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Germ-free rats were monoinfected with Porphyromonas gingivalis strains 381 or A7A1-28 for 42 or 84 days. Both strains induced substantial destruction of alveolar bone and soft tissue when compared to non-infected controls, but the patterns were different. Strain A7A1-28 was associated with increased activity of host collagenase and gelatinase at 42 days, whereas the activity was elevated to a lesser extent at 84 days. Strain 381 showed a moderate increase in host proteinase activity at 42 days, and this remained unchanged until day 84. Strain A7A1-28 was associated with more bone loss than strain 381 by a morphometric analysis that detects horizontal bone loss in the maxilla. Strain 381 was associated with more bone loss than strain A7A1-28 by a radiographic method that detects vertical intrabony defects in the mandible. Infection with one strain gave rise to serum and salivary antibodies strongly reactive to the infecting strain and moderately reactive to antigens from the other strain. This indicates that some antigenic similarity exists between the strains and that there are also strain or perhaps serotype differences in antibody responses induced by infection. Thus two strains of P. gingivalis differing in antigenicity and pathogenicity in the mouse model of the subcutaneous abscess cause substantial periodontal destruction in the germ-free rat. The disease pattern is, however, different, with strain A7A1-28 inducing mostly horizontal bone loss and strain 381 mostly vertical.

Adhesive fimbriae from Porphyromonas gingivalis are cell surface structures which may be important in the virulence of this oral pathogen and thus may serve as a critical or target antigen. Immunization with highly purified 43-kDa fimbrial protein protected against periodontal tissue destruction when tested in the P. gingivalis-infected gnotobiotic rat model. A similarly highly purified 75-kDa cell surface component did not provide protection. Heat-killed whole-cell and sonicated cell surface extracts which contain the 43-kDa protein as well as the 75-kDa component were protective also. This study indicates that the fimbrial protein may serve as a model for the development of effective vaccines against periodontitis, a major human oral disease.

The mitogenic, chemotactic, and synthetic responses of rat periodontal ligament (PDL) fibroblastic cells to epidermal growth factor (EGF), transforming growth factor-beta (TGF-beta), recombinant human platelet-derived growth factor (rhPDGF)-AB, rhPDGF-BB, natural (n) PDGF-AB, and insulin-like growth factor-I (IGF-I) were examined in vitro using PDL cells obtained from the coagulum of healing tooth sockets. PDGFs and IGF-I have potent and comparable mitogenic effects on PDL fibroblastic cells. The maximum mitogenic effect of PDGFs was observed at the concentration of 10 ng/ml, whereas that of IGF-I was seen at concentrations higher than 100 ng/ml. In contrast, EGF induced moderate, and TGF-beta inhibitory mitogenic responses. The combination of rhPDGF-AB with either EGF or TGF-beta demonstrated comparable mitogenic potency, equivalent to the level of PDGF alone regardless of the mitogenic effect of other growth factors. The combination of rhPDGF-AB and IGF-I, however, showed a synergistic effect revealing the highest mitogenic effect among all individual growth factors as well as any combinations of the growth factors tested. Similarly, PDL fibroblastic cells demonstrated strong chemotactic responses to both IGF-I and PDGFs. The maximum effect was observed by IGF-I at concentrations higher than 10 ng/ml, followed by rhPDGF-BB at 0.1 ng/ml, rhPDGF-AB and nPDGF at concentrations ranging from 0.1 to 1 ng/ml. TGF-beta revealed no, and EGF slightly increased, chemotactic effects. IGF-I slightly enhanced the synthesis of total protein, whereas other factors had no significant effect. However, both rhPDGF-AB and TGF-beta stimulated collagen synthesis. On the other hand, IGF-I showed no effect on collagen synthesis, while EGF suppressed collagen synthesis. These findings suggest that rhPDGF-BB and IGF-I stimulate proliferation and chemotaxis of PDL fibroblastic cells. In addition, the combination of these growth factors further increases the mitogenic effect. rhPDGF-AB also stimulates collagen synthesis by PDL fibroblastic cells. Thus, rhPDGF-BB and IGF-I may have important roles in promotion of PDL healing, and consequently, may be useful for clinical application in periodontal regenerative procedures.

Fimbriae are important in the adherence of many bacterial species to the surfaces they eventually colonize. Porphyromonas (Bacteroides) gingivalis fimbriae appear to mediate adherence to oral epithelial cells and the pellicle-coated tooth surface. The role and contribution of fimbriae in the binding of P. gingivalis to hydroxyapatite (HAP) coated with saliva as a model for the pellicle-coated tooth surface were investigated. 3H-labeled P. gingivalis or the radioiodinated purified fimbriae were incubated with 2 mg of HAP beads coated with whole human saliva (sHAP) and layered on 100% Percoll to separate unbound from sHAP-bound components. The radioactivity of the washed beads was a measure of the bound components. The binding of P. gingivalis 2561 (381) cells and that of purified fimbriae were concentration dependent and saturable at approximately 10(8) cells and 40 micrograms of fimbriae added, respectively. The addition of fimbriae inhibited binding of P. gingivalis to sHAP beads by 65%, while the 75-kDa protein, which is another major surface component of P. gingivalis 2561, did not show significant inhibition, suggesting that the fimbriae are important in adherence. Encapsulated and sparsely fimbriated P. gingivalis W50 did not bind to sHAP beads. On the basis of the predicted sequence of the fimbrillin, a structural subunit of fimbriae, a series of peptides were synthesized and used to localize the active fimbrillin domains involved in P. gingivalis adherence to sHAP beads. Peptides from the carboxyl-terminal one-third of the fimbrillin strongly inhibited P. gingivalis binding to sHAP beads. Active residues within the sequence of inhibitory peptide 226-245 (peptide containing residues 226 to 245) and peptide 293-306 were identified by using smaller fragments prepared either by trypsin cleavage of the peptide 226-245 or by synthesis of smaller segments of peptide 293-306. Hemagglutinin activity, lectinlike binding, and ionic interaction did not seem to be involved in this binding since lysine, arginine, carbohydrates, and calcium ions failed to affect the binding of P. gingivalis. The observation that poly-L-lysine, bovine serum albumin, and defatted bovine serum albumin, even at high concentrations, only partially blocked the binding of P. gingivalis indicates that hydrophobic interactions are not the major forces involved in P. gingivalis binding to sHAP beads. Protease inhibitors such as EDTA, leupeptin, pepstatin, 1,10-phenanthroline, and phenylmethylsulfonyl fluoride did not interfere with the binding of P. gingivalis. However, the binding of P. gingivalis to trypsin- or chymotrypsin-pretreated sHAP beads was reduced.(ABSTRACT TRUNCATED AT 400 WORDS)

