Media- and method-dependent variations in minimal inhibitory concentrations of antiplaque agents on oral bacteria.

AIMS: To determine minimal inhibitory concentrations (MICs) and the percentage of nonsusceptible bacteria-- those still cultivable above a threshold concentration--in human supragingival dental plaque and saliva for antiplaque/antimicrobial agents including triclosan (TCS) and trichlorocarbanilide (TCC), and a new potential antimicrobial, 2-t-butyl-5-(4-t-butylphenyl)-phenol (DTBBP). METHODS AND RESULTS: Broth and agar dilution-based MIC tests were performed using 28 oral and nonoral bacterial strains representing 17 species. MICs for TCS were lowest and more than 100-fold lower than DTBBP (P < 0.0005) by both methods. MICs for TCS were lower in broth-based tests compared with TCC. The additions of defibrinated blood to agar and horse serum to broth increased MICs--in the case of TCS, 10- to 15-fold. Significantly higher proportions of nonsusceptible plaque and salivary bacteria were recovered from agar media containing DTBBP or TCC compared with TCS (P < 0.05). CONCLUSIONS: TCS is a more effective antimicrobial agent than either TCC or DTBBP as determined by in vitro testing. SIGNIFICANCE AND IMPACT OF THE STUDY: The utility of in vitro testing for antiplaque agents as a predictor of in vivo efficacy is affected by the methods used.

Epidemiology and diagnosis of HIV-associated periodontal diseases.

A review of periodontal disease as a manifestation of HIV infection suggests a shift in emphasis over the past 5 years. Initially the focus was on newly described forms of periodontal disease (i.e., HIV-associated gingivitis or linear gingival erythema (LGE); HIV-associated periodontitis or necrotizing ulcerative periodontitis (NUP). While the clinical definition of LGE varies from study to study, an association between LGE and Candida infection has been described. Furthermore, the prevalence of NUP is quite low and this disorder is associated with severe immunosuppression. In contrast, the focus today is on the accelerated rate of chronic adult periodontitis occurring in seropositive patients. While the organisms that characterize adult periodontitis in seronegative individuals are present in subgingival plaque from seropositive individuals, reports suggest that atypical pathogens are also present (i.e., Mycoplasma salivarium, Enterobacter cloacae). Recent studies from our laboratory have identified a novel strain of Clostridium isolated from the subgingival plaque of injecting drug users that has pathologic potential. This organism, however, was found in both seropositive and seronegative individuals in this cohort, suggesting an association with lifestyle rather than serostatus. In addition, data has been published examining the local host response in periodontitis in seropositive individuals. Distinctly elevated levels of IgG in gingival crevicular fluid (GCF) have been observed in seropositive patients. Furthermore, data from our laboratory examining inflammatory mediators in GCF (polymorphonuclear leukocyte lysosomal enzyme beta-glucuronidase and the pro-inflammatory cytokine interleukin-1 beta) suggests an altered response in patients with HIV infection. The alteration manifests as the absence of the expected strong correlation between polymorphonuclear leukocyte activity in the gingival crevice and clinical measures of existing periodontal disease, as well as elevated levels of interleukin-1 beta in sites with deeper probing depths. Therefore, it can be concluded that the progression of periodontal disease in the presence of HIV infection is dependent upon the immunologic competency of the host as well as the local inflammatory response to typical and atypical subgingival microorganisms.

Analytical performance of an immunologic-based periodontal bacterial test for simultaneous detection and differentiation of Actinobacillus actinomycetemcomitans, Porphyromonas gingivalis, and Prevotella intermedia.

The analytical performance of a membrane-based immunoassay for the simultaneous detection and differentiation of Actinobacillus actinomycetemcomitans, Porphyromonas gingivalis, and Prevotella intermedia (including Prevotella nigrescens) was investigated. Positive reactions were observed for 71 of 71 reference strains and recent oral isolates of A. actinomycetemcomitans, P. gingivalis, and P. intermedia. No cross-reactivity was observed with 39 other common oral and environmental species. The specificity of the test was unaffected by the presence of potential oral interferents including whole blood, white blood cells, mucin, saliva, toothpastes, and oral rinses. A proficiency test by dental professionals using a standardized set of unknown simulated samples yielded a sensitivity of 97% (116/120) and a 100% specificity (240/ 240). An additional group including dental professionals and high school students was shown to be 99% proficient (1385/1397) in distinguishing proper from improper test function when processing control samples with normal test devices and devices with simulated error conditions. Comparisons to a culture standard for 104 subgingival plaque samples collected from 26 adult periodontitis patients yielded > 98% specificity for each of the test bacteria. In addition, the detection threshold for the test was determined to be equivalent to 10(4) cultivable test bacteria when compared to the culture standard. The data indicate that this membrane immunoassay is a valid and easy-to-use method for the detection of A. actinomycetemcomitans, P. gingivalis, and P. intermedia in subgingival plaque, at levels above the detection threshold of the test.

