bluuebunny/crossref_metadata_2025 huggingface dataset:
Involvement of treponemal surface‐located protein and carbohydrate moieties in the attachment of Treponema denticola ATCC 33520 to cultured rat palatal epithelial cells
Authors: Keulers R. A. C. , Maltha J. C. , Mikx F. H. M. , Wolters‐Lutgerhorst J. M. L.
Published: December 2007

We studied the nature of attachment of Treponema denticola ATCC 33520 to a microscopically distinct population of rounded rat palatal epithelial cells. The motility of the freshly harvested spirochetes appeared not to be a prerequisite for attachment. Treatment of T. denticola ATCC 33520 with proteinase‐K, heat, glutaraldehyde, formaldehyde and periodate oxidation decreased the attachment to the rounded rat palatal epithelial cells, indicating the involvement of protein and carbohydrate moieties. Trypsin treatment had no effect on the attachment. The attachment of T. denticola ATCC 33520 was decreased after treatment with native non‐immune rabbit serum, native polyclonal rabbit serum, D‐mannose, N‐acetyl‐D‐galactosamine and sialic acid. The results indicate that the attachment of T. denticola ATCC 33520 to rounded rat palatal epithelial cells is mediated by trypsin‐resistant adhesin(s) of protein and carbohydrate nature, with affinity for D‐mannose, N‐acetyl‐D‐galactosamine and sialic acid.
Modification of experimental Porphyromonas gingivalis murine infection by immunization with a polysaccharide‐protein conjugate
Authors: Schifferle R. E. , Chen P. B. , Davern L. B. , Aguirre A. , Genco R. J. , Levine M. J.
Published: December 2007

To better understand the role of the capsular polysaccharide in the virulence of Porphyromonas gingivalis, the effect of immunization with a polysaccharide‐protein conjugate on experimental murine infection was evaluated. The conjugate was prepared using polysaccharide isolated from P. gingivalis strain ATCC 53977 and bovine scrum albumin. One group of 22 mice was immunized by intraperitoneal injection with the conjugate and a control group of 25 mice was similarly immunized with bovine serum albumin. Serum antibody reactive to the polysaccharide, as determined by enzyme‐linked immunosorbent assay, was elevated in the group of mice immunized with the polysaccharide‐protein conjugate but not in the mice immunized with bovine serum albumin. Both groups of mice were challenged with P. gingivalis strain ATCC 53977 (1010 cells) administered subcutaneously on the dorsal surface. Following challenge, the mice immunized with the polysaccharide‐protein conjugate appeared healthier and demonstrated less weight loss than did the control group of mice. Ulcerative lesions at secondary locations were smaller in mice immunized with the polysaccharide‐protein conjugate. Thus, immunization of mice with a conjugate containing P. gingivalis polysaccharide could reduce the severity of but not prevent an invasive infection with P. gingivalis.
Transmission of oral bacterial species between spouses
Authors: Saarela M. , von Troll‐Lindén B. , Torkko H. , Stucki A.‐M. , Alaluusua S. , Jousimies‐Somer H. , Asikainen S.
Published: December 2007

The transmission of Actinobacillus actinomycetemcomitans, Porphyromonas gingivalis and mutans streptococci was studied between 4 married couples who suffered from advanced periodontitis. Of the 20 couples investigated, the 4 in which both spouses harbored A. actinomycetemcomitans and P. gingivalis were chosen for the transmission study. Three of these couples also harbored mutans streptococci. A. actinomycetemcomitans isolates (8–24 per subject) and mutans streptococcal isolates (5–23 per subject) were serotyped by immunodiffusion technique. For ribotyping, chromosomal DNA from A. actinomycetemcomitans isolates (4–5 per subject) and mutans streptococcal isolates (4–11 per subject) was digested with restriction endonucleases Clo I or Bgl I and Hind III or Sma I, respectively. P. gingivalis isolates (2–15 per subject) were ribotyped by using Cla I, Bgl I and Sma I. The blotted restriction fragments were hybridized to the plasmid pKK3535, which contains the rRNA operon of the E. coli chromosome. The spouses in 2 couples shared the same sero‐ and ribotypes of A. actinomycetemcomitans and S. mutans. P. gingivalis ribotypes were identical in 2 couples. The result suggests transmission of oral bacteria between spouses.
Significance of serum antibody against surface antigens of Actinobacillus actinomycetemcomitans in patients with adult periodontitis
Authors: Saito A. , Hosaka Y. , Nakagawa T. , Seida K. , Yamada S. , Takazoe I. , Okuda K.
Published: December 2007

This study was undertaken to examine the prevalence of Actinobacillus actinomycetemcomitans, its serotype distribution and the serum immune responses against its surface antigens in 41 Japanese patients with adult periodontitis. The dominant A. actinomycetemcomitans serotype isolated was serotype c. Immunoblot analysis of 3 serotypes of A. actinomycetemcomitans‐sonicated antigens and the patient sera revealed that the reactivities with serotype c were the most frequent and that heat‐stable surface serotype‐specific antigen appeared to be immunodominanl. Elevated serum immunoglobulin G titers to extracted lipopolysaccharide and fimbriae antigen of A. actinomycetemcomitans were noted for the patient sera by enzyme‐linked immunosorbent assay. High serum immunoglobulin G tilers to the fimbriae antigen detected in patients without cultivable A. actinomycetemcomitans suggested Ihe possibilily that the clicked antibody to the antigen played a role in eliminating A. actinomycetemcomitans from the periodontal lesions.
Fimbria‐specific immune response in various inbred mice inoculated with Porphyromonas gingivalis 381
Authors: Isogai E. , Isogai H. , Takagi S. , Ishii N. , Fujii N. , Kimura K. , Hayashi M. , Yoshimura F.
Published: December 2007

