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1.
A M Lo Bue G Nicoletti M A Toscano B Rossetti G Calì F Condorelli 《The new microbiologica》1999,22(3):209-218
Forty-six adult periodontal patients, selected on the basis of clinical examination, and 46 adult healthy subjects were examined. The subgingival plaque samples from one inflammatory and one non-inflammatory site of each periodontal patient were studied to determine Porphyromonas gingivalis prevalence related to other periodontal micro-organisms and to periodontal tissue destruction. The results showed Porphyromonas gingivalis as the main pathogenic micro-organism isolated in the inflammatory sites together with Bacteroides forsythus. Peptostreptococcus sp., Actinomyces sp. and Prevotella sp. were found as a normal oral flora in the healthy subjects. Fusobacterium nucleatum, Prevotella intermedia, Campylobacter rectus and Eikenella corrodens were detected both in inflammatory and in non-inflammatory sites of periodontal patients as well as in the healthy subjects. 相似文献
2.
Detection and comparison of specific hemin binding by Porphyromonas gingivalis and Prevotella intermedia.
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A radioligand assay was designed to detect and compare specific hemin binding by the periodontal anaerobic black-pigmenting bacteria (BPB) Porphyromonas gingivalis and Prevotella intermedia. The assay included physiological concentrations of the hemin-binding protein rabbit serum albumin (RSA) to prevent self-aggregation and nonspecific interaction of hemin with cellular components. Under these conditions, heme-starved P. intermedia cells (two strains) expressed a single binding site species (4,100 to 4,600 sites/cell) with a dissociation constant (Kd) of 1.0 x 10(-9) M. Heme-starved P. gingivalis cells (two strains) expressed two binding site species; the higher-affinity site (1,000 to 1,500 sites/cell) displayed a Kd of between 3.6 x 10(-11) and 9.6 x 10(-11) M, whereas the estimated Kd of the lower-affinity site (1.9 x 10(5) to 6.3 x 10(5) sites/cell) ranged between 2.6 x 10(-7) and 6.5 x 10(-8) M. Specific binding was greatly diminished in heme-replete cells of either BPB species and was not displayed by iron-replete Escherichia coli cells, which bound as much hemin in the absence of RSA as did P. intermedia. Hemin binding by BPB was reduced following treatment with protein-modifying agents (heat, pronase, and N-bromosuccinimide) and was blocked by protoporphyrin IX and hemoglobin but not by Congo red. Hemopexin also inhibited bacterial hemin binding. These findings indicate that both P. gingivalis and P. intermedia express heme-repressible proteinaceous hemin-binding sites with affinities intermediate between those of serum albumin and hemopexin. P. gingivalis exhibited a 10-fold-greater specific binding affinity and greater heme storage capacity than did P. intermedia, suggesting that the former would be ecologically advantaged with respect to heme acquisition. 相似文献
3.
During isolation of Porphyromonas gingivalis from periodontal pockets of patients, the appearance of an unusual rough colony form, designated NUM 114, was observed. The NUM 114 strain grew in aggregated cell form in a liquid culture and formed a light-beige rough colony on blood agar medium. The identifications and DNA studies confirmed that the NUM 114 strain was P. gingivalis. The enzymatic activities and fatty acid end products were in lower levels than found in P. gingivalis 381, a representative strain. The NUM 114 strain had enhanced hydrophobicity, hemagglutination of human erythrocytes and adherence to human buccal epithelial cells. The NUM 114 cells were phagocytized at a two-fold higher rate compared with the 381 strain. NUM 114 cells were also more susceptible to killing by phagocytosis than the 381 cells. The carbohydrates of the outer membrane and crude lipopolysaccharide preparation from the NUM 114 strain were in larger amounts than those of 381 strain. LPS from NUM 114 were observed to be smooth-type. 相似文献
4.