Colonization by Actinobacillus actinomycetemcomitans, Porphyromonas gingivalis and Prevotella intermedia in adult periodontitis patients as detected by the antibody-based Evalusite Test.

Studies were performed to evaluate the detection of disease-associated bacterial colonization in adult periodontitis patients by the antibody-based Evalusite TestTM (Eastman Kodak Company). The association of test results with disease was assessed by collecting 104 duplicate subgingival plaque samples from 26 patients. Samples were tested for Actinobacillus actinomycetemcomitans, Porphyromonas gingivalis and Prevotella intermedia using both microbiological culture and the immunoassay test. The sensitivity and specificity of the 2 methods was calculated using %s of positive results in deep periodontal pockets and negative results in shallow subgingival sites. A cutoff >10(4) cultivable counts yielded the greatest discrimination between health and disease on a cross-sectional basis and established this threshold as clinically relevant for the detection of disease-associated levels of bacterial colonization by these three microorganisms. The clinical detection limit of the immunoassay test was observed to coincide with this threshold of >10(4) cultivable counts. Microbiological testing of the 4 deepest pockets using the immunoassay test was determined to be sufficient to yield a 90% confidence of detecting positive patients in a study with 59 adult subjects. The immunoassay test method was also demonstrated to be effective at detecting bacterial colonization in sets of paper point samples that were pooled for analysis. An overall agreement of 94% (288 of 306) was observed when comparing test results for duplicate sets of pooled and individual samples collected from 51 patients. These studies demonstrate that the Evalusite Test is an effective method for detecting clinically relevant colonization by the test bacteria in patients at risk for periodontal disease.

Microbial alterations in supragingival dental plaque in response to a triclosan-containing dentifrice.

A total of 325 subjects were entered into a double-blind, stratified 2-treatment clinical study that examined the effects of a dentifrice containing 0.3% triclosan, 2% Gantrez copolymer and 0.243% sodium fluoride on supragingival dental plaque and gingivitis. A subset of 159 subjects including 72 men and 87 women participated in the microbiological component of this study, which was designed to detect shifts in supragingival bacterial species in response to triclosan. Subjects were divided into two groups: one performed normal oral hygiene with the triclosan/copolymer dentifrice and a control group used a placebo dentifrice without triclosan. At baseline, 3 and 6 months during treatment and at 6, 12, 18 and 24 weeks post-treatment, supragingival dental plaque was collected from the buccal and lingual surfaces of the 4 first molar teeth and assayed for: 1) bacterial morphotypes by phase-contrast microscopy; 2) select bacterial groups and bacterial species by culture; and 3) target periodontal pathogens including Actinobacillus actinomycetemcomitans, Bacteroides forsythus, Porphyromonas gingivalis and Prevotella intermedia by immunofluorescence microscopy. There were few statistically significant differences between treatment groups in any of the 90 microbiological parameters measured at the 7 different time points. The control group demonstrated significantly higher levels of cultivable Neisseria and higher proportions at the 3-month treatment period of P. gingivalis-infected subjects and mean cells. After 6 months of treatment, the triclosan group demonstrated higher levels of fusiforms. Analysis of triclosan resistance data failed to detect a shift towards increased proportions of bacteria resistant to triclosan, and both treatment groups demonstrated triclosan-resistant strains, predominantly Veillonella dispar. This study confirms the microbiological safety of triclosan-containing dentifrices and suggests that continued use can be associated with beneficial alterations in the bacterial composition of supragingival dental plaque.

Analysis of in vitro lymphoproliferative responses and antibody formation following subcutaneous injection of Actinobacillus actinomycetemcomitans and Wolinella recta in a murine model.