In periodontal diseases, bacteria trigger inflammatory host responses which, along with the direct destructive effects of the bacteria, cause most of the tissue destruction. Periodontal inflammatory responses are, by and large, immunologic, and our understanding of these reactions has been advanced by the explosion of knowledge in immunobiology, some of which is discussed in this review. Understanding the role of immune cells and their regulatory cell surface molecules such as the MHC, CD antigens, and receptors, as well as knowledge of effector systems set into motion such as phagocytes and cytotoxic T-cells, and the effector molecules such as antibodies, complement, and cytokines, have led to better understanding of the complex pathogenesis of periodontal disease. The role of mediators including the matrix metalloproteinases, proteoglycans, the kinins and anaphylatoxins, and low molecular weight mediators including products of arachidonic metabolism is beginning to be elucidated in periodontal disease. Important avenues of research for development of diagnostic tests based upon host response are apparent. For example, tissue products released during periodontal inflammation including the metalloproteinases, elastase, cytokines, prostaglandins, antibodies, and complement components may provide the basis for future diagnostic indicator tests. The recognition that the neutrophil/antibody/complement axis is critical for protection against periodontal bacteria and that abnormalities in this system often lead to increased periodontal susceptibility provide approaches for the development of diagnostic tests assessing risk. A group of factors which are negative regulators of inflammation including TGF-β, gamma-interferon, and IL-1 receptor antagonist provide potential for assessment of periodontal disease in remission or in the healing phase. Finally, factors such as HLA associations and the molecular basis for neutrophil abnormalities may provide genetic markers for periodontal disease susceptibility. Diagnostic factors based upon host response measures offer great potential for predicting host susceptibility and will likely be used in combination with microbial diagnostics which identify specific infecting organisms. J Periodontol 1992;63:338-355.

The objective of The Present study was to define criteria for the diagnosis of "established periodontitis." This term will define subjects who have demonstrated clinical attachment loss, and as such can be considered to have periodontitis. Using these criteria, healthy and established periodontitis subjects were compared with respect to gender, race, and age. Five hundred and eight subjects including 248 females and 260 males between the ages of 25 to 73 (mean 44.6 years), were examined in this study. The clinical examination included: plaque assessment index (PAI); gingival assessment index (GAI); probing pocket depth (PPD); and clinical attachment level (CAL). The mean and frequency distribution of these parameters were analyzed by age and gender. CAL (mean 2.12 mm) showed constant and significant increases with age, ranging from a mean of 1.63 mm in subjects 25 to 34 years of age to a mean of 2.65 mm in subjects 65 to 74 years of age. Males exhibited higher mean values than females for all the measured parameters, which were statistically significant for PAI, PPD, and CAL. The frequency distribution of subjects with PPD and CAL beyond certain threshold levels showed an exponential decline and was correlated to both the severity of the most involved site as well as the number of sites beyond threshold levels. The clinical entity of "established periodontitis" is suggested based on the presence of CAL greater than or equal to 6 mm in 2 or more teeth and one or more sites with PPD greater than or equal to 5 mm. In the present study, 30.5% of the subjects fell into this category.(ABSTRACT TRUNCATED AT 250 WORDS)

Previous studies have suggested that infections with Porphyromonas gingivalis, associated with periodontal disease, may consist of one clonal type. It has also been shown that each individual patient carries a unique clonal type of P. gingivalis, as assessed by DNA fingerprinting. This issue was further examined by random collection of multiple isolates ofP. gingivalis from multiple sites in several patients, and characterization of these isolates by DNA fingerprinting. Although most patients appeared to be infected exclusively by one clonal type of P. gingivalis, at least one patient was found to harbor two distinct clonal types. This indicates that the simultaneous presence of different clonal types of P. gingivalis can occur. A statistical method was developed for retrospective analysis of these data for estimation of whether the remainder of these patients were actually infected with single or multiple clonal types ofP. gingivalis. With this statistical method, a confidence interval was calculated for estimation of the true proportion of a single observed clonal type in the potential population of P. gingivalis that might be recovered from an infected patient. Statistically, the sampling of small numbers of sites per patient or isolates per site leads to a wide confidence interval for the estimated true proportion of the observed clonal type in the infecting P. gingivalis population. For example, when five sites in an oral cavity yield indistinguishable P. gingivalis isolates, then the true proportion of this clonal type in the total P. gingivalis population in the infected oral cavity is estimated to be in the interval between 55% and 100% (at a 95% confidence level). If the isolates from all sampled sites from a periodontal patient appear to be of the same clonal type, it can be calculated that at least 29 different sites must be sampled for this observed clonal type to represent at least 90% of the P. gingivalis population in this patient (95% confidence level); this was termed the LTP, or lowest true proportion. Such calculations can also be used for estimation of the LTP of a clonal type infecting a single oral site in a periodontal patient. These results confirm the commonly held opinion that studies investigating the ecology of P. gingivalis and other suspected pathogens must be designed on a large scale in order for statistically meaningful results to be obtained.

The prevalence of people and sites with attachment loss, pocket depth, Actinobacillus actinomycetemcomitans, Prevotella intermedia, and Porphyromonas gingivalis are described for a random sample of 366 black and 297 white community-dwelling adults, aged 65 or over, residing in five counties in North Carolina. In addition, relationships between sites harboring these microorganisms and loss of attachment (LA) and pocket depth (PD) are presented in a manner that considers the lack of independence of sites within each person. Pocket depths and recession were measured on all teeth by trained examiners during household visits. Immunofluorescent assays for A. actinomycetecomitans, P. intermedia, and P. gingivalis were conducted on subgingival plaque samples obtained from the mesiobuccal aspect of the four first molar teeth using paper points. The prevalences of A. actinomycetemcomitans, P. intermedia, and P. gingivalis were greater in blacks than in whites. The most striking difference was seen for P. gingivalis, which was found in 38.8% of blacks and 9.4% of whites. Similar relationships were found when the percent of sites with these organisms were assessed. Blacks with P. gingivalis or P. intermedia had a higher prevalence of sites with LA greater than or equal to 7 mm as compared to blacks not infected with P. gingivalis or P. intermedia. The same was true for whites. Similar relationships between P. gingivalis or P. intermedia and PD greater than or equal to 6 mm were found for both blacks and whites.(ABSTRACT TRUNCATED AT 250 WORDS)

In these studies we have attempted to show that cell surface structures are critical antigens for protection against P. gingivalis-induced periodontal destruction. Fimbrillin, and in particular a synthetic 20-amino-acid fimbrillin peptide, exerts a protective effect in gnotobiotic rats, thus identifying them as potentially useful in the development of a vaccine.