Rabbit antibodies to cell wall mannans of various microbial strains and their mutants were found to be cross-reactive to cell carbohydrates of mammalian sperm and 4–6-day-old blastocysts. Immunochemical studies indicate that oligomers of α1→2, α1→3, α1→6, and probably also α→4 linked mannose residues of sperm carbohydrates are available for antibody binding. At least 80% of binding activity of a yeast mannan antibody to sperm can be effectively inhibited by specific haptens or digestion with exo-α-D-mannosidase, an enzyme activity highest in testicular tissue. In order to determine the role of this enzyme in the metabolism of the cross-reactive mannan antigens of sperm, the relative amount of a specific α-linked oligomannosyl determinant of bovine sperm from homozygous normals was compared to that of heterozygous carriers of α-mannosidase deficiency. Extensive cross-reactivity between the microbial and mammalian oligomannosyl determinants suggest that these are conserved structures in cell carbohydrates, although the organization of these units in the microbial cell wall lipopolysaccharide has very little similarity to the carbohydrate moieties of mammalian glycoproteins. 相似文献
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6.
Julie Fletcher Sean Nair Steven Poole Brian Henderson Michael Wilson 《Current microbiology》1998,36(4):216-219
The aim of this study was to determine whether biofilms of Porphyromonas gingivalis could proteolytically degrade the cytokines interleukin (IL)-1β, IL-6, or IL-1 receptor antagonist (IL-1ra). Biofilms were
grown on membrane filters on the surface of Wilkins-Chalgren blood agar. The biofilms were removed from the plates, and solutions
containing 2.5 μg/ml of each cytokine were added. Following incubation for up to 4.0 h, supernatants from the biofilms were
subjected to SDS-PAGE. The separated proteins were transferred by Western blotting to PVDF membranes and probed with peroxidase-conjugated
antibodies recognizing both the intact cytokines and their degradation products. After 2 h, no intact IL-1β, IL-6, or IL-1ra
were detectable. Cytokine proteolysis also occurred in the presence of horse serum. These results demonstrate that biofilm-grown
P. gingivalis can degrade both pro- and anti-inflammatory cytokines and so may be able to perturb cytokine networks in vivo by eliminating
cytokines from the local environment.
Received: 12 August 1997 / Accepted: 15 October 1997 相似文献
7.
Bullon P Cordero MD Quiles JL Morillo JM del Carmen Ramirez-Tortosa M Battino M 《Free radical biology & medicine》2011,50(10):1336-1343
Oxidative stress is one of the factors that could explain the pathophysiological mechanism of inflammatory conditions that occur in cardiovascular disease (CVD) and periodontitis. Such inflammatory response is often evoked by specific bacteria, as the lipopolysaccharide (LPS) of Porphyromonas gingivalis is a key factor in this process. The aim of this research was to study the role of mitochondrial dysfunction in peripheral blood mononuclear cells (PBMCs) from periodontitis patients and to evaluate the influence of LPS on fibroblasts to better understand the pathophysiology of periodontitis and its relationship with CVD. PBMCs from patients showed lower CoQ10 levels and citrate synthase activity, together with high levels of ROS production. LPS-treated fibroblasts provoked increased oxidative stress and mitochondrial dysfunction by a decrease in mitochondrial protein expression, mitochondrial mass, and mitochondrial membrane potential. Our study supports the hypothesis that LPS-mediated mitochondrial dysfunction could be at the origin of oxidative stress in periodontal patients. Abnormal PBMC performance may promote oxidative stress and alter cytokine homeostasis. In conclusion, mitochondrial dysfunction could represent a possible link to understanding the interrelationships between two prominent inflammatory diseases: periodontitis and CVD. 相似文献
8.