We studied the genetic control of Porphyromonas gingivalis fimbriae response. Inbred mice with different H‐2 haplotypes and/or different genetic backgrounds were inoculated with viable P. gingivalis 381 cells and tested for fimbria‐specific T cell responses in vivo (delayed‐type hypersensitivity). H‐2d mice showed a strong footpad response, whereas H‐2b mice showed a weak response to fimbriae from P. gingivalis. Similar evidence of genetic control was obtained with an enzyme‐linked immunosorbent assay for the detection of IgG antibody in inbred mice (BALB/c, C3H/HeN and C57BL/6). Several immunoglobulin G (IgG) subclass responses were associated with H‐2 in these B10 congenic mice. However, quantification of IgG antibody to fimbriae was not controlled by H‐2 in B10 congenic mice. The results indicate that, in mice, the responsiveness to fimbriae of P. gingivalis can be controlled by several genes, including the H‐2 complex. C3H/HeN mice were inoculated with the fimbriae intravenously, and the expression of surface antigens on spleen T cells was measured in a fluorescent antibody cell sorter. Stimulation by fimbriae resulted in a changed expression of surface antigens on T cells. Thus, the fimbriae can induce T cell activation.
The antimicrobial activity of essential oils and essential oil components towards oral bacteria
Authors: Shapiro S. , Meier A. , Guggenheim B.
Published: December 2007

A method for reproducibly determining minimal inhibitory concentrations and minimal bactericidal concentrations of plant extracts towards fastidiously and facultatively anaerobic oral bacteria, predicated upon measurements of optical densities in microtitre plate wells, was devised. The antimicrobial properties of some botanical oils were surveyed; of these, Australian lea tree oil, peppermint oil, and sage oil proved to be the most potent essential oils, whereas thymol and eugenol were potent essential oil components.
The in vitro proteolytic and saccharolytic activity of Candida species cultured in human saliva
Authors: Samaranayake Y. H. , MacFarlane T. W. , Samaranayake L. P. , Aitchison T.
Published: December 2007

The proteolytic and saccharolytic activity of 4 Candida species was investigated in batch cultures of pooled, human mixed saliva supplemented with glucose. All the Candida species investigated (Candida albicans, Candida tropicalis, Candida glabrata and Candida krusei) demonstrated a marked growth in saliva with a concomitant reduction in pH from about 7.5 to 3.3, within 72 h. Isotachophoretic analysis of the culture supernatant revealed the presence of a variety of acid anions of which pyruvate and acetate were the most abundant. Proteolysis of salivary components, evaluated by a biochemical assay and sodium dodecyl sulfate‐polyacrylamide gel electrophoresis, was exhibited by all 4 Candida species, although there was inter‐species variation. Despite the similarity in growth rates, C. tropicalis and C. krusei demonstrated greater proteolytic activity than C. albicans and C. glabrata. Neither candidal growth nor proteolysis was observed in glucose‐free control saliva samples. In contrast, the degree of saccharolytic and proteolytic activity of a single isolate of C. albicans in glucose‐supplemented parotid saliva appeared to be relatively weak compared with mixed saliva. As the oral cavity provides ideal low pH niches periodically supplemented with dietary carbohydrates, the acidic proteinases of Candida species may play a role in the pathogenesis of oral candidiasis.
Neutrophil lysosomal nonoxidative microbicidal proteins in early‐onset periodontitis
Authors: Flemmig T. F. , Miyasaki K. T.
Published: December 2007

The purpose of this study was to evaluate whether the reduced microbicidal activity of polymorphonuclear leukocytes (neutrophils) in patients with early‐onset periodontitis is associated with a deficiency of nonoxidative microbicidal proteins. Neutrophils from 10 patients with early‐onset periodontitis and 8 healthy control subjects were assessed for elastase isozymes 1 through 4, cathepsin G isozymes 1 through 4 and defensins (HNP‐1, HNP‐2 and HNP‐3) using cationic and acid urea polyacrylamide gel electrophoresis. The results showed that both the total content and the relative distribution of elastase and cathepsin G isozymes was normal in neutrophils of patients with early‐onset periodontitis. However, the HNP‐3 content was significantly reduced in neutrophils from patients with generalized early‐onset periodontitis. These findings indicate that the impaired microbicidal activities of neutrophils in patients with early‐onset periodontitis does not appear to be based on an elastase or cathepsin G abnormality in neutrophils. Due to the high variability of HNP‐1+2 and HNP‐3 in neutrophils of control subjects, the reduced HNP‐3 content in neutrophils probably plays a minor role in the pathogenesis of generalized early‐onset periodontitis.
Effect of chlorhexidine and toothbrushing on the presence of bacteria on gingival and buccal epithelial cells
Authors: Vaahtoniemi L. H. , Räisänen S. , Stenfors L.‐E.
Published: December 2007

Swabs of buccal and gingival epithelial cells from healthy young adult donors were washed in physiological saline solution, smeared on glass slides and stained with acridine orange. The presence of bacteria attached onto epithelial cells was examined under a fluorescence microscope. Four hours after a chlorhexidine rinse, the number of cells with > 50 attached bacteria had almost completely vanished. The degree of bacterial colonization seemed to re‐establish at a level exceeding the baseline. One week after chlorhexidine treatment the degree of colonization was still over the control level. Toothbrushing with a conventional toothpaste reduced immediately the number of cells with > 50 bacteria. The colonization had re‐established to the pre‐washing levels at the buccal sites at 1 h and at the gingival sites at 4 h after toothbrushing. A method for the evaluating of the antimicrobial power of oral hygiene products is presented.
Carbohydrate depletion of immunoglobulin A1 by oral species of gram‐positive rods
Authors: Frandsen E. V. G.
Published: December 2007

