bluuebunny/crossref_metadata_2025 huggingface dataset:
Purification and properties of sorbitol‐6‐phosphate dehydrogenase from oral streptococci
Authors: Svensäter G. , Edwardsson S. , Kalfas S.
Published: December 2007

The activity of sorbitol‐6‐phosphate (S6P) dehydrogenase (S6PDH) and the sorbitol transport system were studied in strains of the oral streptococci Streptococcus gordonii, Streptococcus mitts, Streptococcus sanguis and Streptococcus mutans. Genetically transformed (to ferment sorbitol) strains and their DNA donors were included. S6PDH was purified by anion exchange chromatography and gel filtration. The purity of the enzyme was confirmed by polyacrylamide gel electrophoresis. The purified enzyme from all the strains exhibited Michaelis‐Menton saturation kinetics. The Km values for nicotinamide‐adenine dinucleotide (NAD) and S6P ranged between 0.03 and 0.21 mM and 0.07 and 0.20 mM respectively. The relative molecular weights of the native enzyme were 229,000 for one donor‐transformant pair (S. sanguis and S. gordonii), 107,000 for the other pair (S. mitis and S. gordonii) and 129,000 for S. mutans. The molecular weights of the S6PDH subunits ranged from 26,000 to 28,000. The pH optima (>8.5) and the amino acid composition (15 amino acids examined) were similar for the S6PDH from the different strains. However, the chromatographic and electrophoretic patterns as well as the Km values for NAD and S6P were the same only between the S6PDHs from the strains within each donor‐transformant pair. Purified S6PDH from S. mutans also exhibited low mannitol‐1‐phosphate dehydrogenase activity. Sorbitol‐grown decryptified cells of all the strains phosphorylated sorbitol in the presence of phosphoenolpyruvate but not in the presence of adenosine triphosphate (ATP). ATP‐mediated phosphorylation of glucose was observed with the same strains when grown on glucose. No evidence for a non‐phosphotransferase transport system was found for sorbitol in any of the strains. The results suggest that the initial catabolism of sorbitol in oral streptococci involves a phosphotransferase system and the enzyme S6PDH. S6PDH from S. sanguis, S. mitis and S. mutans exhibits similarities in amino acid composition, pH optimum and subunit molecular weight but the Km values for NAD and S6P as well as the electrophoretic and chromatographic patterns of the enzyme differ.
Immunochemical and structural characterization of the antigenic polysaccharide from Eubacterium saburreum T18
Authors: Nakazawa F. , Hoshino E.
Published: December 2007

An antigenic surface polysaccharide produced by Eubacterium saburreum strain T18, isolated from human dental plaque, was purified from formamide extract of whole cells. Methylation analysis, Smith degradation, optical rotation data and nuclear magnetic resonance spectra demonstrated that the purified antigen was a homopolysaccharide composed of D‐glycero‐D‐galacto‐heptose (Hep.) residues. The structure of the repealing unit in the polysaccharide was: ‐(→6)‐[α‐Hep.furanosyl‐(1→4)]‐β‐Hep.pyranosyl‐(1→6)‐[α‐Hep.furanosyl‐(1→2), α‐Hep.furanosyl‐(l→4)]‐β‐Hep.pyranosyl‐(1‐)4→6)‐β‐Hep.pyranosyl‐(l→. No heptose residues were acetylated. Immunodiffusion reactions in agar gel suggested that the immunodeterminant of the antigenic polysaccharide was D‐glycero‐D‐galacto‐heptofuranosyl residues as branched nonreducing terminals.
Frequency and stability of mono‐or poly‐infection by Actinobacillus actinomycetemcomitans serotypes a, b, c, d or e
Authors: Saarela M. , Asikainen S. , Alaluusua S. , Pyhälä L. , Lai C.‐H. , Jousimies‐Somer H.
Published: December 2007

The aims of the study were to determine the prevalence of simultaneously multiple Actinobacillus actinomycetemcomitans serotypes in one individual, stability of infection by the same serotype and the occurrence of previously not described serotypes of A. actinomycetemcomitans. The serotypes of 515 clinical isolates of A. actinomycetemcomitans from 91 Finnish, Caucasian subjects, including 321 follow‐up samples from 51 subjects, were determined with immunodiffusion assay. Most subjects (n= 86, 95%) were infected with one serotype only; 466 (91%) isolates from 80 subjects belonged to serotype a (25%) of isolates/25 subjects), b (25% of isolates/27 subjects) or c (41% of isolates/30 subjects). Fifteen isolates from 4 subjects reacted with the antiserum raised against previously untypable clinical strain IDH 781 (serotype d) and 18 isolates from 5 subjects with the antiserum raised against strain IDH 1705 or IDH 3096 (serotype e). Sixteen (3%) isolates from 5 subjects remained untypable. The same infecting A. actinomycetemcomitans serotype(s) persisted for the 1–6 years of follow‐up. In conclusion, the study indicates a rare simultaneous occurrence of multiple oral A. actinomycetemcomitans serotypes, the stability of infection by the same serotype(s) and the existence of serotypes of A. actinomycetemcomitans not previously described.
Reduced nicotinamide adenine dinucleotide oxidase involvement in defense against oxygen toxicity of Streptococcus mutans
Authors: Higuchi M.
Published: December 2007