Recent studies of microbial epidemiology emphasizing the genetic organization and distribution of organisms associated with orofacial infections have led to new insights into the possible origins of pathogenicity. Studies into genetic heterogeneity, acquisition and transmission of these organisms have been markedly advanced by the utilization of the powerful technique of genomic DNA fingerprinting. Characteristic fingerprints for each bacterial isolate can be produced by cleavage of high molecular weight genomic DNA by restriction endonucleases. It is assumed that each DNA fingerprint represents a clonal type. In this report, we review and analyze studies of the epidemiology of bacteria associated with orofacial infections with an emphasis on periodontal disease. Studies of nontypable (NT) Haemophilus influenzae associated with recurrent otitis media illustrate the utility of this technique. DNA fingerprinting clearly demonstrates genetic heterogeneity of NT H. influenzae isolates, and clonality of infection of any individual. Furthermore, DNA fingerprinting has shown that the same clonal type is seen in siblings concurrently suffering from otitis media, suggesting horizontal transmission within the family. Studies of mutans Streptococci also show extensive genetic heterogeneity and show vertical transmission of a predominant clonal type only from mother to infant, but not from father to infant. Studies of Actinobacillus actinomycetemcomitans show considerable genetic heterogeneity among monkey isolates. Thus far, three clonal types have been reported with DNA fingerprinting among isolates from periodontal patients, but additional genetic heterogeneity can be found using specific DNA fragments as probes in hybridization experiments. Intrafamilial transmission of A. actinomycetemcomitans has been demonstrated. Porphyromonas (Bacteroides) gingivalis shows extensive genetic heterogeneity and case reports suggest clonal infection of any one individual. In contrast, results with DNA fingerprinting of Eikenella corrodens, Fusobacterium nucleatum, and Bacteroides intermedius show that individuals may be infected with 2 or more clonal types. These studies point to the great potential of DNA fingerprinting for investigating the epidemiology of putative orofacial pathogens. Such studies with periodontal microorganisms will likely reveal steps in the acquisition, intraoral and person-to-person transmission, which then could possibly be inhibited or interfered with to prevent periodontal disease or its recurrence.

Rapid and reproducible method is described here for the purification of the 43 kDa fimbrial protein from P. gingivalis by preferential fractionation in the presence of 1% SDS and 0.2M of a bivalent cation at pH 6.5. Homogeneity of the purified 43 kDa was confirmed by SDS-PAGE and Western blot analysis using monoclonal and polyclonal antibodies raised against this protein. Amino acid composition and the amino acid sequence of the first 30 amino acid residues of the purified fimbriae are consistent with the composition and sequence predicted from the cloned gene of the fimbrial subunit. Circular dichroism spectra shows high levels of beta-sheet structure. The purified 43 kDa polymer shows fimbriae-like morphology under the electron microscope. Ultrastructural localization of the 43 kDa protein by the immunogold technique revealed specific labeling of the fimbriae with a diameter of approximately 3.5 to 5.0 nm. Localization of this protein suggest that the 43 kDa component is a fimbrial subunit.

The neutrophil (PMN) receptor for formylated peptides such as N-formyl-l-methionyl-l-leucyl-l-phenylalanine (FMLP) is involved in binding and subsequent response to certain chemotactic stimuli. The receptor on human PMN has been reported to consist of several glycoprotein components, ranging in size from 43-94 kDa. Furthermore, FMLP receptors on human PMN have been shown to contain both high and low affinity states. In this study, the receptor was purified by subjecting solubilized PMN plasma membrane components to FMLP-affinity chromatography, and was found to be comprised of four components, one of 68 kDa, and the others of 94, 48, and approximately 40 kDa. Only the 68, the 94, and the approximately 40 kDa components specifically bound a radioiodinated FMLP analogue. To further characterize these components, a battery of monoclonal antibodies reactive against the FMLP receptor was prepared. Seven monoclonal antibodies were selected on the basis of their reactivity with the 68 kDa receptor component. Some of these antibodies also cross-react with the 48 kDa component, suggesting that the 68 and the 48 kDa receptor moieties are immunologically related. These antibodies reacted with normal human neutrophils, but not with lymphocytes, or unstimulated HL-60 cells. Furthermore, the presence of 20 nmol of FMLP inhibited the binding of five of the anti-receptor antibodies to whole PMN. These results suggest that the epitopes recognized by these five antibodies may possibly be involved in FMLP binding.

Bacterial fimbriae mediate cell adhesion and are important in colonization. Fimbrial proteins from strains of Porphyromonas (Bacteroides) gingivalis isolated from different individuals were compared for their size, amino-terminal sequence, and antigenic diversity. Two major protein components of the crude fimbrial preparations differed in apparent molecular mass, ranging from 41 to 49 kDa for the fimbrillin monomer and from 61 to 78 kDa for the other major protein. The amino-terminal sequence of the antigenically related group of proteins of the fimbrillin monomer in the 41- to 49-kDa range showed significant homology; however, minor sequence heterogeneity was observed, mainly in residues 4 to 6. One of the observed amino-terminal sequences, AFGVGDDESKVAKLTVMVYNG, resembled the deduced sequence of P. gingivalis 381 (D.P. Dickinson, M. K. Kubiniec, F. Yoshimura, and R.J. Genco, J. Bacteriol. 170:1658-1665, 1988). Fimbriae from all the strains of P. gingivalis showing this sequence contained a fimbrillin monomer of 43 kDa and showed a strong reaction with both polyclonal and monoclonal antibodies directed to the fimbriae from P. gingivalis 2561 (381). Fimbriae from strains showing amino-terminal sequence variations in residues 4 to 6 (i.e., substitution of VGD with either E or NAG) were more diverse in their molecular sizes. Most of these variant fimbriae showed weak reactions with the polyclonal antibodies and no reaction with the monoclonal antibodies induced to the fimbriae of strain 2561. No correlation could be established between the molecular size and immunological reactivity of the fimbrillin monomer of P. gingivalis strains. Strains 9-14K-1 and HG 564 not only showed markedly different sequences from the other three amino-terminal sequences but also did not react with either polyclonal or monoclonal antibodies to the fimbriae of strain 2561. Strains W50, W83, and AJW 5 failed to show any immunological reactivity with the antibodies to fimbrillin or fimbriae of strain 2561. Fimbriae from different strains revealed different immunologic reactions with rabbit antisera to each of the synthetic peptides of residues 59-78 (peptide I), 79-100 (peptide J), and 91-108 (peptide E) of strain 381. These results suggest that P. gingivalis fimbrillin subunits have size, sequence, and antigenic heterogeneity among the strains and that these differences may be important in the function and immune reactivities of the fimbriae.

