Leukotoxic effects of Actinobacillus actinomycetemcomitans

A sonic extract of Actinobarillus actinomycetemcomitans Y4, a microorganism originally isolated from a patient with juvenile periodontitis, contains a leukotoxin (Aa-leukotoxin) which specifically kills human polymorphonuclear leukocytes (PMNs) and monocytes. In the presence of normal human sera, the toxicity of the leukotoxin is enhanced, whereas sera from patients with juvenile periodontitis neutralize leukotoxic activity. In juvenile penodontitis serum immunoglobulin G was identified as the specific inhibitor of the Aaleukotoxin. The leukotoxin enhancing factor(s) in normal human serum appeared to be a large molecular weight protein which was not immunoglobulin. Pooled sera from rabbits immunized with A. actinomycetemcomitans Y4 sonic extract also neutralized Aaleukotoxic activity while normal rabbit serum enhanced toxicity. The presence of antileukotoxin antibody in sera from individuals with juvenile periodontilis suggests that these people have been immunized with A. actittotnyCetemeomitans microorganisms. Monitoring of this antibody may be a convenient tool in the study of the etiology of juvenile periodontitis.     

Antibody reactive with Actinobacillus actinomycetemcomitans leukotoxin in early-onset periodontitis patients

The objective of this study was to determine whether a relationship exists between antibody reactive with the Actinobacillus actinomycetemcomitans leukotoxin and the severity of periodontal disease. Serum concentrations of antibody reactive with the leukotoxin were determined for 119 early-onset periodontitis patients and 59 non-periodontitis subjects using limiting dilution analysis on Western blots. Immunoglobulin G (IgG) antibody reactive with the A. actinomycetemcomitans leukotoxin ranged from undetectable to 29 jig/ml (mean = 3.13±0.97 μg/ml for the generalized early-onset periodontitis and 2.17±0.86 μg/ml for the localized juvenile periodontitis patients vs 0.32±0.24 μg/ml for 59 non-periodontitis controls), and the dominant subclass was IgGl. Analysis of the relationship between antibody reactive with A. actinomycetemcomitans sonicate, A. actinomycetemcomitans leukotoxin and attachment loss patterns indicates that seropositive generalized early-onset periodontitis patients had decreased attachment loss compared with patients lacking this antibody. The statistical relationship appeared to be stronger for the sonicate than the purified leukotoxin. These data suggest that antibody reactive with A. actinomycetemcomitans leukotoxin may be protective in early-onset periodontitis, but given that the sonicate appeared better than the leukotoxin alone, it is not likely that leukotoxin is the only antigen of importance to host defense.     

Dynamics of infection by leukotoxic strains of actinobacillus actinomycetemcomitans in juvenile periodontitis

Juvenile periodontitis is associated with a high incidence of infection by Actinobacillus actinomycetemcomitans (Aa). The data presented indicate that the ability of Aa to destroy human PMNs is altered during the course of infection. Leukotoxic strains of Aa are characteristically found in isolates obtained from younger patients (6-12 years of age) but not in older subjects (13-25 years old). This suggests that the leukotoxin may be more important during early as opposed to more advanced phases of juvenile periodontitis.     