The growth inhibition of the Streptococcus mutans group, including Streptococcus mutans. Streptococcus cricetus, Streptococcus rattus and Streptococcus sobrinus, on glucose by oxygen and the properties of reduced nicotinamide adenine dinucleotide (NADH) oxidase induced by oxygen, using a representative oxygen‐tolerant strain, were examined. The growth response to oxygen varied among strains and correlated with the level of NADH oxidase activity in the cell extract. The induced synthesis of NADH oxidase as well as superoxide dismutase was affected by oxygen tension and energy sources. The induced NADH oxidase involved at least two types, major H2O‐forming NADH oxidase and minor H2O2‐forming NADH oxidase activity. In the presence of a scavenger of H2O2, pyruvate, the growth inhibition by oxygen in an oxygen‐sensitive strain (GS5) was protected but not in another oxygen‐sensitive strain (MT8148). The high level of induced H2O‐forming NADH oxidase activity protected against oxygen toxicity.
Correlation between cell‐adherent activity and surface structure in Porphyromonas gingivalis
Authors: Watanabe K. , Yamaji Y. , Umemoto T.
Published: December 2007

The cell‐adherent ability of 6 strains of Porphyromonas gingivalis (381, ATCC 33277, SU63, KD1, W50 and W83) was compared by using radiolabeled bacterial cells and human gingival fibroblasts (Gin 1), human periodontal ligament fibroblasts (HPLF) and human epithelial cells (Ca9‐22) that had been grown on collagen beads. The cell‐adherent activity of these organisms varied among strains; P. gingivalis strains 381, ATCC 33277 and SU63 bound to the target cells at a range of 14% to 72%, but the other 3 strains (KD1, W50 and W83) were scarcely bound (0.6% to 3.5%). On the other hand, whole bacterial cells and culture supernatants of all strains showed distinct hemagglutinating activity. The 3 strains showing high cell‐adherent activity were hydrophobic and the other strains showing less activity were relatively hydrophilic. Furthermore, a number of peritrichous fimbriae were found on the surface of P. gingivalis strains 381, ATCC 33277 and SU63, which showed high adherent activity, whereas, fimbriae on the other 3 strains showing low adherent ability were barely apparent. Therefore, it was assumed that the cell‐adherent activity of P. gingivalis was related to the hydrophobicity of the cell surface, which was related to the number of fimbriae.
An analysis of the subgingival microflora in randomly selected subjects
Authors: Papapanou P. N. , Sellén A. , Wennström J. L. , Dahlén G.
Published: December 2007

The aim of this study was to describe the presence of some microbial species in the subgingival microflora of a randomly selected subject sample. A further objective was to analyze the association between some microbiological characteristics and the periodontal conditions of the subjects. A total of 171 individuals aged 30–65 years were included. A pooled subgingival plaque sample was obtained from the mesial aspect of the 6 Ramfjord teeth by the use of the paper‐point method. The samples were processed and analyzed according to standardized procedures. The periodontal condition of the subjects was examined clinically and included assessment of plaque, gingivitis, probing pocket depth and probing attachment level. The results showed that 81% of the subjects were positive for Campylobacter rectus, 80% for Fusobacterium nucleatum, 77% for Streptococcus sanguis, 72% for Streptococcus mitis, 66% for Eikenella corrodens, 58% for Prevotella intermedia, 32% for Capnocytophaga, 21% for Selenomonas, 25% for Actinobacillus actinomycetemcomitans, 14% for Porphyromonas gingivalis and 13% for Streptococcus mutans. Small differences in the periodontal conditions were observed between subjects harboring P. gingivalis, A. actinomytecomitans or >5%P. intermedia and the rest of the sample. Similarly small differences in the clinical parameters were revealed when the subjects were grouped according to proportions of disease‐associated and health‐associated species in their subgingival samples.
Bacteria reactive to plaque‐toxin‐neutralizing monoclonal antibodies are related to the severity of gingivitis at the sampled site
Authors: McAnally J. R. , Levine M.
Published: December 2007

Certain monoclonal antibodies (Mabs) neutralize the inhibition of HL60 cell growth by bacterial plaque extracts. The aim of this study was to enumerate and identify the bacteria reactive to these Mabs at sites where periodontal disease was measured by gingival index, pocket depth and attachment loss. The bacterial colonies were grown on blood agar and the colonies counted directly and after immunoblotting. Sites that bled on probing (n= 15) had a mean of 3.35% Mab‐positive colonies (SEM = 0.52%) and sites that did not bleed (n= 9) had a mean of 1.61% (SEM = 0.41%). The percentage of Mab‐positive colonies also correlated with gingival index but not with the total number of colonies sampled, attachment loss or pocket depth. Antigen was expressed by 4 genera: strains of Streptococcus sanguis and/or Eikenella corrodens (8 sites), Staphylacoccus epidermidis (4 sites) and Corynebacterium minutissimus (1 site). The results suggest that the antigens detected on these bacteria by the Mabs may be processed to form toxins and maintain gingivitis by irritating the surrounding cells.
Modulation of herpes simplex virus type 1 replication by human salivary secretions
Authors: Bergey E. J. , Gu M. , Collins A. R. , Bradway S. D. , Levine M. J.
Published: December 2007

Saliva functions to protect the oral cavity from pathogenic invasion by modulating the ability of microbes to colonize the oral surfaces or limiting their growth and/or viability. Although the role of salivary secretions in the modulation of the oral bacteria flora has received considerable attention, little is known concerning its role in viral pathogenesis. Accordingly, the purpose of this study was to assess the effect of salivary secretions on herpes simplex virus type 1 (HSV‐1) replication. Initially, HSV‐1 plaque and liter reduction assays were performed to determine the ability of human submandibular/sublingual (HSMSL) and parotid (HPS) salivas to inhibit the early stages of HSV‐1 infection (adsorption and penetration). Our results suggested that both HSMSL and HPS possess cell‐protective and virus neutralization activities, with HSMSL being more active than HPS. Additional experiments were performed to determine the effect of saliva on the yield of virus progeny. Again, HSMSL caused a greater reduction of HSV‐1 replication than did HPS. A similar effect could not be obtained using vaccinia, suggesting that this inhibitory activity of human saliva is selective. Collectively, these results suggest that human salivary secretions can modulate the replication of HSV‐I in vitro.
Adherence and hydrophobicity of invasive and noninvasive strains of Porphyromonas gingivalis
Authors: Naito Y. , Tohda H. , Okuda K. , Takazoe I.
Published: December 2007