The relationship between type 2 diabetes mellitus and periodontal disease was evaluated in 2,878 Pima Indians of the southwestern United States. Two independent measures of periodontal disease, probing attachment loss and radiographic bone loss, were used to compare prevalence and severity of periodontal disease in diabetic and nondiabetic subjects. In all age groups studied, subjects with diabetes had a higher prevalence of periodontal disease, indicating that diabetes may be a risk factor for periodontal disease.

This study describes the use of total genomic DNA fingerprinting with the use of restriction endonucleases to characterize clinical isolates of Porphyromonas gingivalis (Bacteroides gingivalis) obtained from patients with periodontitis or with root-canal infections. The majority of independent isolates had a unique DNA fingerprint, indicating extensive genetic heterogeneity within this species. Twenty-nine distinct DNA fingerprints were found among the 33 isolates investigated. This is in contrast to biotyping and serotyping, where only one type and three types, respectively, have been reported. The observed heterogeneity indicates that DNA fingerprinting is a sensitive measure of genetic dissimilarity between P. gingivalis isolates and is able to characterize individual isolates. These results have ecological implications, indicating that there is considerable natural diversity in the global population of P. gingivalis, and that there are likely to be relatively large numbers of genetically distinct clonal lines. Furthermore, DNA fingerprinting is a sensitive and powerful tool for longitudinal and cross-sectional epidemiological studies. This technique provides far greater discrimination between isolates than either biotyping or serotyping, and will be most helpful in, for example, the analysis of distribution of clonal lines within one periodontal patient, or the analysis of the transmission to and turnover of strain populations within a patient population, since the probability of two strains with the same DNA fingerprint being found by chance is small.

The goal of this study was to determine the prevalence and incidence of periodontal disease and its relationship with non-insulin-dependent diabetes mellitus (NIDDM). Two thousand two hundred seventy-three Pima Indians (949 men, 1324 women) aged greater than or equal to 15 yr from the Gila River Indian Community in Arizona were examined between 1983 and 1989. Periodontal disease was diagnosed by tooth loss and by percentage of interproximal crestal alveolar bone loss ascertained from panoramic radiography. Subjects with little or no evidence of periodontal disease were classified as nondiseased. Thus, the incidence of advanced periodontal disease was determined. The age- and sex-adjusted prevalence of periodontal disease at first dental examination was 60% in subjects with NIDDM and 36% in those without. Twenty-two new cases developed in a subset of 701 subjects (272 men, 429 women) aged 15-54 yr who initially had little or no evidence of periodontal disease and had at least one additional dental examination. The incidence of periodontal disease in this group was similar in men and women (incidence-rate ratio 1.0, 95% confidence interval [Cl] 0.5-1.9, controlled for age and diabetes). Higher age predicted a greater incidence of periodontal disease (chi 2 = 30.6, df = 3, P less than 0.001, controlled for sex and diabetes). The rate of periodontal disease in subjects with diabetes was 2.6 times (95% Cl 1.0-6.6, controlled for age and sex) that observed in those without. Although periodontal disease was common in nondiabetic Pima Indians, in whom most of the incident cases occurred, diabetes clearly conferred a substantially increased risk. Thus, periodontal disease should be considered a nonspecific complication of NIDDM.

In order to study the role of membrane proteins in mucosal colonization by Actinobacillus actinomycetemcomitans, a plasmid library was constructed by ligating EcoRI-digested genomic DNA from A. actinomycetemcomitans strain Y4 into EcoRI-digested and dephosphorylated pUC13 DNA. A second library was constructed by ligating Sau3A-digested and size-fractionated A. actinomycetemcomitans strain Y4 DNA into BamHI-cleaved and dephosphorylated pUC13 DNA. The DNA was transformed into Escherichia coli strain MC1022 and plated onto Luria-Bertani agar containing 100 micrograms/ml ampicillin and X-gal. Recombinant colonies were examined for the expression of A. actinomycetemcomitans antigens by colony immunoscreening using rabbit antiserum to strain Y4. Nine positive clones were isolated. Three of these clones produced proteins that reacted with rabbit antiserum to strain Y4 on a Western transfer, and seven of these clones reacted with rabbit antiserum to strain Y4 as measured by indirect immunofluorescence. One reactive clone contained an 8.4 kb plasmid, which directed the synthesis of a peptide with an Mr of 20,000 as measured by SDS-PAGE. Partial DNA sequence analysis revealed the presence of a signal-peptide leader sequence at the amino terminus suggesting that the native protein is a membrane component of A. actinomycetemcomitans. Thus A. actinomycetemcomitans antigens can be expressed in E. coli, and some of these proteins may be expressed on the E. coli cell surface.

P33 Epidemiological studies have demonstrated a significant link between periodontal disease (PD) and increased risk of cardiovascular disease (CVD). This study was conducted to evaluate if lipid metabolism and peroxidation may represent potential pathways linking PD to CVD. The study sample included 747 randomly selected residents of Erie and Niagara counties of New York who had 6 or more teeth and no history of heart attack. Periodontal assessments included pocket depth (POD) and attachment loss (ATL). Periodontitis was defined by 2+ teeth with ATL≥6mm and 1+ teeth with POD≥5mm. Subgingival plaque was sampled and tested using immunofluorescence microscopic technique for the presence of four putative periodontal pathogens: Bacteriodes forsythus (BF), Porphyromouas gingivilis (PG), Prevotella intermedia (PI), and Campyglobacter recta (CR). Fasting blood sample was obtained and determinations of total and LDL cholesterol, triglycerides, and plasma thiobarbituric acid reacting substance (TBARS) were performed. In the sample as a whole, prevalence of specific bacteria in the subgingival plaque were 29.6% for BF, 8.3% for PG, 31.6% for PI, and 4.4% for CR. As expected, prevalence was much higher in participants with periodontitis. Presence of BF and CR (but not PI and PG) was associated significantly with higher levels of LDL cholesterol (Mean LDL cholesterol (mg/dl), 151.3 for BF + and 142.8 for BF -, p

P34 Epidemiological studies have demonstrated a significant association between periodontal disease and increased risk of cardiovascular disease. Periodontal organisms were hypothesized to be the underlying factor for the association. This study investigated the relation of myocardial infarction (MI) with both periodontal health status and presence of periodontal pathogens in subgingival plaque. The study sample included 171 MI cases and 574 controls who had at least 6 teeth and were residents of Erie and Niagara counties of New York. Periodontal assessments included pocket depth and attachment loss. Subgingival plaque was sampled and tested using immunofluorescence microscopic technique for the presence of Bacteriodes forsythus (BF), Porphyromouas gingivilis (PG), Prevotella intermedia (PI), and Campyglobacter recta (CR). Results: Mean (SD) pocket depth (mm) was 2.2 (0.7) for MI cases and 2.0 (0.5) for controls (p