Bacterial deglycosylation of immunoglobulin Al (IgAl), the dominant isotype of antibody in the oral cavity, probably provides both nutrition as well as protection to the oral bacterial community. Representative strains of oral gram‐positive rods were tested for their ability to remove carbohydrates from IgAl. Detection of sialic acids was performed by means of high‐performance liquid chromatography (HPLC) separation (Aminex HPX‐87H) and ultraviolet light absorption at 190 nm, and neutral carbohydrates were measured by HPLC separation (Capcell Pak C‐18 SG 120) and ultraviolet light absorption at 245 nm after derivatization. Four strains of Actinomyces naeslundii, two strains of Corynebacterium matruchotii and one of two strains of Actinomyces odontolyticus partially or totally removed sialic acid, while two strains of Propionibacterium propionicus and the other strain of A. odontolyticus did not. Complete correlation was observed between sialic acid removal, neuraminidase activity measured with fluorogenic substrate and with one exception, altered immunoelectrophoretic mobility of IgAl. Only limited removal of other carbohydrates was observed with poor correlation to exoglycosidase activities measured with chromogenic substrates. Desialylation increases the susceptibility of glycoproteins, including IgAl, to proteolysis. Therefore, the desialylation of IgAl by oral gram‐positive rods may facilitate the proteolytic activities of other oral bacteria, and the concerted action may positively influence the survival of the bacteria in the oral community.
Initial catabolism of sorbitol in Actinomyces naeslundii and Actinomyces viscosus
Authors: Kalfas S. , Takahashi N. , Yamada T.
Published: December 2007

The initial steps of sorbitol catabolism were studied in 4 strains of Actinomyces naeslundii and Actinomyces viscosus that had been isolated from human dental plaque. Cell‐free extracts were prepared from cells grown in the presence of either sorbitol, xylitol or glucose. The extracts from all strains grown on sorbitol had nicotinamide adenine dinucleotide‐linked dehydrogenase activities for sorbitol and xylitol and reduced nicotinamide adenine dinucleotide‐linked reductase activities for fructose and xylulose. Two of the strains also exhibited these activities when grown in the presence of xylitol, and all glucose‐grown cells lacked them. The results indicate that sorbitol metabolism in oral actinomyces involve oxidation of sorbitol to fructose by an inducible enzyme, nicotinamide adenine dinucleotide‐linked sorbitol dehydrogenase. This step is followed by the phosphorylation of fructose with guanosine triphosphate as a main phosphoryl donor. Thus, the initial catabolic pathway of sorbitol in A. naeslundii and A. viscosus is different from those described for other oral bacteria.
Intrafamilial similarity in immunoblot profiles of salivary immunoglobulin A antibody activity to oral streptococci
Authors: Widerström L. , Hamberg K. , Bratthall D.
Published: December 2007

Analysis of the salivary immunoglobulin A (IgA) antibody activity to antigens separated from oral streptococci was carried out on 49 individuals in 11 families. The aim of this study were to i) study the human salivary IgA activity within families to antigens separated from reference strains ofStreptococcus mutans, Streptococcus sobrinusandStreptococcus parasanguisand ii) to search for potential differences in the salivary IgA activity to the streptococcal antigens several years apart. The immunoblot revealed similarity in saliva IgA response within the families. A majority of the bands, ∼7 (median) for each extract was found in both child and parent. A few bands (∼3) were found in parents but not in children. Only one to two bands were found in children's saliva without a corresponding band in any of the parents. The antibody activity of saliva samples obtained several years apart was essentially unchanged, especially considering the total number of bands. Differences in the relative intensity of the bands could be seen, and a few bands appeared only in some immunoblots for a specific subject. For 3 of 4 children participating in the longitudinal study, more bands against mutans streptococci appeared with age.
Lactoferrin Interaction with Actinobacillus actinomycetemcomitans
Authors: Afugupalli K. R. , Kalfas S. , Edwardsson S. , Naidu A. S.
Published: December 2007

The interaction of lactoferrin with Actinobacillus actinomycetemcomitans was examined in a 125I‐labeled protein binding assay. The binding of human and bovine lactoferrins reached maximum within 1 h. Lactoferrin binding to the bacterium was pH‐dependent and reversible. Scatchard analysis indicated the existence of two different types of binding sites on the bacterium, one with a high affinity constant kα=8.8×10−7 M) and the other with a low one (kα=1.8×10−6 M). Bacteria in the exponential phase of growth showed higher binding than cells in the stationary phase. Bacteria grown in medium containing serum and/or lysed erythrocytes bound lactoferrin to a lesser extent. Heat‐inactivated serum, lysed erythrocytes and other proteins such as mucin and laminin inhibited lactoferrin binding to A. actinomycetemcomitans in a competitive binding assay. Sodium dodecyl sulfate polyacrylamidegel electrophoresis and Western blot analysis of the cell envelope as well as the outer membrane of A. actinomycetemcomitans revealed lactoferrin‐reactive protein bands at 29 kDa and 16.5 kDa. The 29‐kDa band displayed a heat‐modifiable lactoferrin‐reactive form with a molecular weight of 34 kDa. Neither proteinase K‐treated cell envelope nor lipopolysaccharide of this bacterium showed reactivity with lactoferrin. These data suggests a specific interaction of lactoferrin with outer membrane proteins of A. actinomycetemcomitans.
Some observations on the nutritional requirements of Eikenella corrodens ATCC 23834T grown in continuous culture
Authors: Gully N. J. , Rogers A. H.
Published: December 2007