The adherence of Porphyromonas gingivalis to cell surfaces of periodontal tissues may play an important role in its pathogenicity. In this study, 12 strains of P. gingivalis, including both invasive and noninvasive strains, were investigated for adherence to gingival ligament components. The test strains of P. gingivalis adhered to collagen, fibronectin, and laminin to significantly different degrees. An overall positive correlation was noted between hydrophobicity and the number of cells that attached to collagen coated on hydroxyapatite beads. Invasive strains had low hydrophobicity and bound less to collagen than did noninvasive strains that possessed strong hydrophobic properties. 3H‐fimbriae extracted from P. gingivalis were found to attach to collagen‐coated hydroxyapatite. The fimbriae extracted from noninvasive strains bound strongly to collagen, whereas invasive strains' fimbriae with low hydrophobicity bound weakly to collagen or saliva‐coated hydroxyapatite. These data suggest that 1) fimbriae play an important role in colonization through their hydrophobic activity; 2) fimbriae of noninvasive strains are associated with the major adhesin for attachment to gingival tissue, whereas fimbriae of invasive strains are weakly involved in adherence; and 3) there is no correlation between colonization and the invasiveness of P. gingivalis.
Ribotyping shows intrafamilial similarity in Actinobacillus actinomycetemcomitans isolates
Authors: Alaluusua S. , Saarela M. , Jousimies‐Somer H. , Asikainen S.
Published: December 2007

This study reports ribosomal RNA gene restriction patterns of 54 Actinobacillus actinomycetemcomitans isolates obtained from 9 families (12 children and 11 parents). The isolates represented serotypes a, b, c and d. The chromosomal DNA extracted from A. actinomycetemcomitans isolates was digested with the restriction endonucleases EroRI, BamHI, HindIII, ClaI and BglI. The DNA fragments were hybridized to the rrnB ribosomal RNA operon of the Escherichia coli chromosome. In 5 families, isolates belonging to the same family (mother and/or father and the children) had identical hybridization patterns when analyzed with all 5 enzymes. In 3 families, each family member harbored only one ribotype of A. actinomycetemcomitans, but at least one member harbored isolates that were of a different ribotype than the other members of the family. In one family, the mother harbored 2 ribotypes (and serotypes), one common with the daughter and one different. In conclusion, the study confirms the previous results that A. actinomycetemcomitans is transmitted intrafamilially.
Binding of hemin and Congo red by oral hemolytic spirochetes
Authors: Scott D. , Siboo I. R. , Chan E. C. S. , Klitorinos A. , Siboo R.
Published: December 2007

Colony‐forming units or cells in suspension of oral anaerobic spirochetes (Treponema denticola, Treponema vincentii and Treponema socranskii) bind hemin and Congo red. Hemin or Congo red binds to a hydrophobia polypeptide receptor that is located in the outer membrane of the bacterial cells and it has a relative molecular mass of 47 kDa. These oral spirochetes also lyse sheep erythrocytes to produce beta‐hemolytic zones around colony‐forming units. The oral spirochetes may acquire iron for growth when they lyse erythrocytes and bind heme from which they may sequester and transport iron into the cells.
K‐antigens in Porphyromonas gingivalis are associated with virulence
Authors: van Winkelhoff A. J. , Appelmelk B. J. , Kippuw N. , de Graaff J.
Published: December 2007

We investigated antigens in spreading and non‐spreading Porphyromonas gingivalis strains. On the basis of differences in virulence in the mouse model, 8 strains were selected for antiserum production in rabbits. Hyperimmune sera were tested by double immunoprecipitation and immunoelectrophoresis. Besides a common antigen, differences in antigenic composition were observed in the thermolabile antigens between all strains tested. Two different heat‐stable antigens were found after heating at 120°C. One such antigen was detected after sonication of the pellet fraction of autoclaved P. gingivalis cells. This antigen cross‐reacted with 6 of the 8 immune sera. This somatic antigen was almost neutrally charged and sensitive to sodium periodate treatment, suggestive of lipopolysaccharide. A second heat‐stable antigen was detected in the supernatant of autoclaved strains of W 83, W50, HG 184 and A7A1‐28. These non‐somatic antigens were strain‐specific, i.e., no cross‐reactivity was found with heterologous hyperimmune sera. An exception was strain W50, which had a non‐somatic heat‐stable antigen which was recognized by W83 antiserum. These antigens were resistant to DNAse, RNAse and proteinase‐K treatment but were degraded by sodium periodate. In immunoelectrophoresis, these antigens appeared to be negatively charged. These properties are characteristics of a K‐antigen, which likely represent a thermostable carbohydrate capsule. The presence of K‐antigen correlates very well with the serum resistance, the low chemiluminescence, the resistance to phagocytosis and the need for opsonization with specific antibodies for complement‐mediated killing of virulent P. gingivalis strains. We propose to designate these antigens in P. gingivalis W83 and W50 as K1, in HG 184 as K2 and in A7A1‐28 as K3.
Involvement of Porphyromonas gingivalis fimbriae in adherence to Streptococcus gordonii
Authors: Lamont R. J. , Bevan C. A. , Gil S. , Persson R. E. , Rosan B.
Published: December 2007