Actinobacillus actinomycetemcomitans in human periodontal disease

Recent evidence implicates Actinobacillus actinomycetemcomitans in the etiology of localized juvenile periodontitis. This paper reviews the morphological, biochemical and serological charcteristics of A. actinomycetemcomitans, evidence incriminating it as a periodontopathogen, its importance in human nonoral infections, and virulence factors which may be involved in the pathogenesis of A. actinomycetemcomitans infections. A. actinomycetemcomitans is a non-motile, gram-negative, capnophilic, fermentative coccobacillus which closely resembles several Haemophilus species but which does not require X or V growth factors. The organism has been categorized into 10 biotypes based on the variable fermentation of dextrin, maltose, mannitol, and xylose and into 3 serotypes on the basis of heat stable, cell surface antigens. A. actinomycetemcomitans' primary human ecologic niche is the oral cavity. It is found in dental plaque, in periodontal pockets, and buccal mucosa in up to 36% of the normal population. The organism can apparently seed from these sites to cause severe infections throughout the human body such as brain abscesses and endocarditis. There is a large body of evidence which implicates A. actinomycetemcomitans as an important micro-organism in the etiology of localized juvenile periodontitis including: (1) an increased prevalence of the organism in almost all localized juvenile periodontitis patients and their families compared to other patient groups; (2) the observation that localized juvenile periodontitis patients exhibit elevated antibody levels to A. actinomycetemcomitans in serum, saliva and gingival crevicular fluid; (3) the finding that localized juvenile periodontitis can be successfully treated by eliminating A. actinomycetemcomitans from periodontal pockets; (4) histopathologic investigations showing that A. actinomycetemcomitans invades the gingival connective tissue in localized juvenile periodontitis lesions; (5) the demonstration of several pathogenic products from A. actinomycetemcomitans including factors which may: (a) facilitate its adherence to mucosal surfaces such as capsular polysaccharides; (b) inhibit host defense mechanisms including leukotoxin, a polymorphonuclear leukocyte chemotaxis inhibiting factor, and a lymphocyte suppressing factor (c) cause tissue destruction such as lipopolysaccharide endotoxin, a bone resorption-inducing toxin, acid and alkaline phosphatases, collagenase, a fibroblast inhibiting factor and an epitheliotoxin.(ABSTRACT TRUNCATED AT 400 WORDS)     

Cytopathic effects of Actinobacillus actinomycetemcomitans on monkey blood leukocytes

The suitability of cynomolgus monkeys (Macaca fascicularis) for studies concerned with the biologic properties of Actinobacillus actinomycetemcomitans is the subject of the present investigation. We found that normal monkeys harbored leukotoxic strains of A. actinomycetemcomitans in subgingival plaque samples. Monkey peripheral blood PMNs and monocytes were killed following in vitro exposure to sonic extracts of leukotoxic strains of A. actinomycetemcomitans . Monkey sera were capable of inhibiting the leukotoxic properties of A. actinomycetemcomitans sonic extracts due to the presence of IgG antibodies which neutralized the leukotoxin. Similarly, sera from patients with juvenile periodontitis (but not normal human sera) abolished leukotoxin-mediated killing of monkey PMNs. Monkey peripheral blood lymphoid cells were not killed by A. actinomycetemcomitans but demonstrated depressed responses to mitogens following pre-incubation with A. actinomycetemcomitans sonicates prepared from either leukotoxic or "non-leukotoxic" human strains. These studies suggest that cynomolgus monkeys may serve as a suitable in vitro and in vivo model for delineating more about host- A. actinomycetemcomitans interrelationships in the etiology of human periodontal disease.     

Actinobacillus actinomycetemcomitans in human periodontal disease. Prevalence in patient groups and distribution of biotypes and serotypes within families

Actinobacillus actinomycetemcomitans is a Gram-negative oral bacterium which has been implicated in the etiology of localized juvenile periodontitis. In this study, 403 subjects from four study groups were examined for A actinomycetemcomitans in subgingival dental plaque. Samples pooled from at least six periodontal sites were included from each subject. A actinomycetemcomitans was detected in 28 of 29 localized juvenile periodontitis patients but in only 15% of the other subjects including 28 of 134 adult periodontitis patients, 24 of 142 periodontally healthy subjects and 5 of 98 insulin dependent juvenile diabetics with varying degrees of gingivitis. A actinomycetemcomitans isolates from members of five families with localized juvenile periodontitis patients were biotyped on the basis of variable fermentation of dextrin, maltose, mannitol and xylose and serotyped by indirect immunofluorescence using serotype specific rabbit antisera. Individuals within a family all harbored A actinomycetemcomitans of the same biotype and serotype. However, even in families with individuals heavily infected with A actinomycetemcomitans, some family members did not appear to be infected with the organism. The apparent poor transmissibility of A actinomycetemcomitans between individuals may, in part, explain the overall low prevalence of localized juvenile periodontitis and the familial pattern of the disease. The high prevalence of A actinomycetemcomitans in the subgingival plaque of localized juvenile periodontitis patients, compared to the much lower prevalence in other patient groups, supports the hypothesis that A actinomycetemcomitans is an etiologic agent in this periodontal disease.(ABSTRACT TRUNCATED AT 250 WORDS)     