The potential of Wolinella recta and Actinobacillus actinomycetemcomitans to cause abscesses and induce an immune response was tested in BALB/c mice. Mice were injected subcutaneously with W. recta, A. actinomycetemcomitans or a mixture of these 2 microorganisms. Mice injected with A. actinomycetemcomitans alone, or with both organisms, demonstrated abscesses at the injection site 2 days later, from which pure cultures of A. actinomycetemcomitans were isolated. Mice injected with W. recta had small, flat abscesses at the injection site from which no bacteria could be cultured. W. recta was cultured from injection sites only when associated with A. actinomycetemcomitans. Mice developed positive serum IgG antibody responses to W. recta by 20 days post-injection but not to A. actinomycetemcomitans whether injected in pure culture or mixed infection. In vitro lymphoproliferative responses following injection of W. recta and/or A. actinomycetemcomitans resulted in increased lymphocyte reactivity in unstimulated cultures and decreased in vitro responses to phytohemagglutinin. In vitro lymphoproliferative responses to Escherichia coli LPS or Salmonella typhimurium LPS were depressed in mice injected with A. actinomycetemcomitans, but not in mice injected with W. recta.

Studies of the subgingival microflora in patients with acquired immunodeficiency syndrome.

Two unique forms of periodontal disease, HIV-gingivitis and HIV-periodontitis, have been described in patients with Acquired Immunodeficiency Syndrome (AIDS). In order to determine the bacterial species associated with periodontitis in AIDS patients, the predominant cultivable microflora was examined in 21 subgingival plaque samples from 11 AIDS patients with periodontitis. The presence of putative periodontal pathogens including Actinobacillus actinomycetemcomitans, Bacteroides intermedius, Porphyromonas gingivalis (formerly B. gingivalis), and Wolinella recta was examined by immunofluorescence in 128 subgingival dental plaque samples from 50 AIDS patients including 32 patients with periodontitis. Of 666 bacterial strains isolated from the 21 subgingival plaque samples, Streptococcus sanguis II was the most frequently recovered species comprising 18.5% of the total number of isolates followed by Lactobacillus acidophilus (12.2%), Porphyromonas gingivalis (12%), Fusobacterium nucleatum (11.4%), Staphylococcus epidermidis (8.7%), Actinomyces naeslundii (7.5%), and Actinomyces viscosus (4.7%). Fusobacterium nucleatum was the most prevalent species and was found in 76% of the sites and 91% of the patients. Enteric species including Enterococcus avium and Enterococcus faecalis, Clostridium clostridiiforme and Clostridium difficle as well as Klebsiella pneumoniae also were recovered. Immunofluorescence assays detected similar carriage rates of A. actinomycetemcomitans, B. intermedius, and P. gingivalis in both gingivitis patients and periodontitis patients, while four times more periodontitis patients demonstrated W. recta. Subgingival yeast was a frequent finding in these AIDS patients, present in 62% of the subjects and 55% of the sites. This study indicates that subgingival plaque in AIDS patients with periodontitis can harbor high proportions of the same periodontal pathogens as are associated with periodontitis in non-HIV infected subjects as well as high proportions of opportunistic pathogens.

Effect of a triclosan/copolymer/fluoride dentifrice on the oral microflora.

Eighty-one human subjects completed a double-blind study which examined the effects of a 0.3% triclosan/2% Gantrez copolymer/0.243% sodium fluoride dentifrice on the microflora of supragingival dental plaque. Subjects were divided into an experimental group which performed normal oral hygiene with the triclosan/copolymer/fluoride dentifrice and a control group which also performed normal oral hygiene with the same dentifrice minus the triclosan/copolymer. At baseline, 10 weeks, and 28 weeks, supragingival dental plaque was collected from buccal and lingual surfaces of the four first molar teeth and assayed for: 1) bacterial morphotypes by phase contrast microscopy, 2) Actinobacillus actinomycetemcomitans, Actinomyces species, Bacteroides forsythus, Bacteroides gingivalis, Bacteroides intermedius, Streptococcus mutans, Streptococcus sanguis, and Wolinella recta by immunofluorescence microscopy, and 3) Lactobacillus, yeast, enterics, Staphylococcus, aerobes and anaerobes by bacterial culture. After 28 weeks' use of their respective dentifrices, changes in the supragingival plaque microflora of the subjects were similar between the triclosan/copolymer/fluoride dentifrice group and the control dentifrice group, except for statistically significant reductions in fusiforms, spirochetes and staphylococci and significant increases in S. sanguis in the triclosan/copolymer/fluoride dentifrice group, as compared to the control dentifrice group. The subject population was unusual in the presence of enteric species and anaerobes found in supragingival plaque sites. This study indicates that the use of a dentifrice containing 0.3% triclosan and 2% Gantrez copolymer over an extended period of time (28 weeks) does not result in shifts in the microflora of supragingival plaque favoring the growth of either opportunistic or pathogenic bacterial species.