Adherence of Porphyromonas gingivalis to early plaque bacteria, such as Streptococcus gordonii, is considered an important colonization mechanism. The molecules that mediate this interspecies binding have not been determined. Fimbriae were prepared from P. gingivalis 33277 by mild agitation, ammonium sulfate precipitation and DEAE‐Sepharose chromatography. In a nitrocellulose blot adherence assay, purified fimbriae inhibited S. gordonii G9B‐P. gingivalis 33277 binding by up to 54%. In addition, fimbriae bound to S. gordonii cells in a dot‐blot assay. Incubation of fimbriae with S. gordonii cells followed by washing, sodium dodecyl sulfate‐polyacrylamide gel electrophoresis (SDS‐PAGE), electroblotting and probing with P. gingivalis antibodies also revealed that the fimbriae bind to S. gordonii. In contrast, S. gordonii did not interact with fimbriae that were first subjected to SDS‐PAGE and electroblotting or deposited on a nitrocellulose membrane, suggesting that conformational determinants of the fimbriae may be important in binding. The results indicate that binding between P. gingivalis and S. gordonii is mediated, at least in part, by the porphyromonads' fimbriae.
Fimbria damage and removal of adherent bacteria after exposure to acoustic energy
Authors: McInnes C. , Engel D. , Martin R. W.
Published: December 2007

The physical effects of low‐frequency acoustic energy on Actinomyces viscosus were studied with electron microscopy to explore both acoustically induced damage to fimbriae on the surface of these bacteria and acoustic removal of bacteria from saliva‐treated hydroxyapatite disks. A bacterial suspension was exposed to acoustic energy from a laboratory acoustic generator (50 kPa, 200 Hz) and from a new electronic toothbrush, the SonicareTM. The exposed bacteria were examined with electron microscopy after negative staining. A decrease in both the percentage of bacterial surface covered with fimbriae and the fimbria length was observed after acoustic exposure. To study the acoustic effects on adherent bacteria, A. viscosus bound to hydroxyapatite disks were exposed to acoustic energy and examined with scanning electron microscopy. Quantitative evaluation of the micrographs for the number of bacteria present after exposure revealed that acoustic energy removed both bacteria adherent to the hydroxyapatite surface and adherent to each other. The results support the concept that an electronic toothbrush employing low‐frequency acoustic energy may help prevent and control periodontal J diseases by altering bacterial adherence.
Coaggregation of oral lactobacilli with streptococci from the oral cavity
Authors: Willcox M. D. P. , Patrikakis M. , Harty D. W. S. , Loo C. Y. , Knox K. W.
Published: December 2007

The ability of oral lactobacilli to coaggregate with streptococci and actinomycetes was investigated. Of the 7 species of lactobacilli studied, only two were capable of Coaggregation and the Coaggregation was restricted to streptococci. Lactobacillus salivarius strains (2/4) coaggregated with Streptococcus salivarius, Streptococcus gordonii, Streptococcus crista and tufted Streptococcus sanguis II strains. Lactobacillus fermentum (2/3) coaggregated with S. gordonii and S. sanguis. The Coaggregation between L. salivarius and S. salivarius, S. gordonii or tufted S. sanguis II strains was mediated by a protein on the surface of the lactobacilli and was not inhibited by lactose. The Coaggregation between L. fermentum and the streptococci was mediated by protein on the surface of the streptococci and was inhibited by lactose.
Mutans streptococci in caries‐active and caries‐free infants in Tanzania
Authors: Matee M. I. N. , Mikx F. H. M. , de Soet J. S. , Maselle S. Y. , de Graaff J. , van Palenstein Helderman W. H.
Published: December 2007

The occurrence of mutans streptococci species was investigated in Tanzanian infants aged between 1.0 and 3.5 years with rampant caries and without caries using biochemical and immunological techniques. These infants came from Turiani and Singida, where the caries prevalence for this age group was 12.8% and 1.6% respectively. Streptococcus mutans was found to be the only mutans streptococcal species. The other species, including the previously reported Streptococcus rattus, were not found. Consequently, the differences in caries experience observed between and within the two populations could not be explained by differences in mutans streptococcal species.
A comparison of two transport media for saliva and subgingival samples
Authors: Dahlén G. , Pipattanagovit P. , Rosling B. , Möller Å. J. R.
Published: December 2007

This study has shown that microorganisms associated with caries (mutans streptococci, lactobacilli) and marginal periodontitis (Porphyromonas gingivalis, Prevotella intermedia and Actinobacillus actinomycetemcomitans) in pure culture can be stored in VMGA or reduced transport fluid and be recovered sufficiently after 24 h. Sufficient recovery after 24 h of mail transport was also obtained in 37 saliva samples obtained from adults and 80 subgingival samples from a group of patients before and 3, 6 and 15 months after periodontal treatment. The samples transported in VMGA III showed in comparison to the samples transported in reduced transport fluid a higher recovery rate as well as a higher percentage of the total viable count for investigated anaerobic species. This was explained by the different composition of the two media and also by the gel consistency of VMGA III, which maintains a low redox‐potential even during transport after transferring a sample into the medium. In a few samples aerobic bacteria increased in both media.
Immunochemical characterization of the carbohydrate antigens of serotype c/Lancefield group C “Streptococcus milleri”
Authors: Kitada K. , Yakushiji T. , Inoue M.
Published: December 2007