Porphyromonas gingivalis is considered an important pathogen in periodontal disease. While this organism expresses a number of virulence factors, no study combining different virulence polymorphisms has, so far, been conducted. The occurrence of combined virulence (Cv) genotypes in 62 isolates of P. gingivalis was investigated from subjects displaying either chronic periodontitis or periodontal abscess. The Cv genotypes, based on gene variation of fimbriae (fimA), Lys-specific cystein proteinase (kgp) and Arg-specific cystein proteinase (prpR1/rgpA), were evaluated by PCR. The isolates were also subjected to capsular polysaccharide K-serotyping. A total of 18 Cv genotype variants based on fimA: kgp: rgpA were identified, of which II:I:A and II:II:A Cv genotypes (53.3%) were the two most frequently detected combinations. Moreover, 36% of the isolates were K-typeable, with the K6 serotype being the most prevalent (23%). Two isolates had the same genotype as the virulent strain W83. The results indicate that chronic periodontitis is not associated with a particularly virulent clonal type. A highly virulent genotype (e.g. strain W83) of P. gingivalis can be found in certain periodontitis patients. 相似文献
9.
Multilocus sequence typing and fimA genotyping were performed on Porphyromonas gingivalis isolates from 15 subjects with "refractory" periodontitis. Several sequence types were detected for most individual pockets. The variation indicated recombination at the recA and pepO genes. The prevalence of fimA genotypes II and IV confirmed their association with periodontitis. 相似文献
10.
Immunological characterization of embryonic cell surface antigens recognized by antiblastocyst serum
Stage-specific cell surface antigens expressed during mouse preimplantation development and detected by a rabbit antiserum prepared against mouse blastocysts (A-BL2) have been characterized by serological and biochemical techniques. Immunofluorescence, immunoradiolabeling, and complement-mediated cytotoxicity assays reveal the expression of A-BL2 surface antigens beginning at the 4-cell stage and reaching a maximum at the 8-cell to morula stages. At earlier times in development A-BL2 antigens are not detectable, and there is a decline in expression at the blastocyst stage. No antibody reactivity is detected against adult mouse tissues or teratocarcinoma cell lines. The presence of A-BL2 antibodies during in vitro embryo culture interferes with normal development. Treatment of embryos with β-N-acetylglucosaminidase, but not other glycosidases, proteases, or lipases, results in a quantitative decrease in the binding of A-BL2 antibodies to surface antigens. Immunoprecipitation and electrophoretic analyses of A-BL2 antigens demonstrate specific antibody activity against a pair of embryonic glycoproteins of 65,000 to 70,000 daltons which can be metabolically labeled with 35S-methionine and 3H-glucosamine. Tunicamycin treatment alters the form of the A-BL2 immunoprecipitate to a single 60,000-dalton protein. 相似文献
11.
Koelle DM 《Methods (San Diego, Calif.)》2003,29(3):213-226
Knowledge of the antigens that are recognized by virus-specific T cells can identify candidate subunit vaccine compounds. Viral antigens are frequently used in cross-sectional and longitudinal studies of the immune response in pathogenesis research. Peptide epitopes are required for the construction of fluorescent major histocompatibility complex-peptide tetramers, which are used to track specific T-cell responses. Expression cloning is a family of methods of antigen discovery that are particularly suitable for DNA viruses with large genomes. Libraries of viral nucleic acid are expressed in formats suitable for presentation to either CD4 or CD8 T cells. Pools of antigens representing fragments of viral open reading frames are loaded into cells expressing the necessary antigen processing and presentation machinery. Highly sensitive T-cell readouts such as lymphokine secretion, proliferation, and gene activation are used to detect active pools, which are then broken down in a reiterative process until a single active clone can be isolated and sequenced. These methods are most applicable if T-cell clones reactive with the whole virus can be obtained by in vitro restimulation or sampling of infected tissues. Alternative methods of antigen discovery are better suited for nonculturable viruses. Expression cloning methods are somewhat generic and are adaptable between infectious diseases, autoimmunity, and tumor immunology research projects. 相似文献
12.