17 antibiotics, with potential for topical use, were tested for their activity against the human oral flora. Concentrations (mumol/l) required to inhibit 90% of test strains are presented and drug activities are compared. The total cultivable oral flora was susceptible to the tetracyclines including tetracycline itself, minocycline, doxycycline, and oxytetracycline and to erythromycin. On the other hand, actinobolin, kanamycin, neomycin, streptomycin, spiramycin, tyrothricin, vancomycin, clindamycin, and chloramphenicol were ineffective against many of the human oral anaerobic bacteria even at high concentration. Penicillin was effective at high concentrations but could not be recommended because organisms which are not inhibited by low concentrations are penicillinase producers. Carbenicillin was effective against all organisms except Actinobacillus actinomycetemcomitans. The gram-negative organisms involved in adult periodontitis were most susceptible to the tetracyclines, tyrothricin, carbenicillin and clindamycin, while those associated with localized juvenile periodontitis were susceptible to the tetracyclines or erythromycin. These data, combined with the previous findings that some tetracyclines exhibit marked substantivity and collagenase inhibition activity, indicate that tetracycline or minocycline are likely to be good choices in the treatment or prevention of oral diseases.

An actinomycetemcomitans can cause localized juvenile periodontitis and certain types of adult periodontitis. Optimal treatment of periodontal disease caused by this microorganism requires systemic antibiotic therapy in addition to mechanical debridement of the infected gingival tissues. Laboratory techniques are available to assist the practitioner in identifying this microorganism in dental plaque samples.

Invasion of periodontal tissues by different bacterial morphotypes has been reported in human periodontitis; however, limited information is available as to prevalence, localization and the bacterial species involved. The present study determined prevalence and gingival localization of Actinobacillus (Haemophilus) actinomycetemcomitans in periodontal lesions of juvenile periodontitis patients. Thirty-five gingival biopsies were obtained from 12 juvenile periodontitis patients at the time of periodontal therapy. One additional control biopsy was obtained from each of two adult periodontally healthy subjects, one adult periodontitis patient and one periodontally healthy monkey (Macaca fosibolius). The biopsies were carefully processed to avoid mechanical introduction of bacteria into the tissues and were examined using light and electron microscopy. Rabbit antisera specific for the three A. actinomycetemcomitans serotypes were used for immunofluorescence microscopic localization of A. actinomycetemcomitans antigens in the gingival sections. Immunofluorescence microscopy showed A. actinomycetemcomitans specific antigens in the gingival tissues of 11 of the 12 juvenile patients examined. None of the control specimens showed evidence of A. actinomycetemcomitans antigens in the gingival connective tissue. One specimen from a periodontally healthy subject and the monkey biopsy, however, showed A. actinomycetemcomitans antigens in bacterial plaque on the surface of the crevicular epithelium. Transmission electron microscopic examination showed microcolonies of small gram-negative rods in the connective tissue, as well as single bacterial cells between collagen fibers and in areas of cell debris. In addition to these extracellular bacterial cells, evidence of bacterial cells was also found within gingival connective tissue phagocytic cells. The data from the present study suggest that the gingival tissue in juvenile periodontitis lesions harbors A. actinomycetemcomitans.

Human oral spirochetes are prominent inhabitants of subgingival plaque in patients with periodontal disease. Measurements of serum antibody titers to these micro-organisms have been used to illuminate the role of human oral spirochetes in periodontal disease. In the present study, rabbit antisera to four oral spirochetes (including Treponema denticola ATCC33520 and three clinical isolates) were examined for reactivity to cell lysates. Western blotting demonstrated that the major treponemal antigens reactive with the rabbit antisera to T. denticola ATCC33520 and to strains 42, 48, and 57 possessed 53-kDa, 53-kDa, 56-kDa, and 56-kDa molecular weights, respectively. Human sera from patients with acute necrotizing ulcerative gingivitis (ANUG) and localized juvenile periodontitis (LJP) were also reactive with these antigens, particularly the 53-kDa antigen of T. denticola ATCC33520. A membrane-rich preparation was obtained from the cell lysate of T. denticola ATCC33520 by column chromatography and centrifugation, and applied to an SDS-polyacrylamide gel. The 53-kDa major peptide band was found. The membrane vesicles in an axial filament-membrane-containing fraction were agglutinated in the presence of the rabbit antiserum to T. denticola ATCC33520. Western blot analysis indicated that the 53-kDa antigen reacted strongly with the rabbit antiserum to T. denticola ATCC33520. These findings suggest that polypeptide antigens, such as the 53-kDa antigen from human oral spirochetes, play an important role in production of humoral antibodies associated with periodontal disease.

A water-soluble extract of Actinomyces viscosus was tested for its capacity to induce deoxyribonucleic acid synthesis in spleen cells from normal mice and to augment the in vitro antibody-forming cell (AFC) response to the T-cell-dependent antigen, sheep erythrocytes (SRBC) by T-cell-deficient adherent spleen cell monolayers from the same mice. Our results indicated that the water-soluble extract induced an in vitro blastogenic response, as assessed by tritiated thymidine incorporation into mouse spleen cells. The water-soluble extract also augmented the antibody response to SRBC. However, after fractionation of the water-soluble extract on Biogel P-300, not all of the fractions induced a blastogenic response and augmented an AFC response to SRBC. This difference in the ability of A. viscosus fractions to induce in vitro responses suggests that the water-soluble extract has multiple immunological properties. Chemical analysis of the water-soluble extract indicated an enrichment for amino acids and amino sugar common to peptidoglycan.

The adhesive interaction between the ectosymbionts of the corn cob configuration (CCC), a naturally occurring bacterial consortium in human dental plaque, was studied. In vitro association was produced in mixed cultures consisting ofBacterionema matruchotii and streptococci resemblingStreptococcus sanguis previously isolated from human CCC. Phase-contrast, selective immunofluorescence, and scanning electron microscopy showed that in this aggregative system, a tuftlike polar structure typical of the coccal epibiont mediated binding to the core filament. This aggregative model provides evidence that a specialized structure on the cell surface of epiphytic organisms may participate in interbacterial adherence.

A cocci-filament association was discovered in bacterial cultural studies of a subgingival plaque sample. Components were isolated and identified as Veillonella parvula and Eubacterium saburreum. A E. saburreum cell-associated material consisting of approximately 25% glucose and 70% protein was found which may play a role in the adherence of V. parvula to this filament. Acid end product analysis via gas liquid chromatography showed an uptake of lactic and succinic acids by V. parvula resulting in increased levels of acetic, propionic and n-butyric acids in mixed culture of the E. saburreum and the V. parvula strains. The ability of E. saburreum to adhere to nichrome wire and to glass surfaces as well as the secondary plaque forming ability of V. parvula indicates that these organisms may play a role in the maturation of human dental plaque.