Eikenella corrodens in the human oral cavity.

The prevalence and distribution of the putative periodontal pathogen Eikenella corrodens in the human oral cavity was examined. A total of 508 oral bacterial samples were taken from 10 periodontally healthy adults (PH), 11 adult periodontitis patients (AP), and 6 localized juvenile periodontitis patients (LJP). From each subject, samples of supra- and subgingival plaque were obtained from six to eight teeth as well as samples from buccal mucosa, lateral and dorsal surfaces of tongue, tonsil, and saliva. E. corrodens was cultured from 70% of healthy subjects and 100% of periodontitis patients. Dental plaque appears to be the main oral ecological niche of E. corrodens in PH subjects since it was found in, respectively, 26% and 31% of supra- and subgingival plaque samples and rarely found in other oral sites in these subjects. It was found in 59% of both supra- and subgingival plaque samples from AP subjects, as well as 48% and 64% of supra- and subgingival plaque samples of LJP subjects. In contrast to healthy subjects, E. corrodens was found on the buccal mucosa, tongue, tonsil and in the saliva of patients with periodontitis. The microorganism constituted, on average, 1% to 2% of the total cultivable bacteria in supra- and subgingival plaque samples. The prevalence of E. corrodens in plaque samples was higher in AP and LJP subjects and was significantly different from PH subjects. Within the AP group, the prevalence of E. corrodens in subgingival plaque is significantly higher from sites with GI greater than or equal to 2. These data suggest that E. corrodens is an indigenous oral microorganism which may be an opportunistic pathogen associated with gingival inflammation.

Effect of a chemotherapeutic agent delivered by an oral irrigation device on plaque, gingivitis, and subgingival microflora.

Sixty-six adults were examined in a double-bind study which examined the effect of an antimicrobial agent delivered by an oral irrigating device. Each subject received a randomized half mouth dental prophylaxis. The Gingival Index, gingival crevicular fluid volume, Plaque Index, Modified Papillary Bleeding Index, probing pocket depth, and probing attachment levels were determined at baseline, 3 weeks, and 6 weeks. The composition of the subgingival microflora in the prophied and non-prophied quadrants was examined by phase contrast microscopy and by immunofluorescence. This study demonstrates that an antimicrobial product delivered by an oral irrigating device could result in significant reductions in plaque, bacterial cell counts, and gingival bleeding and may, therefore, be an effective adjunct to normal oral hygiene.

Heterogeneity of virulence among strains of Bacteroides gingivalis.

The ability of fresh isolates of B. gingivalis to establish abscesses in the mouse model was studied by comparing them with established laboratory strains of B. gingivalis. Eight fresh isolates obtained from plaque associated with periodontal disease and grown under similar conditions as established strains were injected subcutaneously on the back of the mouse. All of these strains produced secondary lesions on the abdomen. Septicemia was associated with seven of the strains. Two commonly used laboratory strains, W50 and W83, also produced secondary lesions and septicemia. Five other laboratory strains produced only localized abscesses. On histologic examination, the strains that produced disseminated disease showed invasion of connective disease by individual bacteria that were not in clumps. The strains that produced localized abscesses were characterized by growing in colonies or clumps in the abscess cavity. Four synthetic enzyme substrates were examined to determine whether the differences between invasive and non-invasive strains were due to differences in proteolytic enzyme production. No differences in enzyme production could be demonstrated with the selected substrates.

Effects of subgingival irrigation on A. actinomycetemcomitans.

The effects of repeated subgingival irrigation on Actinobacillus actinomycetemcomitans was examined. 24 periodontal pockets harboring A. actinomycetemcomitans in 3 juvenile and 4 adult periodontitis patients were studied. The protocol included bi-weekly subgingival irrigation with hydrogen peroxide of the periodontal sites until the micro-organism was no longer detected by selective culture, or for 6 months. A. actinomycetemcomitans was gradually suppressed to below detection following the irrigation regime and could no longer be detected in 46% of the sites at completion of the irrigation protocol. The sites were microbiologically re-examined 5 months after cessation of the irrigation regime. A. actinomycetemcomitans re-occurred in only 2 of the sites from which it had originally been suppressed below detection. The results indicate: (1) that the irrigation regime tested has some potential to suppress A. actinomycetemcomitans in periodontal pockets; (2) that the effect of the irrigation protocol generally lasted for 5 months; (3) that the reduction rate of A. actinomycetemcomitans to below detectable levels seems related to the initial number of cultivable bacteria from the periodontal pocket.