Abstract Recent work in our laboratory indicates that selected protease/peptidase inhibitors interfere with the growth of Porphyromonas gingivalis . The aim of the present study was to further investigate the inhibitory effect of bestatin on the growth of P. gingivalis . Complete growth inhibition of P. gingivalis (11 strains) was observed when bestatin was incorporated at 2.5 μg ml−1 in a complex broth medium. Fifty percent inhibition was still obtained with bestatin at a final concentration of 0.5 μg ml−1 . The inhibitory effect of bestatin was highly specific as the growth of 20 different oral bacterial species, including Gram-positive and Gram-negative as well as saccharolytic and asacharolytic bacteria, was not affected even at bestatin concentrations up to 50 μg ml−1 . Bestatin did not significantly affect the viability of P. gingivalis indicating that it has a bacteriostatic rather than a bactericidal effect. Growth assays using other specific inhibitors suggested that the effect of bestatin on the growth of P. gingivalis was unlikely to be related to its aminopeptidase inhibitor activity. Cultivation of P. gingivalis with a subinhibitory concentration of bestatin did not modify the cell envelope protein profile, as determined by SDS-PAGE analysis, but significantly decreased the number of extracellular vesicles produced. The present study indicated that bestasin is a highly effective inhibitor of cell growth of P. gingivalis . Additional studies will indicate whether bestatin should be considered as a potential drug in the control of P. gingivalis , a suspected pathogen in adult chronic periodontitis. 相似文献
13.
Onoue S Imai T Kumada H Umemoto T Kaca W Isshiki Y Kaneko A Kawahara K 《Microbiology and immunology》2003,47(1):51-55
Serum antibody titers against the lipopolysaccharides (LPSs) of Porphyromonas gingivalis and Fusobacterium nucleatum were compared between 9 periodontitis patients and 24 healthy persons. The IgG titers against the LPSs of P. gingivalis ATCC 33277(T) and W50 were clearly higher in the patients than in the healthy persons. However, IgM titers against the LPSs of P. gingivalis strains were relatively low, and no significant difference was observed between the patients and healthy persons. On the other hand, IgG and IgM titers against the LPS of Fusobacterium nucleatum JCM 8532(T) in some patients were significantly higher than those in the healthy persons, although the difference in IgG titers was not large compared to that of the LPS of P. gingivalis. These results suggest that the antibody measurement of patients' sera against the LPS of periodontal bacteria can be applied for the diagnosis of periodontitis. 相似文献
14.
A Kamaguch K Nakayama T Ohyama T Watanabe M Okamoto H Baba 《Microbiology and immunology》2001,45(9):649-656
Porphyromonas gingivalis cells coaggregated with Prevotella intermedia cells. The coaggregation was inhibited with L-arginine, L-lysine, Nalpha-p-tosyl-L-lysine chloromethyl ketone, trypsin inhibitor, and leupeptin. Heat- and proteinase K-treated P. gingivalis cells showed no coaggregation with P. intermedia cells, whereas heat and proteinase K treatments of P. intermedia cells did not affect the coaggregation. The vesicles from P. gingivalis culture supernatant aggregated with P. intermedia cells, and this aggregation was also inhibited by addition of L-arginine or L-lysine and by heat treatment of the vesicles. The rgpA rgpB, rgpA kgp, rgpA rgpB kgp, and rgpA kgp hagA mutants of P. gingivalis did not coaggregate with P. intermedia. On the other hand, the fimA mutant lacking the FimA fimbriae showed coaggregation with P. intermedia as well as the wild type parent. These results strongly imply that a heat-labile and proteinous factor on the cell surface of P gingivalis, most likely the gingipain-adhesin complex, is involved in coaggregation of P. gingivalis and P. intermedia. 相似文献
15.