Clonal diversity of Actinobacillus actinomycetemcomitans isolates from young adults with minimal periodontal disease

Actinobacillus actinomycetemcomitans is a major periodontal pathogen which is associated with both early-onset periodontitis and adult cases refractory to conventional periodontal therapy, although the organism has also been shown to be widely distributed among dentate healthy individuals. The observed disease status may be associated with a variation in virulence of different strains or clones. The aim of the present study was to analyse genotype distribution as assessed by an arbitrarily primed polymerase chain reaction (AP-PCR) among 51 isolates of A. actinomycetemcomitans recovered from more than 200 young adult recruits with no or minor periodontal disease. In addition, isolates from 25 periodontitis patients as well as reference strains were genotyped. Primers amplifying (i) a specific sequence in the ltxA region, (ii) a specific 16S rRNA sequence and (iii) sequences in the leukotoxin promoter region were used to verify species identity of the strains. Three random oligonucleotide primers were employed to analyse genomic polymorphisms of the organism by means of PCR. A total of 19 genotypes could be distinguished, which were grouped by cluster analysis into 5 major clusters based on genetic similarity and a complete linkage sort. Whereas 3 clusters assembled A. actinomycetemcomitans genotypes isolated from both healthy subjects and periodontitis patients, one cluster containing 4 different genotypes exclusively comprised isolates from healthy or gingivitis subjects. Another cluster with 2 genotypes consisted of strains originating from periodontitis patients (p < 0.05). One strain characterized by a specific 530 bp deletion in the promoter region of the leukotoxin region was identified in a Ghanese patient with localized juvenile periodontitis. It was concluded that there is considerable clonal diversity of A. actinomycetemcomitans strains isolated from healthy or periodontally diseased subjects, and that genetically closely related groups might be associated with health or disease.     

High-titer antisera from patients with periodontal disease inhibit bacterial capsule-induced bone breakdown

Solubilized surface-associated material (SAM) form a number of periodonotopathogenic bacteria have been shown to be potent stimulators of bone resorption in vitro in the murine calvarial bone culture assay. Antibodies to the constituents of SAM are also found in patients with periodontal disease. Serum from patients with severe generalized periodontitis (SGP) containing high titers of antibodies to the SAM of Porphyromonas gingivalis completely inhibited the bone resorption induced by SAM from this organism. In contrast, serum from patients with low titers of antibodies to SAM form P. gingivalis failed to inhibit bone resorption. High-titer sera (containing antibodies to SAM from Actinobacillus actinomycetemcomitans) from patients with localized juvenile periodontitis (LJP) were added to calvarial cultures stimulated with SAM from A. actinomycetemcomitans . Of 6 high-titer sera tested, only 4 inhibited bone breakdown, the other 2 sera having no effect on resorption. Low-titer sera were also ineffective at blocking bone resorption. This suggests that the antibody response to SAM may have a protective effect in patients with periodontal disease.     