The type strain, ATCC 23834, of Eikenella corrodens was grown anaerobically in continuous culture in chemically defined media. Initial experiments showed that glucose was not utilized and it was subsequently omitted from all media. The initial chemically defined medium contained varying levels of 14 amino acids and 20 mM potassium nitrate, an essential nutrient for growth under these conditions. In this medium, the doubling time was 2.1 h, the optimum growth temperature 34°C and the pH 7.2. The key growth‐promoting amino acids were glutamate and serine, and the culture appeared to be nitrate limited. By growing the organism in a series of chemically defined media containing 40 mM nitrate and varying levels of these amino acids, glutamate was found to be the major contributor to biomass increase. Its catabolism may be linked to a respiratory chain involving nitrate as the ultimate electron acceptor.
Patterns of antibody response in subjects with periodontitis
Authors: Haffajee A. D. , Socransky S. S. , Taubman M. A. , Sioson J. , Smith D. J.
Published: December 2007

Periodontal diseases comprise a heterogeneous group of infections that are difficult to distinguish on a clinical basis alone. The purpose of the present investigation was to group periodontitis subjects according to their elevated serum antibody levels to specific subgingival species. A total of 119 subjects (19‐70 years) with evidence of prior periodontal destruction were monitored at 2‐month intervals (maximum 8 visits), prior to therapy, using clinical parameters measured at 6 sites per tooth. The probing attachment level was measured twice at each visit, and an increase of >2.5 mm at a site was used to define subjects with progressing disease. Serum samples were obtained from each subject at each visit and the level of antibody determined by enzyme‐linked immunosorbent assay to 12 subgingival species. Subgingival plaque samples were taken from the mesial aspect of all teeth in each subject at each visit, and the levels of 14 different subgingival species were determined using a colony‐lift method and DNA probes. Subjects were grouped by cluster analysis of their elevated antibody levels using a simple matching coefficient. Ninety‐two subjects fell into 9 clusters with 100% similarity; 29 subjects in one cluster group exhibited elevated antibody to none of the test species. Seven subjects in a second cluster group showed elevated antibody to Bacteroides forsythus. Subjects in the other 7 clusters showed elevated antibody to Actinobacillus actinomycetemcomitans serotype a only or in combination with B. forsythus, A. actinomycetemcomitans serotype b, Prevotella intermedia or Porphyromonas gingivalis. Subjects in different cluster groups differed with respect to age, number of active sites, levels of gingivitis, prior periodontal destruction and levels of 11 of 14 microbial species tested. Subjects with multiple elevated antibody levels had significantly more active sites than subjects with 0 or 1 elevated response. The 23 early‐onset periodontal disease subjects were confined primarily to 2 clusters or were unclustered and 78% had multiple elevated antibody levels. There appeared to be a finite number of frequently detected antibody patterns that may be useful in categorizing periodontitis.
Sucrose‐dependent accumulation of oral streptococci and their adhesion‐defective mutants on saliva‐coated hydroxyapatite
Authors: Vickerman M. M. , Jones G. W.
Published: December 2007

The adhesion and accumulation of oral streptococci on saliva‐coated hydroxyapatite was examined in strains representing species that appear in initial plaque (Streptococcus sanguise FC1 and Streptococcus oralis C5) and in more mature plaque (Streptococcus gordonii G9B). Washed cells of strains FC1 and C5 did not attach better to saliva‐coated hydroxyapatite than did strain G9B, suggesting that the degree of initial adhesiveness does not alone account for the temporal appearance of these bacteria in dental plaque. Growing cells of each strain were also examined for their ability to accumulate on saliva‐coated hydroxyapatite. The addition of sucrose to the medium promoted the accumulation of strain G9B more than it promoted the accumulation of strains FC1 and C5. Sucrose also enhanced the accumulation of adhesion‐defective mutants of each strain to levels similar to those of the respective parent strains. These results suggest that sucrose‐dependent accumulation may facilitate the colonization of the tooth surface by these species of oral streptococci when adhesion is limited by reduced bacterial adhesiveness or limited pellicle‐binding sites.
Primer extension analysis of Streptococcus mutans promoter structures
Authors: Smorawinska M. , Kuramitsu H. K.
Published: December 2007

To characterize the promoter structures of Streptococcus mutans genes, primer extension analysis was carried out with 4 genes previously isolated in this laboratory. An analysis of the transciption start sites for the gtfB, gtfC, gtfD and ftf genes revealed that each transcript was initiated at a purine residue. Based on these sites, the ‐10 and ‐35 sequences for each transcript were deduced and compared. For all 4 transcripts, the ‐10, but not the ‐35, sequences were homologous to the Escherichia coli promoter consensus sequences.
Effect of acetate on sorbitol fermentation by oral lactobacilli
Authors: Takahashi N. , Kalfas S. , Yamada T.
Published: December 2007

