocin, which is similar in size, antimicrobial activity, and biochemical properties to that purified from Pr. nigrescens ATCC 25261.
CONCLUSION
nig Locus is a chromosomal fragment of Pr. nigrescens ATCC 25261, consisting of 4868 base pairs, and has been proved to be important for bacteriocin production.
SIGNIFICANCE AND IMPACT OF THE STUDY
This is the first report of the successful cloning and expression of the bacteriocin from Pr. nigrescens ATCC 25261 into E. coli. This will facilitate the construction of bacteriocin analogues and permit investigation of their structure/function relationships.
Topics: Bacterial Proteins; Bacteriocins; Cloning, Molecular; Escherichia coli; Genomic Library; Molecular Sequence Data; Operon; Prevotella nigrescens; Sequence Analysis, DNA
PubMed: 15644114
DOI: 10.1111/j.1472-765X.2004.01639.x -
International Journal of Systematic... Oct 1992A total of 31 strains of Prevotella intermedia were subjected to DNA-DNA hybridization and were characterized by performing physiological tests and by performing a...
A total of 31 strains of Prevotella intermedia were subjected to DNA-DNA hybridization and were characterized by performing physiological tests and by performing a multilocus enzyme analysis, using malate dehydrogenase and glutamate dehydrogenase. All of the strains assigned to P. intermedia fermented glucose and sucrose, hydrolyzed starch but not esculin, and produced indole, acetic, isobutyric, isovaleric, and succinic acids as metabolic end products. The results of DNA reassociation experiments performed with the reference probe permitted separation of the strains into two well-defined homology groups. In addition, strains with DNAs that hybridized with DNA from strain ATCC 25611T (T = type strain) had high levels of peptidase activity and cleaved lipid substrates (4-methylumbelliferyl laurate and 4-methylumbellifelyl elaidate). Multilocus enzyme electrophoresis revealed two electromorphic profiles, one characteristic of strain ATCC 25611T and the other characteristic of strain ATCC 33563T. We propose that a new species, Prevotella nigrescens, should be created for the genetically distinct group of strains that hybridized with strain ATCC 33563T. Strain ATCC 33563 is designated the type strain of P. nigrescens.
Topics: Bacterial Typing Techniques; Bacteroides; Cellobiose; Culture Media; DNA, Bacterial; Fermentation; Nucleic Acid Hybridization; Phenotype; Pigmentation; Xylans
PubMed: 1390107
DOI: 10.1099/00207713-42-4-542 -
Letters in Applied Microbiology 2004To characterize the antimicrobial activity produced by Prevotella nigrescens ATCC 25261, and to evaluate its safety on cultured gingival fibroblasts.
AIMS
To characterize the antimicrobial activity produced by Prevotella nigrescens ATCC 25261, and to evaluate its safety on cultured gingival fibroblasts.
METHODS AND RESULTS
An antimicrobial activity was obtained from purifying the culture supernatant of Pr. nigrescens ATCC 25261. Purification of the active compound was achieved with ammonium sulphate precipitation followed by anion-exchange and gel filtration chromatography. As revealed by SDS-PAGE, the active fraction was relatively homogeneous, showing a protein with an approximate molecular weight of 41 kDa. The antimicrobial compound, named nigrescin, exhibited a bactericidal mode of action against Porphyromonas gingivalis, Prevotella intermedia, Tannerella forsythensis, and Actinomyces spp. Nigrescin was stable in a pH range between 6.5 and 9.5, at 100 degrees C for 10 min, and resistant to lyophilization. But its activity was lost after proteinase K treatment. Despite at very high concentrations beyond the minimum inhibitory concentration (MIC), nigrescin was not toxic to the gingival fibroblasts.
CONCLUSION
Nigrescin is a novel bacteriocin produced by Pr. nigrescens ATCC 25261. It exhibits antimicrobial activity against species that are implicated in periodontal diseases. The absence of toxicity on the gingival fibroblasts suggests the possibility in using of nigrescin for an application in periodontal treatment.
SIGNIFICANCE AND IMPACT OF THE STUDY
Novel evidence on nigrescin would make Pr. nigrescens ATCC 25261 attractive in biotechnological applications as an antimicrobial agent in clinical dentistry.
Topics: Actinomyces; Ammonium Sulfate; Anti-Bacterial Agents; Bacteriocins; Bacteroides; Cells, Cultured; Chromatography, Gel; Chromatography, Ion Exchange; Electrophoresis, Polyacrylamide Gel; Endopeptidase K; Enzyme Stability; Fibroblasts; Fractional Precipitation; Freeze Drying; Gingiva; Humans; Hydrogen-Ion Concentration; Molecular Weight; Porphyromonas gingivalis; Prevotella intermedia; Prevotella nigrescens; Temperature
PubMed: 15482437
DOI: 10.1111/j.1472-765X.2004.01608.x -
Oral Microbiology and Immunology Dec 1998The pathogenicity of obligate and facultative anaerobic bacteria commonly found in endodontic infections was tested using a mouse model. The capacity of inducing... (Comparative Study)
Comparative Study
The pathogenicity of obligate and facultative anaerobic bacteria commonly found in endodontic infections was tested using a mouse model. The capacity of inducing abscesses was evaluated seven days after subcutaneous injection of the bacteria in pure culture and in combinations with either Prevotella intermedia or Prevotella nigrescens. Nine of the fifteen bacterial strains tested were pathogenic in pure culture. No statistically significant differences were detected between these strains in pure culture and in mixtures with either P. intermedia or P. nigrescens. Synergism between the bacterial strains was only apparent when associating Porphyromonas endodontalis with P. intermedia or P. nigrescens. Histopathological examination of tissue sections from induced abscesses revealed an acute inflammatory reaction, dominated by polymorphonuclear leukocytes. Sections from the control group using sterile medium showed no evidence of inflammatory reaction.