Evidence for the role of highly leukotoxic Actinobacillus actinomycetemcomitans in the pathogenesis of localized juvenile and other forms of early-onset periodontitis

BACKGROUND: Actinobacillus actinomycetemcomitans leukotoxin is thought to be an important virulence factor in the pathogenesis of localized juvenile and other forms of early-onset periodontitis. Some highly leukotoxic A. actinomycetemcomitans strains produce 10 to 20 times more leukotoxin than other minimally leukotoxic strains. The distribution, clonality, and intrafamilial transmission of highly leukotoxic A. actinomycetemcomitans were examined in order to determine the importance of leukotoxin in the pathogenesis of periodontitis. METHODS: The polymerase chain reaction (PCR) was used to differentiate highly leukotoxic from minimally leukotoxic strains in examining 1,023 fresh A. actinomycetemcomitans isolates and strains from our culture collection. These were obtained from 146 subjects including 71 with localized juvenile periodontitis (LJP), 4 with early-onset periodontitis, 11 with post-localized juvenile periodontitis, 41 with adult periodontitis, and 19 periodontally normal subjects. The arbitrarily primed polymerase chain reaction (AP-PCR) analysis of 30 oral isolates from each of 25 subjects was used to determine the intraoral distribution of A. actinomycetemcomitans clones. AP-PCR was also used to examine the transmission of A. actinomycetemcomitans in 30 members of 6 families. The clonality of 41 highly leukotoxic A. actinomycetemcomitans strains was evaluated by both AP-PCR and ribotyping. RESULTS: Highly leukotoxic A. actinomycetemcomitans was found only in subjects with localized juvenile and early-onset periodontitis. Fifty-five percent of the LJP subjects harbored highly leukotoxic A. actinomycetemcomitans isolates. Seventy-three percent of the A. actinomycetemcomitans isolates in these subjects were highly leukotoxic. Highly leukotoxic A. actinomycetemcomitans infected younger subjects (mean age 13.95 years, range 5 to 28 years) than minimally leukotoxic (mean age 35.47 years, range 6 to 65 years). Most subjects were infected with only one A. actinomycetemcomitans genotype. However, PCR of whole dental plaques and subsequent analysis of up to 130 individual oral isolates suggested a possible shift in A. actinomycetemcomitans over time in that a few subjects harbored both highly leukotoxic and minimally leukotoxic strains. AP-PCR analysis was consistent with intrafamilial A. actinomycetemcomitans transmission. Ribotyping and AP-PCR analysis confirmed a previous report that highly leukotoxic A. actinomycetemcomitans consists of a single clonal type. CONCLUSIONS: This study suggests that localized juvenile and other forms of Actinobacillus-associated periodontitis are primarily associated with the highly leukotoxic clone of A. actinomycetemcomitans.     

Tissue localization of Actinobacillus actinomycetemcomitans in human periodontitis. I. Light, immunofluorescence and electron microscopic studies

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.     

Transmission and colonization of Actinobacillus actinomycetemcomitans in localized juvenile periodontitis patients

Actinobacillus actinomycetemcomitans is a Gram-negative oral microorganism, which has been implicated in the etiology of localized juvenile periodontitis and in severe medical infections such as bacterial endocarditis. This study evaluated the ability of periodontal probes to transmit A actinomycetemcomitans from juvenile periodontitis lesions to healthy gingival sulci in the same patient. Localized juvenile periodontitis patients exhibiting first molar and incisor alveolar bone loss and with large numbers of A actinomycetemcomitans in deep periodontal pockets were included in this study. A periodontal probe was inserted into periodontal pockets of 6 mm or greater depth. The probe was then placed into a healthy gingival sulcus of 3 mm or less, in the same subject. Fifty-five transfers by probing were made and A actinomycetemcomitans in both the donor and recipient sites was assessed by a selective culture technique. The results indicate that periodontal probes can become contaminated with A actinomycetemcomitans from juvenile periodontitis lesions during routine dental examinations and can transfer this microorganism from infected to previously uninfected sites. However, A actinomycetemcomitans inoculated into the healthy gingival sulci did not permanently colonize these sites since the organisms were eliminated within 3 weeks.     