The rate of acid production and end‐products from sorbitol were measured under anaerobic conditions in washed‐cell suspensions of oral strains of Lactobacillus casei subsp, casei and Lactobacillus casei subsp, rhamnosus. The enzymatic activities were assayed in cell extracts of these strains. The cells fermented sorbitol to lactate, formate, ethanol and acetate under anaerobic conditions. Exposure of the cells to air (oxygen) led to inactivation of pyruvate formatelyase and inhibition of anaerobic sorbitol fermentation. In the presence of acetate, air‐exposed cells fermented sorbitol with a concomitant consumption of acetate and production of ethanol and lactate. Acetate also enhanced acid production from sorbitol in cells kept under anaerobic conditions and resulted in formation of lactate and ethanol. Cell extracts of all the strains had NADH‐coupled acetate‐reducing activity, which consisted of sequential reactions of acetate kinase, phosphotrans‐acetylase, acylating aldehyde dehydrogenase and alcohol dehydrogenase. These findings indicate that oral lactobacilli can utilize acetate as an electron acceptor for maintaining their intracellular redox balance during anaerobic sorbitol fermentation in the absence of pyruvate formate‐lyase activity.
Differentiation of the serotype b and species‐specific antigens of Actinobacillus actinomycetemcomitans recognized by monoclonal antibodies
Authors: McArthur W. P. , Stroup S. , McClellan S. , Leung K.‐P.
Published: December 2007

The serotype b antigens have been reported to be associated with lipopolysaccharide. Using murine monoclonal antibodies specific for either a serotype b antigen or the Actinobacillus actinomycetemcomitans species, the relationship of the two epitopes to lipopolysaccharide was determined. Both the species‐specific and serotype b‐specific monoclonal antibodies bound to whole cells, vesicles and conventionally isolated lipopolysaccharide and polysaccharide material derived from A. actinomycetemcomitans culture supernatants. Serotype b‐specific monoclonal antibodies bound to the polysaccharide of acid‐hydrolyzed lipopolysaccharide. Species‐specific monoclonal antibodies bound to both the polysaccharide and the lipid A fraction of lipopolysaccharide after acid hydrolysis. Polymyxin b partially inhibited the binding of the species‐specific monoclonal antibodies to lipopolysaccharide and had no effect on the binding of the serotype b‐specific monoclonal antibodies to lipopolysaccharide. Lipopolysaccharide from whole bacteria and polysaccharide material isolated from culture supernatants were separated by gel filtration chromatography in deoxycholate into fractions that contained serotype b antigen, both serotype b and species‐specific antigens, or species‐specific antigen. SDS‐polyacrylamide gel electrophoresis and Western blotting analysis of the fractions revealed that the serotype b antigen was on a high‐molecular‐weight polysaccharide material. The species‐specific antigen was on a ladder of lower‐molecular‐weight polysaccharides identical to the blot pattern of lipopolysaccharide molecules separated by polyaerylamide gel electrophoresis and stained with silver stain. Chemical analysis of the polysaccharide containing serotype b antigen revealed 85% ribose, 11% glucose, and no lipid. Chemical content of the species‐specific antigenic material revealed a composition typical of lipopolysaccharide. Immunoelectron microscopy using the species‐ or serotype b‐specific monoclonal antibodies confirmed the biochemical and immunological characterization of the two antigens, showing that the species‐specific epitopes were on the surface of the A. actinomycetemcomitans cell membrane and the serotype b‐specific epitopes on the amorphous material extending from the cell surface. The data indicated that the serotype b antigen, detected by the antibody, was separable from lipopolysaccharide and was an A. actinomycetemcomitans capsular material. The species‐specific antigen, being more conserved than the serotype antigen, was on all the lipopolysaccharide molecular species.
Shared antigens of Porphymmonas gingivalis and Bacteroides forsythus
Authors: Vasel D. , Sims T. J. , Bainbridge B. , Houston L. , Darveau R. , Page R. C.
Published: December 2007

Periodontitis in humans is caused by a group of predominantly gram‐negative, anaerobic bacteria among which Porphyromonas gingivalis and Bacteroides forsythus are prominent. A similar group is present and presumably plays a similar role in experimental periodontitis in the primate Macaca fascicularis. Nevertheless, immunization using a vaccine containing only killed P. gingivalis suppresses the progress of experimental periodontitis in M. fascicularis. We investigated the hypothesis that gram‐negative period ontopathic bacteria may share antigens, and immunization with one species may induce antibodies reactive with other gram‐negative species. Using enzyme‐linked immunosorbent assay (ELISA), Western and dot immunoblots with nonabsorbed and absorbed and immune and preimmune sera we show that monkeys immunized with P. gingivalis produce antibodies reactive not only with antigens of P. gingivalis but also with those of B. forsythus. Similarly, rabbits immunized with P. gingivalis or with B. forsythus produce antibodies that react with antigens of both bacteria. Cross‐reactive antibodies bind to epitopes in lipid A and possibly in core carbohydrate of lipopolysaccharide. Using complexes of lipopolysaccharide with polymyxin B, bovine serum albumin and apolipoprotein A1 specificity of binding was documented. Using sera from monkeys immunized with P. gingivalis, cross‐reactivity with Actinobacillus actinomycetemcomitans could not be demonstrated by ELISA, although binding to lipopolysaccharide but not to lipid A was demonstrated by Western and dot immunoblots. Antibodies to shared lipopolysaccharide epitopes of periodontopathic bacteria may account, at least in part, for the immune protection observed in immunized monkeys, and shared epitopes may have potential as a vaccine for periodontitis in humans.
Direct detection of Streptococcus mutans in human dental plaque by polymerase chain reaction
Authors: Igarashi T. , Yamamoto A. , Goto N.
Published: December 2007

Igarashi T, Yamamoto AA, Goto N. Direct detection of Streptococcus mutans in human dental plaque by polymerase chain reaction.