Topics: Abscess; Animals; Bacteria, Anaerobic; Dental Pulp Diseases; Disease Models, Animal; Ecosystem; Male; Mice; Porphyromonas; Prevotella; Prevotella intermedia; Virulence
PubMed: 9872113
DOI: 10.1111/j.1399-302x.1998.tb00693.x -
Oral Diseases Mar 1995Eight oligonucleotides based upon regions of the small subunit 16S ribosomal RNA gene sequences were analysed against a background of their position within the molecule...
Oligonucleotide probes to the 16S ribosomal RNA: implications of sequence homology and secondary structure with particular reference to the oral species Prevotella intermedia and Prevotella nigrescens.
Eight oligonucleotides based upon regions of the small subunit 16S ribosomal RNA gene sequences were analysed against a background of their position within the molecule and their two-dimensional structure to rationalise their use in recognising Prevotella intermedia and Prevotella nigrescens. The 41 clinical isolates from both oral and respiratory sites and two reference strains were subjected to DNA-DNA hybridisation and multilocus enzyme electrophoresis to confirm their identity. Alignment of oligonucleotide probes designated I Bi-2 to I Bi-6 (for P. intermedia) and 2Bi-2 (for P. nigrescens) with the 16S rRNA suggested that these probes lacked specificity or were constructed from hypervariable regions. A 52-mer oligonucleotide (designated Bi) reliably detected both species. Because of the high degree of concordance between the 16S rRNAs of both species, it was necessary to vary the stringency of hybridisation conditions for detection of both species. Thus probe I Bi-I recognised P. intermedia while I Bi-I detected both P. intermedia and P. nigrescens at low stringency. However, under conditions of high stringency only P. nigrescens was recognised by probe 2Bi-I. These probes were highly specific and did not hybridise with DNA from the closely related P. corporis, nor other periodontal pathogens such as Fusobacterium nucleatum, Actinobacillus actinomycetemcomitans, Treponema denticola and several pigmented species such as Prevotella melaninogenica, P. denticola, P. loescheii, Porphyromonas asaccharolytica, Py. endodontalis, Py. gingivalis, Py. levii, and Py. macacae.
Topics: Bacterial Typing Techniques; Base Sequence; DNA, Bacterial; Humans; Molecular Sequence Data; Nucleic Acid Conformation; Nucleic Acid Hybridization; Oligonucleotide Probes; Prevotella; Prevotella intermedia; RNA, Bacterial; RNA, Ribosomal, 16S; Sequence Alignment; Sequence Analysis, RNA; Sequence Homology, Nucleic Acid
PubMed: 7553378
DOI: 10.1111/j.1601-0825.1995.tb00154.x -
Journal of Clinical Microbiology Apr 1999Primers were designed from 16S rRNA sequences of Prevotella intermedia sensu stricto and Prevotella nigrescens and were used to discriminate these two species by PCR.... (Comparative Study)
Comparative Study
Primers were designed from 16S rRNA sequences of Prevotella intermedia sensu stricto and Prevotella nigrescens and were used to discriminate these two species by PCR. The results were compared with those from the PCR technique using primers designed from arbitrarily primed PCR products by Guillot and Mouton (E. Guillot and C. Mouton, J. Clin. Microbiol. 35:1876-1882, 1997). The specificities of both assays were studied by using P. intermedia ATCC 25611, P. nigrescens ATCC 33563, 174 clinical isolates of P. intermedia sensu lato, and 59 reference strains and 58 clinical isolates of other Prevotella species and/or common oral flora. In addition, the usefulness and reliability of sodium dodecyl sulfate-polyacrylamide gel electrophoresis (SDS-PAGE) in the differentiation of the two species were examined by comparing the results with those from PCR assays. The controversial lipase test for distinguishing these species was also carried out. Unambiguous differentiation was made by both PCR assays, and the results matched each other. The SDS-PAGE assay was found to misidentify a few strains tested, compared with the results of PCR assays. The lipase test was positive for both species, including the reference strains of P. intermedia and P. nigrescens. We conclude that both PCR assays are simple, rapid, reliable, and specific methods which could be used in clinical studies and that the lipase test is not valuable in the differentiation. The reliable discrimination of the two species by SDS-PAGE is questionable.
Topics: Bacterial Typing Techniques; Base Sequence; DNA Primers; Electrophoresis, Polyacrylamide Gel; Evaluation Studies as Topic; Humans; Polymerase Chain Reaction; Prevotella; Prevotella intermedia; RNA, Bacterial; RNA, Ribosomal, 16S; Reproducibility of Results; Sodium Dodecyl Sulfate; Species Specificity
PubMed: 10074526
DOI: 10.1128/JCM.37.4.1057-1061.1999