Collagenase activity in gingival crevicular fluid of patients with juvenile periodontitis

The nature and origin of collagenases in gingival crevicular fluid of juvenile periodontitis patients was investigated. Gingival crevicular fluid collected from deep untreated periodontal pockets of juvenile periodontitis patients was found to contain only vertebrate collagenase (EC 3.4.24.7) activity that cleaved soluble type-I and -III collagens into 3/4 and 1/4 length fragments, as analyzed by SDS-polyacrylamide gel electrophoresis. Type II collagen was degraded at a markedly slower rate. This substrate specificity is indicative of collagenases produced by fibroblasts, epithelial cells and macrophages. We have previously found that collagenase in gingival crevicular fluid of adult periodontics patients appears to be mainly derived from polymorphonuclear leukocytes (PMN). The reasons for the apparent difference in collagenase source between the groups were investigated. We examined whether the pathogen characteristic for juvenile periodontitis, Actinobacillus actinomycetemcomitans, can release collagenase from normal human PMNs. All 10 A. actinomycetemcomitans strains tested, freshly isolated from the subgingival plaque of juvenile periodontitis patients, caused release of collagenase from PMNs in vitro. These results suggest that the lack of normally functioning PMNs in the periodontium of juvenile periodontitis patients may result in a colonization of bacteria that activate the resident periodontal cells to produce increased amounts of collagenase.     

Serum IgG antibodies to lipopolysaccharides in various forms of periodontal disease in man

Serum IgG antibody titres to lipopolysaccharides (LPS) from two strains of Actinobacillus actinomycetemcomitans were significantly elevated in juvenile periodontitis compared with other types of periodontal disease and with controls (p less than 0.05). The highest antibody titres to Bacteroides gingivalis LPS were in juvenile periodontitis, but this difference was significant only against the control group (p less than 0.01). In adult mild periodontitis there were higher antibody levels to LPS from Veillonella parvula compared with all other groups and controls (p less than 0.05). The possibility that high antibody titres to LPS from A. actinomycetemcomitans may play a protective role in juvenile periodontitis needs further investigation.     

Serological investigation of various forms of inflammatory periodontitis

Antibody levels towards sonicated whole cell extracts of selected oral Gram negative bacteria and Actinomyces viscosus were determined in sera from patients with various inflammatory forms of periodontal disease and from healthy control individuals. Antibody titres to two (non-leukotoxin producing) strains of Actinobacillus actinomycetemcomitans were significantly elevated in patients with juvenile periodontitis. Adult patients with severe periodontitis had significantly lower IgG antibody titres to Veillonella parvula (p < 0.001) or A. actinomycetemco-mitans strain NCTC 9710 (p < 0.01) and patients with mild periodontitis had decreased IgM antibody titres to A. actinomycetemcomitans strain NCTC 10979 (p < 0.05) when compared with the control subjects. Young adults with severe periodontitis showed marked individual differences in their humoral responses. Analysis of data revealed that only patients with a history of juvenile periodontitis had elevated IgG antibody titres to Bacteroides gingivalis (p < 0.001) and to V. parvula (p < 0.01). Hence the profiles of antibody levels to oral microorganisms identified patients who had previously manifested classical "localised" juvenile periodontitis.     

The presence of bacteria in the oral epithelium in periodontal disease. II. Immunohistochemical identification of bacteria

Serial histological sections of gingiva obtained from each of six advanced adult periodontitis, two localized juvenile periodontitis and two periodontally healthy patients were used for specific identification of bacteria within the oral epithelium and adjacent connective tissue. Healthy gingival biopsies served as controls. Sections from patients and control biopsies were Gram-stained and also screened with antibacterial sera associated with the peroxidase immunocytochemical technique for specific bacterial identification. The "Pop-off" electron microscopic technique was also used to further demonstrate the bacterial nature of peroxidase-stained material. In addition, the possible correlation between bacteria and areas of possible reduced keratinization was investigated. The results showed that sections of orthokeratinized healthy gingiva did not contain bacteria. Gram-stained sections from diseased sites contained large numbers of bacteria in the oral epithelium and adjacent connective tissue. Bacteroides gingivalis and to a lesser extent Capnocytophaga gingivalis were found in periodontitis, and Actinobacillus actinomycetemcomitans was found in juvenile periodontitis when the immunoperoxidase technique was used. The bacterial nature of peroxidase-stained material was confirmed by the "pop-off" technique. In the disease biopsies, bacterial presence was correlated with areas of reduced amounts of keratin suggesting that the oral epithelium may be a portal of entry for bacteria into gingival tissues.     