Streptococcus mutans is an etiological agent in human dental caries. A method for the detection of S. mutans directly from human dental plaque by polymerase chain reaction has been developed. Oligonucleotide primers specific for a portion of the dextranase gene (dexA) of S. mutans Ingbritt (serotype c) were designed to amplify a 1272‐bp DNA fragment by polymerase chain reaction. The present method specifically detected S. mutans (serotypes c, e and f), but none of the other mutans streptococci: S. cricetus (serotype a), S. rattus (serotype b), S. sobrinus (serotypes d and g), and S. downei (serotype h), other gram‐positive bacteria (16 strains of 12 species of cocci and 18 strains of 12 species of bacilli) nor gram‐negative bacteria (1 strain of 1 species of cocci and 20 strains of 18 species of bacilli). The method was capable of detecting 1 pg of the chromosomal DNA purified from S. mutans Ingbritt and as few as 12 colony‐forming units of S. mutans cells. The S. mutans cells in human dental plaque were also directly detected. Seventy clinical isolates of S. mutans isolated from the dental plaque of 8 patients were all positive by the polymerase chain reaction. These results suggest that the dexA polymerase chain reaction is suitable for the specific detection and identification of S. mutans.
Tetracycline resistance in Prevotella isolates from periodontally diseased patients is due to the tet(Q) gene
Authors: Olsvik B. , Flynn M. J. , Tenover F. C. , Slots J. , Olsen I.
Published: December 2007

Tetracycline‐resistance in gram‐negative periodontal bacteria is often due to the presence of the tet(Q) gene. In the present study the polymerase chain reaction (PCR) was used to examine 54 isolates of gram‐negative anaerobic rods (Prevotella intermedia, Prevotella nigrescens and related or Bacteroides‐like species) for the presence of the tet(Q) gene. The isolates were recovered from 42 patients with periodontal disease living in northern Europe and North America. An 814 base‐pair segment of the tet(Q) gene was amplified from all 41 isolates resistant to tetracycline with minimal inhibitory concentrations of 4 μg/ml and above. The presence of the tet(Q) gene was verified using hybridization with a specific oligonucleotide internal to the amplified region and restriction endonuclease digestion with DdeI. A PCR product of the same size was also amplified from one tetracycline susceptible isolate (minimal inhibitory concentration=0.5 μg/ml). However, this isolate and the one isolate that was resistant to tetracycline at 4 μg/ml showed a weaker signal than the remaining isolates when hybridized with the internal probe. Typing of the PCR products using restriction endonuclease digests with AluI and HpaII revealed two clusters of distinct electrophoresis patterns, indicating that two different subtypes of the tet(Q) gene were present in this material. A control strain containing the tet(Q) gene from Bacteroides thetaiotaomicron had a different electrophoresis pattern for AluI. This study indicated that subtypes of the tet(Q) gene in tetracycline‐resistant gram‐negative periodontal bacteria exist both within the same patient and within the same species.
Characterization of a 4.2‐kb plasmid isolated from periodontopathic spirochetes
Authors: Chan E. C. S. , Klitorinos A. , Gharbia S. , Caudry S. D. , Rahal M. D. , Siboo R.
Published: December 2007

Oral anaerobic treponemes are associated with active periodontal disease and may comprise up to 57% of the microbiota in periodontal pockets. Four treponeme strains (designated U2a, U2b, U9b, and U9c) isolated from clinical cases were found to harbor a new 4.2‐kb plasmid when plasmid DNA was extracted and purified employing the Qiagen Plasmid Kit. This plasmid differs from the smaller plasmids (2.0‐, 2.6‐, and 2.7‐kb) reported previously by others in Treponema denticola. The newly discovered 4.2‐kb plasmid was found to be the same in all four treponeme strains by restriction endonuclease analysis. It is a circular plasmid since restriction with Pst I, Pvu II, Sma I, Xma I, Ava I or Bam HI produced a single band of the same size. Bacterial strain U2b was shown to be Treponema socranskii and U9c to be T. denticola. The plasmid is designated “pTS1”. The presence of the same plasmid in different species of the treponemes isolated from the same patient suggests the possibility of a naturally occurring genetic transfer system within the oral spirochetes or their ability to take up and maintain mobilizable plasmids.
Lipopolysaccharide increases cell surface‐associated fibronectin in fibroblasts in vitro
Authors: Kubo K. , Kamada T. , Okamoto H. , Izumi Y. , Otsuji S. , Sueda T.
Published: December 2007

This study assessed the fibronectin expression by fibroblasts derived from non‐inflamed and inflamed gingiva by measuring the amount of cell surface‐associated fibronectin and fibronectin released into the medium. The effects of added lipopolysaccharide from Porphyromonas gingivalis, Prevotella nigrescens and Escherichia coli on both types of fibroblasts were also studied. In the absence of lipopolysaccharide, the amounts of the two types of fibronectin were significantly larger in the fibroblasts from inflamed than from non inflamed gingiva. The specific lipopolysaccharide had no effect on the amount of fibronectin released into the medium by either fibroblast type. The amount of cell surface‐associated fibronectin increased significantly when lipopolysaccharide (0.1 and 1 μg/ml) was added to the cells from the non inflamed gingiva (the effect was evident in the order: P. gingivalis >P. nigrescens >E. coli). Lipopolysaccharide from P. gingivalis significantly increased the cell surface‐associated fibronectin even at a low concentration of lipopolysaccharide (0.01 μg/ml). In fibroblasts from the inflamed gingiva, only the lipopolysaccharide from P. gingivalis was effective in increasing the amount of cell surface‐associated fibronectin. Our findings showed that the fibronectin expression was increased in the fibroblasts from inflamed gingiva and that lipopolysaccharide from P. gingivalis increased the cell surface‐associated fibronectin.
Evidence for involvement of superantigens in human periodontal diseases: skewed expression of T cell receptor variable regions by gingival T cells
Authors: Zadeh H. H. , Kreutzer D. L.
Published: December 2007