A series of 22 5-(alkylsulfonyl)salicylanilides was synthesized and evaluated for in vitro antibacterial and antiplaque activity against Actinomyces viscosus and Streptococcus mutans, adherent microorganisms implicated in periodontal disease and dental caries. The minimum inhibitory concentrations of 25 salicylanilides (including 5-acyl-, 5-alkyl-, and 5-(alkylsulfonyl)-4'-bromo- and -4'-(trifluoromethyl)salicylanilides) were found to correlate (r = 0.94) with estimated log D values. Several salicylanilides, such as 5-(decylsulfonyl)- and 5-(dodecylsulfonyl)-4'-(trifluoromethyl)salicylanilides (15 and 19) were found to exhibit high levels of in vitro antibacterial and antiplaque activity against A. viscosus and S. mutans.

All study strains ofBacteroides gingivalis, B. asaccharolyticus, andB. melaninogenicus subspecies possessed numerous pilus-like fibers and capsule-like outer surface structures. The capsular morphology varied between the different species and subspecies.B. gingivalis strongly agglutinated 16 erythrocyte species studied.B. asaccharolyticus showed variable and weak agglutination of only a few erythrocyte species.B. melaninogenicus subsp.intermedius strains strongly agglutinated rabbit erythrocytes and exhibited variable, often weak agglutination of 8 other erythrocyte species. Preparations of capsular polysaccharide or lipopolysaccharide fromB. gingivalis failed to agglutinate human erythrocytes, while pili preparations from the same organisms possessed marked hemagglutinating activity.B. gingivalis cells adhered in high numbers to human buccal epithelial cells, whereas strains ofB. asaccharolyticus failed to show measurable adherence. Oral strains ofB. melaninogenicus subsp.intermedius feebly adhered to the buccal epithelial cells. Pretreatment ofB. gingivalis cells with serum or saliva prevented the adherence to epithelial cells. Our findings suggest that cell surfaces with distinct properties exist on the various black-pigmentedBacteroides species and subspecies and this may accout for markedly differing ability of these organisms to attach to mammalian cells.

Great progress has been made in our understanding of the pathogenesis of periodontal disease, the primary role of bacteria as etiologic agents, and the critical modifying role of host responses. It is useful to consider several stages in the pathogenesis of periodontal disease - (a) colonization, (b) invasion, (c) destruction, and (d) healing - and to place into perspective the various host responses as they may affect each of these four stages (Table 5). With respect to colonization, although very little direct evidence is available, it is reasonable to suggest that antibodies, either secretory or serum-derived, acting by virtue of their ability to block attachment, could inhibit colonization by immune reduction of adherence mechanisms. With respect to invasion of the tissue, it appears that phagocytes, particularly the neutrophils, are important, acting in concert with opsonic antibody and complement in ingesting and killing the periodontal microflora before or during the early invasive process. A major advance in our understanding of the pathogenesis of periodontal diseases is the realization that the virulence of periodontopathic bacteria relates to their leukaggressive properties, allowing them to evade neutrophil protective mechanisms. Invasion of the periodontal tissues by bacterial products may be inhibited by the complexing of these products with antibody with the formation of antigen-antibody complexes that are phagocytosed and digested, particularly by scavenger phagocytes such as the macrophage. With respect to the destructive phase of periodontal disease, it is clear that the direct effect of lymphocytes mediated either through direct cytotoxic activity, or through biologically-active destructive lymphokines (such as alpha-lymphotoxin and osteoclast activating factor), can lead to tissue destruction. Macrophages, through the production of monokines, collagenase, and reactive oxygen species, can also lead to tissue destruction. The direct effects of bacterial toxins or enzymes which can lead to tissue destruction can be inhibited by complexing with antitoxic or enzyme-neutralizing antibodies. With respect to healing and fibrosis, very little direct information is available; however, it is possible that the lymphocytes and macrophages affect fibrosis by the production of chemotactic factors for fibroblasts which would be expected to bring them to the area of periodontal inflammation and also by production of fibroblast-activating factors, which then cause the fibroblasts to proliferate and produce collagen which replaces lost collagen or results in fibrosis.(ABSTRACT TRUNCATED AT 400 WORDS)

The subgingival microflora and serum antibody response was examined in periodontitis patients with noninsulin-dependent diabetes mellitus (NIDDM), impaired glucose tolerance (IGT), and normal glucose tolerance (NGT). The predominant cultivable microflora was determined for subgingival plaque sampled from two deep periodontal pockets in each of eight adult periodontitis patients with NIDDM. Indirect immunofluorescence for Bacteroides intermedius, Bacteroides gingivalis, and Haemophilus actinomycetemcomitans was used to examine these same samples as well as 186 additional subgingival plaque samples from 47 patients with moderate to severe generalized periodontitis including 25 subjects with NIDDM, six subjects with IGT, and 16 subjects with NGT. Serum antibody levels to 13 microorganisms including seven oral bacterial species and one nonoral control species were measured by enzyme-linked immunosorbent assays (ELISA) in 377 subjects including 84 normal subjects without periodontal disease, 112 normal subjects with periodontitis, 19 periodontally normal subjects with IGT, 65 periodontitis patients with IGT, 15 periodontally normal subjects with NIDDM, and 82 periodontitis patients with NIDDM. Three hundred eighty-two bacterial isolates were recovered from the eight patients. B. intermedius was the most frequently isolated microorganism constituting 16% of the total isolates followed by Wolinella recta and B. gingivalis, which each accounted for 13% of the total. Streptococcus sanguis was the most prevalent microorganism, which was found in 75% of the sites. Subgingival plaque samples examined by immunofluorescence demonstrate a high prevalence of black-pigmented Bacteroides and suggest that the proportion of B. gingivalis but not B. intermedius is higher in NIDDM with periodontitis than in other groups.(ABSTRACT TRUNCATED AT 250 WORDS)

Subtraction radiography, a sensitive and accurate technique for identifying alveolar crestal change from standardized pairs of radiographs, is useful in monitoring periodontal therapy. One half of the radiographs were found to be appropriate for subtraction analysis using present technology for taking standardized radiographs. The criterion for usability was identical interpretation of subtraction images made in duplicate from a pair of radiographs. A set of radiographs was analyzed by subtraction radiography as well as by measurement of alveolar bone-crest height. Subtraction radiography was found to be more sensitive in detecting change. Whereas 53% pairs of radiographs showed a change on subtraction radiography, only 14% showed a change in crest height. Comparison of change by subtraction radiography and probing attachment level showed an overall correlation. Since these two measures assess different aspects of the periodontium, perfect correlation was not expected.

