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D Grenier

Publications and source records attributed to D Grenier.

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Functional characterization of extracellular vesicles produced by Bacteroides gingivalis.

Extracellular vesicles of Bacteroides gingivalis (type strain 33277) were isolated, and some of their biological activities were characterized. The vesicles were obtained from a 2-day culture after ammonium sulfate precipitation, differential centrifugation, and dialysis. When viewed by electron microscopy, vesicles of approximately 50 nm predominated. The results indicated that the enriched vesicle fraction had a high proteolytic activity against collagen, Azocoll, and N-alpha-benzoyl-DL-arginine p-nitroanilide. The polypeptide pattern of the vesicles was similar but not identical to that of the outer membrane. The membrane vesicles could also promote bacterial adherence between homologous cells as well as mediate attachment between two noncoaggregating bacterial species. These vesicles could thus play an important role in periodontal diseases by serving as a vehicle for toxins and various proteolytic enzymes, as well as being involved in adherence.

Bacterial Adhesion↗

Isolation of a membrane-associated Bacteroides gingivalis glycylprolyl protease.

A low-molecular-weight proteolytic enzyme was purified 47-fold from outer membranes of Bacteroides gingivalis ATCC 33277 by preparative polyacrylamide gel electrophoresis. The enzyme was present in all B. gingivalis strains tested but was not found in other species of black-pigmented Bacteroides. The molecular weight, determined by sodium dodecyl sulfate (SDS)-polyacrylamide gel electrophoresis, was 19,500 when the enzyme was heated to 100 degrees C in SDS before electrophoresis and 29,000 when it was mixed with SDS but not heated. The optimum pH, with azocasein as the substrate, was between 6.0 and 6.5. The activity was inhibited by phenylmethylsulfonyl fluoride, N-alpha-p-tosyl-L-lysine chloromethyl ketone, Hg2+, and various reducing agents. The enzyme was active against azocasein, azocoll, proline-rich protein from saliva, and the synthetic peptide glycyl-L-proline-p-nitroanilide. The enzyme did not degrade acid-soluble collagen nor did it hydrolyze various arginine- and lysine-containing synthetic substrates.

Bacterial Outer Membrane Proteins↗

Selected characteristics of pathogenic and nonpathogenic strains of Bacteroides gingivalis.

Strains of Bacteroides gingivalis were compared for the presence of properties associated with pathogenicity. Some strains were infectious in pure culture in an in vivo model (guinea pig), and all but one of these were more collagenolytic than those which failed to cause lesions in guinea pigs. However, other factors seem to be necessary for the induction of an infection in this animal model.

Animals↗

Nutritional relationships between oral bacteria.

Nutritional relationships were revealed during the coculturing of Bacteroides gingivalis with Wolinella recta and Bacteroides melaninogenicus with W. recta. W. recta produced a substance that stimulated the growth of B. gingivalis and B. melaninogenicus. Characterization by thin-layer chromatography and absorption spectrometry identified the compound as protoheme. Production of large amounts of formate by B. melaninogenicus stimulated the growth of W. recta. These nutritional relationships could represent examples of mechanisms favoring bacterial succession in periodontal sites.

Bacteroidaceae↗

Detection of collagenase activity in oral bacteria.

Collagenolytic activity of 12 species of oral bacteria was assessed using two methods of detection. Except for two species, all bacterial strains tested were capable of degrading at least one general protein substrate. Results of collagenolytic activity in a growth assay indicate that Bacteroides gingivalis is the only bacterium capable of degrading collagen when the substrate is sterilized using ethylene oxide. However, if the substrate is sterilized by autoclaving, in the presence or absence of the growth medium, other bacterial species could be shown to be collagenolytic. Collagenolytic activity was also demonstrated when whole or broken cells were used in a [14C]collagen assay. Results from this assay and from inhibition studies indicate that collagenolytic activity can either be the result of the combined activities of both a specific collagenase and nonspecific proteases (B. gingivalis) or nonspecific proteases only (other strains in this study), although in the latter case, the time taken to hydrolyze collagen can be 10 times longer than with a specific collagenase.

Bacteria↗

Cytotoxic effects of culture supernatants of oral bacteria and various organic acids on Vero cells.

Dilute culture filtrates of the three asaccharolytic black-pigmented Bacteroides species as well as B. macacae, Fusobacterium nucleatum, and animal strains resembling B. gingivalis were cytotoxic for the African green monkey kidney cell line (Vero cells). Butyric or propionic acid, major metabolic products of these bacteria, seem to be responsible for the characteristic morphological changes observed. Among the organic acids studied, only butyric, propionic, and valeric acids exhibited cytotoxic effects. Finally, cumulative effects (as opposed to synergistic) were observed when mixtures of two organic acids were tested.

Animals↗

Properties of oral asaccharolytic black-pigmented Bacteroides.

Bacteroides endodontalis, a newly described asaccharolytic black-pigmented Bacteroides, along with the other two recognized species of this group (B. gingivalis and B. asaccharolyticus) were studied for their susceptibility to various dyes and inhibitory agents and for some of their enzymatic activities to facilitate differentiating between them. Bacteroides endodontalis resembles B. asaccharolyticus physiologically except for the fact that the former cannot grow on media containing methylene blue, neutral red, or 3% sodium chloride, whereas B. asaccharolyticus can. On the other hand, B. endodontalis and B. gingivalis can grow on a medium containing Congo red while B. asaccharolyticus cannot.

Bacteroides↗

[Studies of mixed anaerobic infections involving Bacteroides gingivalis].

The infectiousness of several combinations of bacteria containing Bacteroides gingivalis and other bacteria associated with periodontal diseases was evaluated by subcutaneous injection to guinea pigs. Among the seven mixtures studied, only one permitted the development of an infection easily transmissible to a second guinea pig; this bacterial mixture was composed of B. gingivalis, Fusobacterium nucleatum, Eubacterium saburreum, and Capnocytophaga ochracea (formerly Bacteroides ochraceus). Owing to its ability to synthesize many lytic enzymes and potentially cytotoxic products, B. gingivalis represented the most virulent species of the mixture. Results from in vitro studies suggest that the development of B. gingivalis in the infected animal depends on the growth of C. ochracea. Succinic acid produced in large amount by C. ochracea seems to be one of the growth factors used by B. gingivalis.

Animals↗