Serological studies of oral Bacteroides intermedius.

Bacteroides intermedius is a gram negative, anaerobic microorganism associated with certain forms of human periodontal disease, including adult periodontitis and acute necrotizing ulcerative gingivitis. Previous studies have indicated the presence of two DNA homology groups which could be distinguished by analysis of protein patterns on polyacrylamide gel electrophoresis, as well as at least two serogroups within B. intermedius. The present study examined the serology of B. intermedius and determined the distribution of B. intermedius serogroups in clinical isolates and patient plaque samples. Serological reactions with unabsorbed rabbit antisera and antisera immunoabsorbed with B. intermedius strains demonstrated a previously unreported antigenic group within B. intermedius, serogroup C, in both immunodiffusion and immunofluorescence assays. Of 79 B. intermedius isolates from 68 subjects examined with specific antisera, 55% of the isolates and 52% of the subjects were categorized in serogroup C, 40% of the isolates and 46% of the subjects were in serogroup B, and 5% of the isolates and 6% of the subjects were in serogroup A. In 31 samples of subgingival dental plaque from adolescents known to harbor B. intermedius, 81% demonstrated serogroup B, 16% had serogroup A, and 3% had serogroup C.

Effect of immunization on experimental Bacteroides gingivalis infection in a murine model.

BALB/c mice were immunized with an invasive (A7A1-28) or noninvasive (381) Bacteroides gingivalis strain, Bacteroides intermedius, or Ringer solution. All immunized mice were subsequently challenged with the invasive B. gingivalis strain and examined for septicemia or secondary spread of the infection or both. Mice immunized with the invasive B. gingivalis strain localized the infection to the challenge site. Mice immunized with the noninvasive B. gingivalis strain, B. intermedius, or Ringer solution developed spreading infections. These data suggest that immunization with an invasive B. gingivalis strain can alter the course of subsequent infections.

Effect of anaerobiosis on the surface ultrastructure and surface proteins of Actinobacillus actinomycetemcomitans (Haemophilus actinomycetemcomitans).

The ultrastructures and surface protein profiles of aerobically cultured Actinobacillus actinomycetemcomitans (Haemophilus actinomycetemcomitans) differed from those of cells cultured anaerobically. Similar ultrastructural differences were also observed when aerobic and anaerobic cultures of a strain of Escherichia coli were compared. These results suggest that oxygen-related variations in the bacterial cell surface may play a role in the adaptation of oral bacteria to different host environments.

The use of monoclonal antibodies to detect Bacteroides gingivalis in biological samples.

Hybridomas were established which produce monoclonal antibodies specific for Bacteroides gingivalis, a pathogen associated with human periodontal disease. Spleen cells from BALB/c mice immunized with formalinized B. gingivalis were fused with Sp2/0-Ag14 myeloma cells. Of 1,050 wells with positive growth, 60 contained antibody reactive with the immunizing strain of B. gingivalis by enzyme-linked immunosorbent assay. Expansion of these cultures and cloning by limited dilution resulted in 28 clones which reacted with B. gingivalis but not with other orals and nonoral black-pigmented Bacteroides species or any of 29 representative strains of other oral bacteria. Of these 28 clones, 14 were also specific for B. gingivalis by indirect immunofluorescence microscopy. One clone, BBG-12 producing immunoglobulin G2b(kappa), was chosen to identify B. gingivalis in subgingival plaque because of its high reactivity in indirect immunofluorescence assays. This antibody reacted strongly with all 17 representative B. gingivalis strains obtained from diverse sources. Furthermore, when this reagent was applied to subgingival plaque samples, B. gingivalis was stained with high specificity and low background fluorescence, indicating that it may be useful for clinical identification of this organism.

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% 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%.(ABSTRACT TRUNCATED AT 250 WORDS)

Lactic acid production by oral Streptococcus mitis inhibits the growth of oral Capnocytophaga.

Various relationships including inhibition or stimulation of growth have been demonstrated among the bacteria present in dental plaque, both in vitro and in vivo. A large number of these relationships involved oral Streptococci. An earlier study found that strains of Streptococcus mitis inhibited the growth of potential periodontopathic microorganisms, such as Actinobacillus actinomycetemcomitans, Capnocytophaga and species of Bacteroides and Fusobacterium. The present investigation showed that this inhibitory effect resulted primarily from lactic acid production.

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.