The name Bacteroides melaninogenicus subsp. macacae is proposed for a subspecies to accommodate strains of gram-negative, anaerobic, catalase-producing rods which produce black colonies on blood agar plates. The organisms weakly ferment galactose, glucose, lactose, and mannose and exhibit no fermentation of 24 other carbohydrates. The metabolic acid end products formed in peptone-yeast extract-glucose broth cultures include acetic, propionic, isobutyric, butyric, isovaleric, and succinic acids. The organisms utilize pyruvate, digest gelatin, and produce indole and hydrogen sulfide. B. melaninogenicus subsp. macacae appears to be serologically distinct from B. asaccharolyticus, B. melaninogenicus subsp. levii, B. melaninogenicus subsp. intermedius, and B. melaninogenicus subsp. melaninogenicus. The type strain of this new subspecies is ATCC 33141.

This report summarizes the rationale and illustrates the use of microbiological assays for specific microorganisms such as Actinobacillus (Haemophilus) actinomycetemcomitans in periodontal diseases. Data on the prevalence and monitoring of A. actinomycetemcomitans in patients with localized juvenile periodontitis (LJP) are presented. A. actinomycetemcomitans was found in 57 of 60 LJP patients for a prevalence of 95%. Two of the three patients not demonstrating the organism had serum antibodies to A. actinomycetemcomitans suggesting that they were once infected. Large numbers of A. actinomycetemcomitans were found in two-thirds or more of the periodontally involved sites in the LJP patients. Microbiologic and clinical data illustrates the critical need for complete eradication of this organism in order to ensure successful therapy. The general use of microbiological tests for specific microorganisms in individual periodontitis patients can be applied to: (a) determine the causative agent, (b) assess disease activity for treatment planning, (c) monitor the effects of treatment, and (d) decide on recall intervals. Bacterial culture of the subgingival flora and determination of antibiotic susceptibility is likely to be useful in patients in whom therapy has not been successful. Rapid assays are also useful in examining subjects in epidemiologic studies to determine the prevalence of pathogens in various populations, in longitudinal studies to assess the importance of various microorganisms in the etiology of periodontal disease, and to identify persons at-risk for either the initial episode of periodontal disease or for recurrent disease. The data presented here and the review of the relevant literature strongly support the use of microbiological assays for specific microorganisms as adjuncts in the diagnosis and management of periodontal disease. It is clear that meticulous, long-term supragingival plaque control and root planing to achieve smooth surfaces of the roots is very important for the management of most forms of periodontal disease, however, identification and monitoring of specific microorganisms represents a new approach to the management of periodontal disease and provides a rationale for treatment of the specific infections of the periodontium. Although all of the specific periodontal pathogens have not been defined, a sufficient number of important pathogens are known and testing for these organisms is, therefore, indicated. Furthermore, technologies have developed to the point where rapid microbiological tests are feasible and practical. Future developments will likely provide more efficient and improved tests. In conclusion, the rationale for assessment of several important periodontal pathogens and the present level of technology making such assessment feasible, strongly support the use of microbiologic assays as adjuncts in the clinical management of various forms of periodontal disease.

Two cases of Papillon-LeF evre are described. Both siblings demonstrated neutrophil dysfunction and severe precocious periodontal disease. The neutrophil locomotion defect was characterized by a decreased migration toward a chemotactic factor and decreased random migration. Binding of the chemotactic factor, FMLP, to the neutrophil surface was unchanged. Both patients harbored Actinobacillus actinomycetemcomitans and showed elevated serum IgG levels. Both patients also demonstrated salivary and serum antibody to A. actinomycetemcomitans. The Papillon-LeF evre syndrome is compared with the more common localized juvenile periodontitis.

M fascicularis monkeys were immunized subcutaneously in the vicinity of the major salivary glands and by retrograde infusion into the parotid duct, with a vaccine containing Formalin-killed S mutans strain 6715 cells and culture-fluid antigens. Indirect immunofluorescent staining was used to titrate and classify antibodies. Subcutaneous immunization induced only a serum response, whereas intraductal infusion stimulated both an IgA antibody response in the parotid fluid and a serum response. Immunized and nonimmunized control groups were orally infected with S mutans strain 6715. The establishment in dental plaque was quantitated by recovery of the infecting organism on selective media and by immunofluorescent staining of plaque smears taken from individual tooth surfaces. The establishment of S mutans strain 6715 was noticeably inhibited in immune monkeys. Immunofluorescent assays for antibody also showed that serum and parotid fluid containing serum IgA antibodies cross reacted with other d serotype and a serotype strains but not representative b and c strains. Immune and control groups were then orally infected with S mutans strain GS-5, a c serotype strain, and no inhibition in establishment was detected of the non-cross-reacting type c organism in the immune group. A latter series of booster immunizations via the intraductal route resulted in a significant decrease in parotid fluid flow. Histological investigations showed inflammatory cell infiltration and replacement of epithelium by connective tissue in the glands from immunized monkeys. A separate group of monkeys, younger than the first, was immunized with the same vaccine via the duct only. In this group, immunizations were given at shorter intervals, but the immunization response was similar to that observed in the first group. The investigations reviewed here and new experiments reported show that immunization of monkeys with S mutan strain 6715 via the parotid duct elicited a reproducible IgA antibody response in the parotid fluid as well as a serum antibody response. Inhibition of colonization on tooth surfaces in immune monkeys showed specificity for the immunizing strain suggesting that inhibition was antibody mediated.

The localization of IgG, IgM, IgA, and several complement components were studied in long-standing human periapical lesions. Sections of 38 Formalin-fixed and ten fast-frozen periapical lesions were stained with fluorescein-conjugated antibodies specific for IgG, IgM, IgA, C3, C4, and C3 proactivator (C3pa). IgG, IgA, and IgM were observed extracellularly as well as in plasma cells. Plasma cells containing IgG were most numerous and those containing IgM least numerous. Numerous non-Ig-containing round cells resembling lymphocytes also were found in most lesions. Five of ten lesions processed by fast freezing showed bright C3 staining of small vessel-like structures. This staining was blocked with unlabeled anti-C3 serums, but was not blocked by prior absorption of the anti-C3 conjugate with AB blood group substances. Deposits of C3 were also identified extracellularly in most of the lesions. C3pa and C4 were not observed in the ten lesions. These experiments suggest that reactions involving complement (C3) play a role in some periapical lesions.