Carbohydrate antigens of the serotype c/Lanceficld group C “Streptococcus milleri” were extracted by autoclaving whole cells of the type c reference strain K51Y. The type c and group C antigen molecules were separated and partially purified by a DEAE‐Sephadex A‐25 column chromatography followed by Sephad‐ex G‐100 gel filtration. The purified type c antigen and group C antigen were homogeneous in the double diffusion and in the immunoelectrophoresis. The type c antigen was composed principally of glycerol, rhamnose, glucose and N‐acetylglucosamine in a molar ratio of 0.22:0.27:1.00:0.48. The quantitative precipitin inhibition test indicated that N‐acetylglucosamine played a major role in immunodeterminant structure. Thus, the type c antigen of “S. milleri” is a new carbohydrate type antigen and is immunochemically different from the Oltens‐type antigen III found occasionally in group C streptococci. In contrast, the group C antigen preparation contained a high proportion of N‐acetylgalactosamine in addition to glycerol, rhamnose, glucose and N‐acelylglucosamine, and the N‐acetylgalactosamine residue was involved in the immunodominant epitope, being in good agreement with the previously proposed chemical structure of the group antigen. N‐acetylgalaclosamine was detected in the autoclaved extracts of a nontypeable/group C strain but not of a type c/ungroupable strain.
Differential pattern of infection and i immune response during experimental oral candidiasis in BALB/c and DBA/2 (H‐2d) mice
Authors: Chakir J. , Côté L. , Coulombe C. , Deslauriers N.
Published: December 2007

We used an experimental model of oral candidiasis in the mouse to investigate the impact of the introduction of Candida albicans into a Candida‐free system. We report that 2 strains of mice with the same major histocompatibility complex haplolype (H‐2d) display different kinetics of primary oral infection after topical application of the same inoculum. The mucosal reactions in both DBA/2 and BALB/c mice involve a similar recruitment of CD4+ and CD8+ T cells and of MAC‐1 + cells in mucosal tissue during the infection. A carrier state is maintained following the resolution of the infection in both strains and is associated with the persistence ofintraepithelial CD4+ T cells. However, there is a time‐specific recruitment of y6 T cells that coincides with a dramatic decrease in viable Candida in the mucosal tissue; this occurs on day 3 in BALB/c mice and on day 6 in DBA/2 mice. The denouement of an oral contact with Candida is also different in the 2 mouse strains, cell‐mediated immunity being triggered in DBA/2 mice but not in BALB/c mice. The different kinetics of Candida clearance in BALB/c vs DBA/2 mice may therefore signal a differential priming of T cell subsets whose modalities do not appear to be associated with the H‐2 complex.
Macaca fascicularis as a model in which to assess the safety and efficacy of a vaccine for periodontitis
Authors: Persson G. R. , Engel L. D. , Whitney C. W. , Weinberg A. , Moncla B. J. , Darveau R. P. , Houston L. , Braham P. , Page R. C.
Published: December 2007

We have assessed Macaca fascicularis as a potential model in which to test the efficacy and safety of a vaccine for periodontitis. Twenty‐eight animals were surveyed and 20 studied in more detail. Clinical periodontal status was assessed, the subgingival microflora analyzed especially for the presence and proportions of Porphyromonas gingivalis and liters and avidities of serum antibodies reactive with P. gingivalis measured. Probing depths ranged from 0.90 mm to 3.80 mm, Gingival Index scores from 0.00 to 4.00 and Plaque Index scores from 0.00 to 3.0About 40% of sites bled on probing. The animals manifested a subgingival flora characteristic of the anaerobic gram‐negative bacteria found in human periodontal pockets, including Actinobacillus actinomycetemcomitans, P. gingivalis, Bacteroides forsytkus, Campylobacter rectus, Prevotella intermedia and Fusobac‐terium niicleatum. P. gingivalis was detected in 70 of 80 samples studied, ranging from 0.01% to 20% of the total flora. Serum antibody reactive with antigens of P. gingivalis was observed in all animals, with titers ranging from 1.0 enzyme‐linked immunosorbent assay (ELISA) unit to 25 ELISA units and avidities from 0.10 M to 2.20 M. Antibody titer and maximum percentage of P. gingivalis were inversely correlated, indicating that a humoral immune response may be effective in reducing P. gingivalis overgrowth. M. fascicularis appears to be an excellent model for use in vaccine development.
Porphyromonas gingivalis lipopolysaccharide stimulation of human monocytes: dependence on serum and CD14 receptor
Authors: Shapira L. , Takashiba S. , Amar S. , Van Dyke T. E.
Published: December 2007

The purpose of this study was to investigate factors influencing the ability of lipopolysaccharide (LPS) derived from Porphyromonas gingivalis to elicit secretion of tumor necrosis factor‐a (TNFα) from human monocytes (adherent mono‐nuclear cells). The results indicate that P. gingivalis LPS stimulation of TNFa from monocytes is comparable to LPS from Escherichia coli. Both LPS, although structurally different, increased TNFα secretion in a dose‐dependent manner. In serum‐free conditions, TNFα secretion was relatively low, but it dramatically increased at human serum concentrations as low as 1%. Maximal secretion was observed in the presence of 10% serum, with a slight decrease at higher serum concentrations. The CD14 molecule is a putative monocyte LPS receptor. When cells were pre‐incubated with a blocking monoclonal antibody (My4) to CD 14, TNFα‐mRNA accumulation and TNFα secretion were reduced to control levels at LPS concentrations of up to 10 ng/ml. At higher LPS concentrations, the blocking effect was only partial, in spite of 50‐fold excess antibody concentration. The blocking effect was observed only in the presence of serum. The effect of the CD14 antibody was dose‐dependent with saturation at 2.5 μg/ml. The results suggest that CD 14 is one of the major receptors for P. gingivalis LPS but highlight the necessity to investigate other cell‐surface receptors mediating P. gingivalis‐LPS interactions. These interactions are believed to be important in the pathogenesis of periodontal destruction.
In vitro modulation of oral bacterial adhesion to saliva‐coated hydroxyapatite beads by milk casein derivatives
Authors: Neeser J.‐R. , Golliard M. , Woltz A. , Rouvet M. , Dillmann M.‐L. , Guggenheim B.
Published: December 2007

