P. B. Chen

Rapid Identification of Periodontal Pathogens in Subgingival Dental Plaque: Comparison of Indirect Immunofluorescence Microscopy with Bacterial Culture for Detection of Bacteroides gingivalis*

A large body of research implicates Bacteroides gingivalis in the etiology of adult periodontitis, however, the application of this information to clinical diagnosis and treatment has been hampered by the need for a simple, rapid, and reliable means of detecting this microorganism. In the present study, indirect immunofluorescence microscopy using species specific, polyclonal antisera and a monoclonal antibody was evaluated in the clinical identification and quantitation of B. gingivalis in human subgingival dental plaque. One hundred and twenty subgingival plaque samples were obtained from predetermined sites by means of sterile paper points from 20 human subjects including 10 adult periodontitis patients and 10 periodontally normal subjects. The proportions of cultivable B. gingivalis in each sample were determined following anaerobic culture on nonselective blood agar media and selective media containing kanamycin. These results were then compared to quantitative estimates of B. gingivalis by indirect immunofluorescent microscopic evaluation of heat-fixed plaque smears. Using both immunofluorescence microscopy and bacterial culture, the present study confirms the importance of B. gingivalis in adult periodontitis previously described by culture. The organism was cultivable from 70% of the adult periodontitis patients but not from any of the normal adults. In contrast, indirect immunofluorescence microscopy detected the organism in up to 40% of the subgingival sites in 100% of the adult periodontitis patients as well as four sites in the periodontally normal subjects. The sensitivity of indirect immunofluorescence microscopy compared to culture ranged from 91 to 100% while the specificity varied from 87 to 89%. The numbers of B. gingivalis identified by immunofluorescence microscopy were directly proportional to the severity of periodontal disease as measured by clinical indices. Linear regression analysis of the immunofluorescent data suggests that the probability of a subject having adult periodontitis approaches 100% when B. gingivalis comprises 9% or more of the subgingival microflora. The present study indicates that indirect immunofluorescence microscopy using speciesspecific serodiagnostic reagents is useful in the clinical detection of B. gingivalis in human subgingival dental plaque. The data also suggests that certain cases of adult periodontitis can be diagnosed solely on the basis of this laboratory test.

Differences in lymphoproliferative responses to the bacterium Actinomyces viscosus in various mammalian species.

A water-soluble extract of Actinomyces viscosus (AVS) was tested for its capacity to induce DNA synthesis in lymphocytes from man, monkeys, mice and guinea pigs. The results indicated that the AVS induced an in-vitro lymphoproliferative response, as assessed by tritiated thymidine incorporation, in mouse-spleen cells, in the majority of human peripheral blood samples tested and in macaque monkey spleen cells. The AVS also elicited a blastogenic response from spleen, lymph node and peripheral blood lymphocytes from guinea pigs immunized with A. viscosus. The AVS did not elicit a lymphoproliferative response from human-cord blood cells, monkey peripheral blood lymphocytes, or peripheral blood and spleen lymphocytes from non-immunized guinea pigs. Thus there was a difference in the ability of A. viscosus to induce DNA synthesis in lymphocytes from the different animal species tested.

Cell-mediated and humoral immune responses of guinea-pigs to the oral bacterium Actinomyces viscosus

Actinomyces viscosus is a Gram-positive facultative rod indigenous to most human mouths. The guinea-pig was evaluated as a model for assessing cellular immune responses to a human strain of A. viscosus. Guinea-pigs, immunized with heat-killed A. viscosus cells or a water-soluble extract from A. viscosus (AVS), were tested for delayed-type hypersensitivity responses to A. viscosus 4 weeks after immunization. A week later, the guinea-pigs were terminated. Lymphocytes from peripheral blood, lymph node and spleen were tested for in-vitro blastogenic responses to heat-killed A. viscosus or the water-soluble extract. Plasma from these guinea-pigs were tested for the presence of precipitating antibodies. Positive in-vivo delayed-type hypersensitivity reactions and antibody responses to AVS occurred only in immunized guinea-pigs. A strong in-vitro blastogenic response to AVS or heat-killed A. viscosus occurred in lymphocytes from immunized guinea-pigs, but a weak response was detected in un-immunized animals. These data suggest that the guinea-pig can be used to evaluate different parameters of the immune response to oral bacteria such as A. viscosus.