A commercially available bacterial colony counter has been modified to allow rapid, accurate, semi-automated evaluation of cell numbers in the micropore filter assay for chemotaxis. The method is valuable for objective, rapid evaluation of cell counts at various levels through the filter, as well as counts on the distal surface of the filter. Coupled with a programmable calculator, this instrument had made feasible a new method of assessing random migration by the regression line analysis, which discriminates between migration rate and mass migration of cells. This combination of equipment may thus serve as a considerable time saving accessory to laboratories engaged in cell locomotion research, but also will allow more rigorous assessment of differences among specific populations of cells.

In order to assess the relationships among strains of the genera Actinobacillus and Haemophilus, DNAs from 50 strains of these genera were isolated and purified. The guanine plus cytosine (G + C) content of DNAs from strains of Haemophilus segnis and Haemophilus parainfluenzae were determined by thermal denaturation. DNA-DNA homologies were measured using labelled probes from one strain representing Haemophilus segnis (strain ATCC 10977), and two strains representing Haemophilus parainfluenzae (strains ATCC 9796 and ATCC 7901). Strains isolated as H. segnis had a G + C content of 39.0 to 42.9% and were 49-92% homologous with the ATCC 10977 DNA probe. All of the strains freshly isolated as H. parainfluenzae were 70-81% homologous with the ATCC 9796 DNA probe and had a G + C content of 34.9 to 38.3%. Strain ATCC 7901 was 11% homologous with the ATCC 9796 DNA probe, had a G + C content of 42.4%, and was 65-78% homologous to DNA from strains identified as Haemophilus aphrophilus and Haemophilus paraphrophilus. From these results we conclude that strain ATCC 7901 is a mislabelled strain of H. paraphrophilus. The results of multiple DNA-DNA hybridizations indicated that separate species designations were appropriate for H. segnis, H. parainfluenzae, Actinobacillus actinomycetemcomitans ("Haemophilus actinomycetemcomitans"), and H. aphrophilus. H. aphrophilus and H. paraphrophilus were closely related organisms and did not fulfill the generally accepted criteria for designation as separate species.

One objective of periodontal therapy after resolution of the infectious process is to facilitate a new connective tissue attachment to the root dentin surface. Here we report that a selective advantage for attachment and growth can be conferred on human gingival fibroblasts by biochemical manipulation of the dentin surface. Tetracycline HCL treatment of the dentin surface increases binding of fibronectin. The adsorbed fibronectin stimulates fibroblast attachment and growth, while suppressing epithelial cell attachment and growth. This biochemical manipulation may provide a useful approach for the treatment of periodontally involved teeth.

We studied the association between post-treatment periodontal disease activity and subgingival Bacteroides gingivalis, Bacteroides intermedius, spirochetes and motile rods. 20 adults, 22-62 years, with moderate-to-severe periodontitis participated in a split-mouth treatment study. All individual quadrants received supragingival cleaning and in addition, subgingival scaling and a NaHCO3-NaCl-H2O2 slurry, subgingival scaling alone, slurry alone, or no subgingival treatment. Post-treatment periodontal disease status was determined over a period of 12 months by changes in probing periodontal pocket depth and probing periodontal attachment level. Subgingival specimens obtained by paper point-sampling were evaluated for B. gingivalis and B. intermedius using indirect immunofluorescence and for spirochetes and motile rods using bright light phase contrast microscopy. A total of 142 periodontitis lesions representing all 4 quadrants of the 20 subjects were studied. The relationship between clinical data and bacteria was analyzed using logistic regression. The probability of the study organisms being present in subgingival sites at 3 to 6 months after treatment increased with increased residual pocket depth. The presence of B. gingivalis showed a strong positive association (p less than 0.004) with loss of periodontal attachment. A significant association was also found for spirochetes (p less than 0.008) but not for motile rods (p greater than 0.35) or B. intermedius (p greater than 0.13). Similar results were obtained at 12 months after therapy, except that the presence of motile rods was significantly associated with loss of periodontal attachment (p less than 0.03). Caution must be exercised when using B. gingivalis or spirochetes to evaluate treatment efficacy. If the presence of these organisms was utilized to indicate progressing periodontitis, many active lesions could be identified, and only 1 to 17% and 13 to 43% of sites in remission at 3-6 months after therapy would be expected to harbor B. gingivalis and spirochetes, respectively. The consequences of treating periodontal sites in remission would mainly be limited to cost and inconvenience. However, since several active periodontitis lesions did not reveal the organisms, treatment decisions based solely on the absence of the organisms may result in the omission of needed therapy. As a practical consideration, periodontal treatment should be continued as long as B. gingivalis and maybe spirochetes are detectable in the subgingival microflora. In the absence of these organisms, and until additional periodontal pathogens have become known, the decision to continue or conclude periodontal therapy must b

These studies demonstrate a unique constellation of organisms populating the subgingival area in periodontitis lesions of patients with juvenile or insulin-dependent diabetes mellitus (IDDM). The cultivable microflora was predominated by Capnocytophaga and anaerobic vibrios in the patients studied. In some patients, Actinobacillus actinomycetemcomitans were also found. This distinguishes the subgingival flora of IDDM patients suffering from periodontitis from that of patients with localized juvenile periodontitis (LJP), and that of adult periodontitis patients. In LJP most patients harbor both A actinomycetemcomitans and Capnocytophaga subgingivally; and in periodontitis lesions from nondiabetic adults, black-pigmented Bacteroides such as B gingivalis or B melaninogenicus subspecies intermedius are often found. Antibiotic susceptibility patterns suggest that penicillin or tetracycline or its analogs such as minocycline may be effective against the predominant cultivable microflora in periodontal lesions of IDDM patients; however, individual patients may harbor flora with significant resistance to these antibiotics.

The C3 cleavage products C3c and C3d were tested for their ability to alter the immunoproliferative response of human peripheral mononuclear cells to the antigens SLO and SK-SD, and to the mitogens PHA and PWM. It was found that both C3c (30 to 120 µg/ml) and C3d (10 to 40 µg/ml) inhibited lymphocyte blastogenesis in the presence on antigens but not mitogens, when cells were cultured in either autologous plasma or FCS. Similarly, the response to antigens of cell populations enriched for T lymphocytes was inhibited, whereas the response to optimal or suboptimal doses of mitogens was unaffected. When nonadherent (NA) cells were reconstituted with increasing numbers of adherent (AD) cells to potentiate the proliferative response of NA cells to the antigen SLO, the addition of either C3c or C3d abolished the potentiation of the response at low levels of reconstitution. However, at a given dose of C3c or C3d, addition of excess AD cells could restore the proliferative response. These results suggest that both C3c and C3d can inhibit T cell proliferation in response to antigen and that they may act at the level of the monocyte-T lymphocyte interaction to modulate cellular immune responses.