Immunomodulation by periodontopathic bacteria has been implicated in the pathogenesis of inflammatory periodontal diseases. A novel class of microbial‐derived T cell mitogens, referred to as superantigens, has recently been described. Superantigens are unique in that they induce a tremendous activation and expansion of specific subsets of T cells in an antigen‐independent manner, thereby causing immune dysfunction. Subsets of superantigen‐expanded T cells can be identified with reagents that discriminate among different families of the variable domains of the T cell antigen receptor β‐chain (Vβ). Since superantigens expand one or a few of these T cell antigen receptor Vβ families, T cell subsets that have been expanded by superantigens have restricted expression of one or a few Vβ families. In the present study, we investigated the presence of putative superantigen‐stimulated T cells in periodontitis sites, utilizing a panel of monoclonal antibodies to T cell antigen receptor Vβ families. Leukocytes were isolated from gingival tissues obtained from 8 periodontitis and 4 non‐periodontitis patients by collagenase digestion. Three‐color flow cytometric analysis of these gingival cells demonstrated that in most periodontitis patients examined, patterns of Vβ expression among T cells are characteristic of superantigen stimulation, i.e., there is an elevation in the proportion of one or a few Vβ families. Specifically, these analyses revealed that T cell subsets expressing Vβ5a and Vβ5b, Vβ6, Vβ8 and Vβ12 were each elevated greater than 2 standard deviations in at least one periodontitis patient compared with the mean of the non‐periodontitis subjects. In some periodontitis patients, a less marked elevation of T cells that express Vβ3, Vβ5a, Vp5b, Vβ6, Vβ8, Vβ12 and Vβ13 was noted (greater than 1 standard deviation higher than the mean of the Vβ families in non‐periodontics subjects). Interestingly, VP8+ T cells were elevated to some degree in all periodontitis patients examined. In contrast, T cells expressing Vβ2, Vβ17 and Vβ19 were not significantly different in any of the subjects studied. In most periodontitis but not non‐periodontitis patients, up to 50% of all gingival T cells expressed one or a few T cell antigen receptor Vp families, suggesting that superantigens constitute a major pathway of T cell activation and expansion. Hence, our data support the hypothesis that a large proportion of T cells in periodontitis sites have been stimulated and expanded by superantigens, presumably produced by periodontitis‐associated bacteria.
Genetic evidence for the relationship of Porphyromonas gingivalis cysteine protease and hemagglutinin activities
Authors: Yoneda M. , Kuramitsu H. K.
Published: December 2007

Cysteine protease and hemagglutinin activities of Porphyromonas gingivalis have been implicated as virulence factors in periodontitis. In addition, a close structural relationship between these factors has been suggested. In order to examine the molecular basis for such a relationship, we constructed an isogenic mutant. G‐102, of P. gingivalis 381 deficient in Arggingipain cysteine protease activity. The mutant displayed not only reduced protease activity but also significantly reduced hemagglutination activity compared with the wild‐type strain. Therefore, this investigation provided genetic evidence for the recently proposed structural relationship between Arg‐gingipain and the hemagglutinin activity of P. gingivalis strains.
Associations between Porphyromonas gingivalis and oral treponemes in subgingival plaque
Authors: Riviere G. R. , Smith K. S. , Carranza N. , Tzagaroulaki E. , Kay S. L. , Dock M. , Zhu X. , DeRouen T. A.
Published: December 2007

Colonization and/or proliferation of Treponema denticola may depend on the presence of Porphyromonas gingivalis. The aims of this study were to confirm this synergistic relationship, to determine whether other oral bacteria were similarly associated with P. gingivalis and to relate coinfection to the periodontal status of plaque donors. Subgingival plaque was collected from every tooth except third molars in 106 subjects who were grouped by their worst periodontal condition. In addition to P. gingivalis, monoclonal antibodies were used to identify Campylobacter rectus, Eikenella corrodens, T. denticola, Treponema socranskii and pathogen‐related oral spirochetes. Associations of these bacteria with coinfection by P. gingivalis were assessed by estimated odds ratios. The results indicate that coinfection with P. gingivalis is linked to all tested bacteria, but each pair was associated with distinct periodontal conditions. The distribution of coinfected sites suggests biased colonization of facial surfaces over lingual surfaces.
Comparison of the indigenous oral microbiota and immunoglobulin responses of athymic (nu/nu) and euthymic (nu/+) mice
Authors: Marcotte H. , Lavoie M. C.
Published: December 2007

The role of the immune system in the homeostasis of indigenous oral bacterial populations is poorly understood. In this study, we compared the evolution of the indigenous oral microbiota of specific pathogen‐free athymic nude (nu/nu) BALB/c mice with that of their corresponding phenotypically normal (nu/+) littermates. We also evaluated corresponding salivary and serum antibody activities (IgA and IgG) against the predominant indigenous oral bacteria. The bacterial species recovered from the two mouse strains were Lactobacillus murinus, Enterococcus faecalis, Streptococcus oralis and Staphylococcus epidermidis. From 27 days of age, nu/+ and nu/nu mice had significantly different proportions of oral bacterial populations. When the microbiota stabilized (at 40 days of age), the total cultivable microbiota of nu/+ mice was dominated by L. murinus (65–85%), while that of nu/nu mice was dominated by E. faecalis (40–60%). The precise factors that alter the oral resident microbiota in nu/nu mice are unknown. We found that total salivary IgA levels were significantly lower in nu/nu mice, but no association were observed between the level of salivary IgA antibody against indigenous bacteria and the proportion of these indigenous bacteria in the oral microbiota. The change in the microbiota of nude mice may have been caused by other factors such as defects in other immune functions or cold stress.
A 4‐year longitudinal study of the oral prevalence of enteric gram–negative rods and yeasts in Chinese children
Authors: Sedgley C. M. , Samaranayake L. P. , Chan J. C. Y. , Wei S. H. Y.
Published: December 2007