John W. Smalley Andrew J. Birss Ailsa S. McKee Philip D. Marsh 《Current microbiology》1998,36(2):102-106
Hemoglobin binding to chemostat-grown hemin-excess and hemin-limited cells of Porphyromonas gingivalis W50, and to cells of the avirulent, beige-pigmenting variant W50/BE1, was quantified. Hemin-excess W50 bound more hemoglobin
than hemin-limited W50, mirroring the hemin-binding ability of these cells [Microb Ecol Health Dis 7:9–15, 1994]. In contrast
to hemin, hemoglobin binding was not enhanced by sodium dithionite. The hemoglobin-binding capacity of hemin-excess W50/BE1
was below that of hemin-limited W50 and only observed under oxidizing conditions. Scatchard analysis revealed similar affinity
constants for hemin-excess and hemin-limited W50, and confirmed a lower binding maximum for the latter. Hemin-excess W50/BE1
displayed cooperative binding of hemoglobin. These differences in binding were reflected in the binding of a horse radish
peroxidase-conjugated hemoglobin (HHRPO) in a dot-blot assay. However, neither the 32-kDa hemin-binding protein, nor its 19-kDa
heat-modified form, from either hemin-limited W50 or hemin-excess W50/BE1, bound this conjugate. These data indicate that
hemoglobin binding by P. gingivalis is hemin-regulated and occurs via a mechanism different from hemin binding.
Received: 2 June 1997 / Accepted: 4 August 1997 相似文献
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17.
Genetic analysis of Porphyromonas gingivalis, an obligately anaerobic gram-negative bacterium, has been hindered by the apparent lack of naturally occurring bacteriophages, transposable elements, and plasmids. Plasmid R751::*omega 4 has previously been used as a suicide vector to demonstrate transposition of Tn4351 in B. uniformis. The erythromycin resistance gene on Tn4351 functions in Bacteroides and Porphyromonas. Erythromycin-resistant transconjugants were obtained at a mean frequency of 1.6 x 10(-7) from matings between Escherichia coli HB101 containing R751::*omega 4 and P. gingivalis 33277. Southern blot hybridization analysis indicated that about half of the erythromycin-resistant P. gingivalis transconjugants contained simple insertions of Tn4351 and half contained both Tn4351 and R751 sequences. The presence of R751 sequences in some P. gingivalis transconjugants most likely occurred from Tn4351-mediated cointegration of R751, since we were unable to detect autonomous plasmid in these P. gingivalis transconjugants. The P. gingivalis-Tn4351 DNA junction fragments from different transconjugants varied in size. These results are consistent with transposition of Tn4351 and with insertion at several different locations in the P. gingivalis chromosome. Tn4351 may be useful as a mutagen to isolate well-defined mutants of P. gingivalis. 相似文献
18.
Hojo K Tamura A Mizoguchi C Kato D Ohshima T Maeda N 《Bioscience, biotechnology, and biochemistry》2008,72(5):1348-1351
We isolated oral bacteria that coexisted with Porphyromonas gingivalis in a hamster periodontitis model. As predominant bacteria in the periodontitis site, Collinsella-reltaed strains, Eubacterium-reltaed strains, Streptococcus suis-related strains, and Veillonella parvula-reltaed strains were detected. In addition, Actinomyces, Bacteroides, and P. gingivalis were also isolated predominantly. The results suggest that the bacterial composition of the periodontitis site in hamsters is complex, as in human periodontitis. 相似文献
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20.
Banbula A Mak P Bugno M Silberring J Dubin A Nelson D Travis J Potempa J 《The Journal of biological chemistry》1999,274(14):9246-9252
Porphyromonas gingivalis possesses a complex proteolytic system, which is essential for both its growth and evasion of host defense mechanisms. In this report we characterized, both at a protein and genomic level, a novel peptidase of this system with prolyl tripeptidyl peptidase activity. The enzyme was purified to homogeneity, and its enzymatic activity and biochemical properties were investigated. The amino acid sequence at the amino terminus and of internal peptide fragments enabled identification of the gene encoding this enzyme, which we refer to as PtpA for prolyl tripeptidyl peptidase A. The gene encodes an 82-kDa protein, which contains a GWSYGG motif, characteristic for members of the S9 prolyl oligopeptidase family of serine proteases. However, it does not share any structural similarity to other tripeptidyl peptidases, which belong to the subtilisin family. The production of prolyl tripeptidyl peptidase may contribute to the pathogenesis of periodontal tissue destruction through the mutual interaction of this enzyme, host and bacterial collagenases, and dipeptidyl peptidases in the degradation of collagen during the course of infection. 相似文献