Frequency of 530-bp deletion in Actinobacillus actinomycetemcomitans leukotoxin promoter region

Actinobacillus actinomycetemcomitans strains showing a 530-bp deletion in the promoter region of the leukotoxin gene operon elaborate high amounts of leukotoxin that may play a role in the pathogenesis of periodontal disease. This study used polymerase chain reaction detection to determine the occurrence of the 530-bp deletion in 94 A. actinomycetemcomitans strains from individuals of various ethnic backgrounds. Eleven blacks and one Hispanic subject but no Caucasian or Asian subjects showed the 530-bp deletion in the leukotoxin promoter region, suggesting that the deletion is mainly a characteristic of individuals of African descent. A. actinomycetemcomitans strains exhibiting a deletion in the leukotoxin promoter region occurred both in individuals having severe periodontitis and in adolescents revealing no evidence of destructive periodontal disease.     

The gingival immune response to periodontal pathogens in juvenile periodontitis

A gingival explant culture system was utilized to evaluate the reactivity of local immunoglobulins produced by juvenile periodontitis tissue. Gingival explant culture supernatant fluids were screened, via a standardized dot-immunobinding assay, for antibodies reactive to: Actinobacillus actinomycetemcomitans, Porphyromonas gingivalis, Prevotella intermedia, Campylobacter rectus, Eikenella corrodens, Peptostreptococcus micros, Peptostreptococcus anaerobius, Capnocytophaga ochracea, Eubacterium nodatum and Fusobacterium nucleatum and one nonoral microorganism, Porphyromonas asaccharolytica. Of the 75 juvenile periodontitis supernatant fluids tested, the organisms that reacted with the highest numbers of supernatant fluids were E. nodatum (72%) and A. actinomycetemcomitans (49%). More juvenile periodontitis than healthy tissue samples showed supernatant fluid reactivity to P. intermedia, C. ochracea, E. nodatum and P. micros. No significant difference was observed between the juvenile periodontitis group supernatant fluids reactivity and the supernatant fluids of the other periodontal disease groups tested. Cluster analysis revealed the association, as determined by supernatant fluid reactivity, of P. micros and C. ochracea in the juvenile periodontitis group. The data from this investigation are consistent with a hypothesis of multiple possible etiologies of periodontal destruction in juvenile periodontitis and other forms of periodontal diseases.     

Identification of periodontopathic bacteria in gingival tissue of Japanese periodontitis patients

INTRODUCTION: The identification of invading periodontopathic bacteria in tissues is important to determine their role in the pathogenesis of periodontal disease. The objective of this study was to identify periodontopathic bacteria in diseased gingival tissue of periodontitis patients. METHODS: Subgingival plaque and gingival tissue were collected from 32 generalized chronic periodontitis (CP), 16 generalized aggressive periodontitis (GAgP) and eight localized aggressive periodontitis (LAgP) patients. Detection frequencies and quantities of Porphyromonas gingivalis, Actinobacillus actinomycetemcomitans and Tannerella forsythensis were investigated by polymerase chain reaction. The prevalences of Streptococcus oralis and Streptococcus sobrinus were also examined and the distribution of A. actinomycetemcomitans serotypes was observed. RESULTS: P. gingivalis and T. forsythensis were detected in approximately 70% of tissue samples and 50% of plaque samples in the three periodontitis groups. Prevalence of A. actinomycetemcomitans in tissue samples was higher in the LAgP (63%) group than in either the CP (16%) or the GAgP (38%) group. A. actinomycetemcomitans serotype c was detected in 50% of LAgP patients. Detection frequencies of S. oralis and S. sobrinus were markedly low in both plaque and tissue samples from all three periodontitis groups. Amounts of P. gingivalis, A. actinomycetemcomitans and T. forsythensis in the tissue samples were not different among the three periodontitis groups. CONCLUSION: P. gingivalis, A. actinomycetemcomitans and T. forsythensis can localize in diseased gingival tissue and may be involved in periodontal tissue destruction. Serotype c is the predominant serotype of A. actinomycetemcomitans in Japanese LAgP patients.     