Bovine caseinate, derivatives of its glycosylated moiety [caseinoglycomacropeptide (CGP)], and caseinophosphopeptides were evaluated as inhibitors of adhesion of oral bacteria to saliva‐coated hydroxyapatite beads (S‐HA). All milk casein‐derived components behaved as potent inhibitors of Streptococcus sanguis OMZ 9 and Streptococcus sobrinus OMZ 176 adhesion to S‐HA, whereas neither bovine serum albumin nor polyethyleneglycol were able to interfere with the adhesion of these strains. By contrast, none of the molecular species tested was able to inhibit the attachment of Actinomyces viscosus Ny 1 to S‐HA. On the other hand, casein derivatives were shown to displace human serum albumin from S‐HA beads. They were also able to bind to the bacterial cell surface of all strains examined. Collectively, these findings suggest that interactions between acidic casein‐derived milk components and the biological surfaces involved in bacterial adhesion to S‐HA result in an inhibitory effect that is selective for the oral streptococci examined.
The tet(Q) gene in bacteria isolated from patients with refractory periodontal disease
Authors: Olsvik B. , Olsen I. , Tenover F. C.
Published: December 2007

Twenty‐two tetracycline‐resistant (tetr) anaerobic and facultative anaerobic bacteria isolated from periodontal pockets of 12 patients with refractory periodontitis were examined for the presence of the Tet Q determinant by DNA‐DNA hybridization. Dot blots of bacterial DNA were tested with an intragenic digoxigenin‐labelled tet(Q) probe consisting of a 1.45 kb EcoRI/PvuII fragment from plasmid pNFD13‐2. Southern blots of chromosomal DNA digested with the restriction enzyme EcoRI were also examined. The tet(Q) probe hybridized with DNA from 8 of the 22 tetr strains, including 2 Prevotella intermedia strains and one strain each of Prevotella nigrescens, Prevotella loescheii, Prevotella veroralis and Prevotella melaninogenica. The tetr strains of Mitsuokella dentalis and Capnocytophaga ochracea also hybridized with the probe. The lack of discernible plasmid DNA in all the probe‐positive isolates suggests that these tetracycline‐resistance genes were chromosomally encoded. The probe hybridized with a different size fragment in all the isolates. This study extends the number of species that carry the tet(Q) gene to include several outside the genera Prevotella and Bacteroides.
Enumeration of viable oral spirochetes from periodontal pockets
Authors: Qiu Y.‐S. , Klitorinos A. , Rahal M. D. , Siboo R. , Chan E. C. S.
Published: December 2007

We recently developed a successful method for quantifying oral anaerobic spirochetes in pure culture by a viable count. New oral spirochete medium was used with low temperature‐gelling agarose in polystyrene tissue‐culture flasks. We have extended the use of this method to determine the viable count of spirochetes from periodontal pockets. Sixteen subgingival plaque samples were obtained by insertion of sterile paper points into deep periodontal pockets. The points were placed into reduced transport medium at chairside, vortexed in the microbiology laboratory and aliquots of the medium inoculated into molten new oral spirochete‐agarose medium (37°C) containing rifampin (20 μg/ml) in a flask. Subsequent dilutions were made from this initial flask to other flasks containing selective medium in sequence. All flasks were incubated anaerobically. Most other subgingival bacteria were selectively inhibited by rifampin. Spirochete colonies were typically spherical and were either dense or cottony. Their identities were checked by darkfield examination. Counts of colony‐forming units of cultivable spirochetes ranged from 12.5% to 28.2% of the total cultivable anaerobic flora by the method described.
Growth‐inhibitory effect of pyrophosphate on oral bacteria
Authors: Drake D. , Grigsby B. , Krotz‐Dieleman D.
Published: December 2007

The purpose of this investigation was to determine whether pyrophosphate, the anticalculus component of tartar‐control dentifrices, exerts antimicrobial activity against oral bacteria commonly found in supragingival plaque. Minimal inhibitory concentrations of pyrophosphate were determined for Streptococcus sanguis, Streptococcus mutans (serotype c), Actinomyces viscosus and Actinomyces naeslundii. All of the bacteria tested were susceptible to pyrophosphate with identical minimal inhibitory concentrations of 0.67% wt/vol (25 mM). Bactericidal kinetics assays revealed that both S. mutans and A. viscosus were killed by pyrophosphate, with the latter being considerably more susceptible. The mechanism of killing was not due to high ionic strength, as comparable controls showed no loss in numbers of viable cells. Brief exposure (two 5‐min incubations) of S. mutans to pyrophosphate and sodium dodecyl sulfate caused pronounced inhibition of growth over the 24‐h test period. Under the constraints of the conditions used, these studies indicate that pyrophosphate and sodium dodecyl sulfate can substantially inhibit the growth of oral bacteria. These compounds may affect the oral microflora of patients who routinely use tartar‐control dentifrices and mouthrinses.
Catabolite modification of acid tolerance of Streptococcus mutans GS‐5
Authors: Belli W. A. , Marquis R. E.
Published: December 2007