A 4‐year longitudinal study of the oral prevalence of enteric gram–negative rods and yeasts in 116 Chinese primary school children in Hong Kong was conducted. The oral prevalence of enteric gram‐negative rods for each consecutive year was 25.3%, 37.0%, 24.0%’and 25.8% respectively, with a weighted mean of 27.9%. Enterobacteriaceae, which comprised 57% of all enteric gram–negative rods, were more common in children with no caries experience. The oral prevalence of yeasts for each consecutive year was 7.7%, 12.0%, 14.4% and 15.5% respectively, with a weighted mean of 12.5%. Candida albicans comprised 84% of all yeasts isolated. Oral yeast carriage was significantly associated with caries prevalence. While the oral prevalence of enteric gram–negative rods in primary school children in Hong Kong may be higher than in other parts of the world, repeated isolation of either enteric gram–negative rods or Candida spp. from individual children over the 4–year study period was rare, suggesting that carnage of these organisms is transient.
Genetic characterization of an oral treponeme isolated from human subgingival plaque
Authors: Nakazawa F. , Umemoto T. , Namikawa I. , Hoshino E.
Published: December 2007

We examined the genetic characteristics of a human oral treponeme isolated from subgingival plaque of a patient with periodontal disease that could not be assigned to any of the previously described oral Treponema species. The guanine‐plus‐cytosine content of its DNA was 51 mole%. The levels of DNA‐DNA relatedness of the organism to other Treponema species, including Treponema denticola, Treponema vincentii, Treponema socranskii, Treponema pallidum and Treponema phagedenis, were less than 30%. This study clearly demonstrated that the isolated treponeme could be completely distinct from the established oral Treponema species genetically. Also, the study indicated remarkable genetic heterogeneity among human oral Treponema species.
Characteristics of systemic antibody responses of nonhuman primates to cell envelope and cell wall antigens from periodontal pathogens
Authors: Cox S. E. , Holt S. C. , Ebersole J. L.
Published: December 2007

The immune response of the primate, Macaca fascicularis, to cell envelope (CEA) or cell wall (CWA) antigens of several periodontal pathogens was examined to develop a strategy to interfere with ligature‐induced periodontitis. Animals were parenterally immunized with CEA of either Porphyromonas gingivalis, Prevotella intermedia or a combination of CEA/CWA of Campylobacter rectus, Fusobac‐terium nucleatum and Actinomyces viscosus. Serum samples were taken every 2–4 weeks over a 4‐month period, which included a 13‐week interval with molar teeth ligated. All of the nonhuman primates in the study exhibited baseline levels of IgG, IgM and IgA antibody to formalinized whole cells of the bacteria. These levels increased significantly following immunization and were elevated above baseline throughout the remainder of the experiment. The largest change in antibody responses was seen in IgA antibody levels to P. gingivalis and C. rectus (42‐fold above baseline), IgM antibody to P. intermedia, (41‐fold increase) and IgG antibody to F. nucleatum and A. viscosus (32 and 63‐fold increases). Moreover, the nonhuman primates exhibited differences in isotope response levels to whole microorganisms compared with the cell envelope antigens. These findings demonstrate the capacity of these nonhuman primates to produce an active immune response to microorganisms chronically colonizing the subgingival microbiota. Additionally, it appears that the bacteria may exhibit some unique differences in their immunogenicity as detected by the nonhuman primate and may contribute to the ability of the immune responses to effectively interact with these pathogens.
Arbitrarily primed polymerase chain reaction fingerprinting and clonal analysis of oral Fusobacterium nucleatum isolates
Authors: George K. S. , Reynolds M. A. , Falkier W. A.
Published: December 2007

F. nucleatum is the most commonly isolated microorganism from subgingival plaque, but the role of this microorganism in periodontal diseases remains undefined. Arbitrarily primed polymerase chain reaction (AP‐PCR) was evaluated as a method for fingerprinting F. nucleatum isolates and for use in clonal analysis. Pulsed field gel electrophoresis was used to further differentiate F. nucleatum isolates, with identical AP‐PCR patterns. Extremely heterogeneous AP‐PCR fingerprints were observed among the 98 F. nucleatum isolates, with 36 different genotypes observed with primer CI and 30 different genotypes detected with primer C2. Combining the results of the AP‐PCR genotype analysis from C1 and C2 primer amplifications revealed that up to 7 different genotypes could be distinguished from isolates from the same oral cavity and that up to 4 different genotypes were observed within a single site. An intense amplicon at approximately 450 bp generated in AP‐PCR amplification with primer C2 was associated with F. nucleatum subsp. nucleatum (ATCC 25586) and with 15 F. nucleatum isolates from diseased sites and 2 isolates from healthy sites. Pulsed field gel electrophoresis confirmed the AP‐PCR genotypes and demonstrated increased discriminatory power over AP‐PCR. The results indicated that AP‐PCR and pulsed field gel electrophoresis provide a simple and sensitive means for differentiating oral F. nucleatum isolates and further demonstrate the heterogeneity of this species. These techniques may serve as useful tools in the clonal and epidemiological analysis of F. nucleatum isolates, which may help define the role of these microorganisms in periodontal diseases.