Prevalence of Actinobacillus actinomycetemcomitans in an ethnic adult Chinese population

AIM: The aim of this study was to determine the prevalence and the structure of the leukotoxin promoter region of Actinobacillus actinomycetemcomitans in an ethnic Chinese population. METHOD: Subgingival plaque samples were collected from 42 patients with moderate to advanced periodontitis and 50 periodontally healthy patients. A. actinomycetemcomitans was detected directly from the crude subgingival plaque by PCR using leukotoxin gene specific primers. The presence of A. actinomycetemcomitans was determined by a single 285 bp PCR amplicon. RESULTS: A. actinomycetemcomitans was found to be present in the subgingival plaque of 68 out of a total of 92 patients examined (74%). 29 out of the 42 periodontitis patients tested were carriers of A. actinomycetemcomitans (69%). Among the periodontally healthy patients studied, 39 out of 50 subjects possessed the bacteria (78%). PCR analysis of the promoter region of the ltx operon revealed that none of the 42 moderate to advanced periodontitis patients examined harboured A. actinomycetemcomitans strains with the JP2-like promoter of the ltx operon, known to enhance leukotoxin expression. 2 out of the 27 advanced periodontitis patients clinically diagnosed as suffering from rapidly progressive periodontitis were found to be carriers of the mildly toxic strain of A. actinomycetemcomitans with the characteristic 652-like promoter. CONCLUSIONS: The high prevalence of A. actinomycetemcomitans, regardless of whether the subgingival samples were analysed from patients with healthy or diseased periodontium suggests that this bacterial species is part of the normal oral flora of ethnic Chinese. Our preliminary results also suggested that subjects who harboured the mildly toxic strain of A. actinomycetemcomitans were potentially susceptible to aggressive forms of periodontitis.     

The immunoreactivity of systemic antibodies to Actinobacillus actinomycetemcomitans and Porphyromonas gingivalis in adult periodontitis

Actinobacillus actinomycetemcomitans and Porphyromonas gingivalis secrete several potent virulence factors and are known to be two of the major periodontal pathogens. In the present case-control study, the systemic immunoreactivity to A. actinomycetemcomitans exotoxins, cytolethal distending toxin (Cdt) and leukotoxin was analyzed in adult subjects with periodontitis and in periodontally healthy controls. Furthermore, systemic immunoreactivity to P. gingivalis was analyzed in these subjects. Reactivity to the A. actinomycetemcomitans toxins was determined in bioassays that quantified neutralizing antibodies, and P. gingivalis antibodies were detected by enzyme-linked immunosorbent assay (ELISA). The results showed a significantly enhanced immunoreactivity to P. gingivalis in the subjects with periodontitis, while the reactivity to A. actinomycetemcomitans leukotoxin showed no significant difference between patients and controls. However, combined immunoreactivity to leukotoxin and Cdt was more prevalent in the subjects with periodontitis than in the controls. In addition, immunoreactivity to leukotoxin correlated to periodontitis in men but not in women. In conclusion, data from the present study indicate that immunoreactivity to P. gingivalis is frequent in adult periodontitis, while the role of A. actinomycetemcomitans seems to be more complex and depends on gender of the infected subject as well as the virulence of the bacteria.