The acid tolerance of Streptococcus mutans GS‐5 in standard pH‐drop assays was found to be affected by the sugar used in the assay and also by the sugar used for growth of the organism. For example, acid tolerance was lower when galactose was used as catabolite than when glucose was used, apparently because galactose/proton symport brought protons extruded by the F‐ATPase back into the cell and thus reduced ?pH across the cell membrane. The acid tolerance of glycolysis was related directly to the capacities of the cells to produce acid glycolytically, or probably more correctly, to their capacities to produce adenosine triphosphate but not to acid tolerance of phosphotransferase systems for sugar uptake. Thus, glycolytic acid tolerance of S. mutans depends not only on environmental factors such as potassium or magnesium levels but also on the specific catabolites the organism is metabolizing or to which it has become metabolically adapted.
Effective method for discriminating between oral bacterial and human alkaline phosphatase activity
Authors: Shibata Y. , Yamashita Y. , Miyazaki H. , Ueno S. , Takehara T.
Published: December 2007

Alkaline phosphatase (ALPase) activity was quantitatively compared in various kinds of oral bacteria. High ALPase activity was detected in 3 species of periodontal bacteria, Porphyromonas gingivalis. Prevotella intermedia and Capno‐cvtophaga sputigena. The ALPase activity detected in these bacteria was almost completely inhibited in the presence of 1% sodium dodecyl sulfate (SDS). By contrast, the activity of mammalian ALPase isoenzymes was not inhibited at all even in the presence of 1% SDS. These results indicate that the ALPase assay in combination with 1% SDS can identify the origin of ALPase detected in gingival crevicular fluid as being from bacteria or from a host response. Clinical examination with adult periodontitis revealed that ALPase activity in gingival crevicular fluid from the patients consisted of a combination of SDS‐sensitive and SDS‐resistant activities. These findings indicate that ALPase activity detected in gingival crevicular fluid originates not only from bacteria but also from a host response.
Innervation of recurrent aphthous ulcers
Authors: Konttinen Y. T. , Häyrinen‐lmmonen R. H. , Nordström D. , Malmström M. J. , Hukkanen M. , Sorsa T. , Polak J.
Published: December 2007

Specimens from nonkeratinized oral mucosa were obtained from diseased and clinically healthy mucosa from 7 patients with minor recurrent aphthous ulcers. The innervation of the specimens was visualized using antibodies to neuron‐specific intermediate cytoskeletal neurofilament fiber, the cytoplasmic protein gene product 9.5 and a 38 kDa integral membrane protein of synaptic vesicles applied in avidin‐biotin‐peroxidase staining. Mapping with these 3 antibodies revealed dense and basically similar pattern of innervation in the specimens of the clinically healthy oral mucosa. In recurrent aphthous ulcers, all 3 general markers disclosed peripheral nerve fibers also in the lesions, apart from the necrotic area, among the inflammatory cells without signs of retraction from the diseased area. Synaptophysin staining suggested that these peripheral nerve fibers in the inflammatory areas still contained synaptic vesicles. Accordingly, they were shown to contain substance P and calcitonin gene‐related peptide, which are known to be relased upon stimulation of the nerve and can exert potent paracrine actions, possibly on the local inflammatory cells as suggested by a close spatial relationship between neuropeptide‐containing nerves and inflammatory cells.
Humoral immune responses to Porphymmonas gingivalis and Actinobacillus actinomycetemcomitans in adult periodontitis and rapidly progressive periodontitis
Authors: Mooney J. , Kinane D. F.
Published: December 2007

The relationships between various forms of periodontal disease and the avidities of serum antibodies of all 3 immunoglobulin (Ig) classes (IgG, IgM and IgA) to Porphyromonas gingivalis and Actinobacillus actinomycetemcomitans were investigated. Twenty‐four patients with untreated adult periodontitis and twelve untreated patients diagnosed as suffering from the early‐onset form of periodontitis, rapidly progressive periodontitis, were studied. The latter group were matched for age and sex to healthy controls. Antibody litres were measured and avidity (expressed as molarity) was further assayed using the thiocyanate elution method. Avidity has previously been shown to relate to the biological function of antibody. IgM avidities to P. gingivalis were lower in the rapidly progressive periodontitis group than in the adult periodontitis group (0.54 M vs 0.74 M). IgG avidities tended to be lower in the former than in the latter group (0.58 M vs 0.92 M). In accordance with other workers, seropositivily was defined as an immunoglobulin titre more than twice the median level of control sera. Only 2 of the rapidly progressive periodontitis group were seropositive. Interestingly, the seronegative rapidly progressive periodontitis patients were significantly different (0.53 M vs 0.92 M). The data that patients with various forms of periodontal disease appear to produce antibodies of differing avidity to P. gingivalis suggest that the quality of the humoral immune response to suspected periodontopathogens may have a bearing on the aetiology of periodontal disease.
Clonal diversity of the Streptococcus mitis biovar 1 population in the human oral cavity and pharynx
Authors: Hohwy J. , Kilian M.
Published: December 2007

A total of 250 isolates of oral streptococci were recovered from swabs of oropharyngeal surfaces of 3 members of one family. All isolates were examined by biochemical and serological means, and 106 isolates were identified as Streptococcus mitis biovar 1. These were typed by restriction endonuclease analysis using the enzymes EcoRI and HaeIII and further characterized by their whole‐cell polypeptide profile patterns in sodium dodecyl sulfate‐polyacrylamide gel electrophoresis. In addition, rabbit antisera raised against 8 reference strains of oral streptococci were used to characterize representative isolates both by their carbohydrate and protein antigens by Ouchterlony and Western blot analyses. Very limited biochemical diversity was observed among the 106 S. mitis biovar 1 isolates. In contrast, 24 different genotypes defined by restriction endonuclease analysis were detected, and each individual carried 6‐13 types. Limited sharing of genotypes was observed between the 3 members of the same family and between the pharyngeal and buccal mucosa of single individuals. The antigenic analyses showed remarkable antigenic diversity between the 24 genotypes. The results provide a basis for studying the population dynamics of an oral commensal species and its interaction with the salivary immune system.
Identification of salivary proteins inhibiting herpes simplex virus 1 replication
Authors: Gu M. , Haraszthy G. G. , Collins A. R. , Bergey E. J.
Published: December 2007

Salivary proteins play an important role in the maintenance of the oral ecology. Previous studies have indicated that human submandibular‐sublingual and parotid salivas can selectively suppress the in vitro infectivity of herpes simplex virus 1. The purpose of this study was to identify the salivary components in human submandibular‐sublingual saliva that modulate in vitro infectivity. Assessment of the interaction of viral particles with salivary components was accomplished using an in vitro solid‐phase assay. These experiments revealed that herpes simplex virus particles selectively interact with the members of the salivary proline‐rich protein and cystatin families. Subsequent yield reduction assays demonstrated the ability of proline‐rich proteins and salivary cystatins to inhibit the viral replication, with basic proline‐rich peptides being more effective. Subsequent assays suggest that basic proline‐rich peptides reduced the virus titer by interfering with penetration and/or cellular processing of virus within the target cell. Collectively, these results further suggest that salivary proteins have an important role in the host defense mechanism against recurrent herpesvirus infection.
Influence of toothbrushing, eating and smoking on Dentocult SM Strip mutans® test scores
Authors: Schlagenhauf U. , Pommerencke K. , Weiger R.
Published: December 2007

The Dentocult SM Strip mutans® test (D‐SM, Vivadent) is a popular method for estimating salivary mutans streptococci and is clinically used for the detection of potential caries risk patients. The influence of toothbrushing, eating and smoking on the reliability of D‐SM test scores was assessed in 30 subjects aged 21‐39 years. All experiments were performed 24 h after professional tooth cleaning in the morning of 3 consecutive days. On day 1 immediately before and 30 min after toothbrushing, D‐SM tests were taken and total salivary colony‐forming units per ml were determined. Following the same protocol, the influence of a standardized breakfast and of smoking was evaluated on day 2 and on day 3 respectively. Although all parameters significantly decreased the salivary colony‐forming units per ml, only the standardized breakfast induced significant changes in the D‐SM test results. Eating therefore should be avoided prior to the performance of the D‐SM test.
Characterization of immunoglobulin G‐degrading proteases of Prevotella intermedia and Prevotella nigrescens
Authors: Jansen H.‐J. , Grenier D. , Van der Hoeven J. S.
Published: December 2007

Degradation of immunoglobulins is thought to be an important factor in the causation of periodontal diseases by hindering local host defenses and by providing nutrients to the periodontal microflora. In this study, we characterized the proteolytic activity against human immunoglobulin G (IgG) of 20 strains of Prevotella intermedia and Prevotella nigrescens isolated from periodontal pockets and oral abscesses. IgG degradation was studied by sodium dodecyl sulphate‐polyacrylamide gel electrophoresis. All strains degraded IgG within 48 h after growth in trypticase‐yeast extract medium (TY) supplemented with 0.3% IgG. Incorporating IgG in TY broth enhanced bacterial growth. Protease profiles (zymography), which revealed the presence of 1‐4 IgG‐degrading proteolytic bands in bacterial cell extracts, became more complex after growth in the presence of IgG. A 38‐kDa protease capable of degrading IgG nonspecifically was present in almost all strains. The proteolytic activity was mainly located on the surface of the cell envelope. Two strains of P. intermedia and P. nigrescens ATCC 33563 were selected for further studies. Bacterial cell suspensions in phosphate‐buffered saline completely degraded human IgG, IgA and IgM within 24 h. This activity depended on reducing conditions and was inhibited at temperatures above 50°C. The pH optimum of immunoglobulin degradation was at pH 7. Strains cultured at 42°C showed a markedly reduced capacity to degrade IgG. Inhibition studies revealed that breakdown of IgG was caused by a cysteine protease(s). The capacity of P. intermedia and P. nigrescens to degrade immunoglobulins may explain their association with polymicrobial oral diseases.
Immunolocalization of bone‐resorptive cytokines in rat pulp and periapical lesions following surgical pulp exposure
Authors: Tani‐Ishii N. , Wang C.‐Y. , Stashenko P.
Published: December 2007

The bone‐resorptive cytokines interleukin 1 (IL‐1) and tumor necrosis factor (TNF) have been implicated in the pathogenesis of many chronic inflammatory diseases, including pulpitis and apical periodontitis. To further elucidate their role in these disorders, we have identified cells that express IL‐1α and TNFα in infected pulps and in developing rat periapical lesions after surgical pulp exposure. As detected by immunohistochemistry, IL‐1α‐ and TNFα‐positive cells were present as early as 2 days after pulp exposure in both the pulp and periapical region. The numbers of cytokine‐expressing cells increased up to day 4 in the pulp and up to day 30 in the periapex. In contrast, cells expressing IL‐1β and TNFβ the homologous forms of these mediators, were not found in pulp or periapical lesions during this period. Cells expressing IL‐1α and TNFα were identified primarily as macrophages and fibroblasts, with occasional staining of polymorphonuclear leukocytes. Osteoblasts and osteoclasts were also positive, whereas lymphocytes were negative. In general, cytokine‐expressing cells were located proximal to abscesses and the root apex. These findings demonstrate that cells that express bone‐resorptive cytokines IL‐1α and TNFα are present immediately after pulp exposure in this model, which supports the hypothesis that these mediators play a key role in pulpal and periapical pathogenesis, including the concomitant bone destruction. They also indicate that both resident connective tissue cells as well as infiltrating cells express bone‐resorptive cytokines in response to infection in these lesions.