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 promotor region occurred both in individuals having severe periodontitis and in adolescents revealing no evidence of destructive periodontal disease.     

Black-pigmented anaerobic rods in closed periapical lesions

AIM: This study determined the frequency of Porphyromonas endodontalis, Porphyromonas gingivalis, Prevotella intermedia and Prevotella nigrescens in 20 closed periapical lesions associated with symptomatic and asymptomatic refractory endodontic disease. METHODOLOGY: To deliniate possible oral sources of P. endodontalis, the presence of the organism was assessed in selected subgingival sites and saliva in the same study patients. Periapical samples were obtained by paper points during surgical endodontic procedures using methods designed to minimize contamination by non-endodontic microorganisms. Subgingival plaque samples were obtained by paper points from three periodontal pockets and from the pocket of the tooth associated with the closed periapical lesion. Unstimulated saliva was collected from the surface of the soft palate. Bacterial identification was performed using a species-specific polymerase chain reaction (PCR) detection method. RESULTS: P. endodontalis was not identified in any periapical lesion, even though subgingival samples from eight patients (40%) revealed the P. endodontalis-specific amplicon. P. gingivalis occurred in one periapical lesion that was associated with moderate pain. P. nigrescens, P. endodontalis and P. intermedia were not detected in any periapical lesion studied. CONCLUSIONS: Black-pigmented anaerobic rods appear to be infrequent inhabitants of the closed periapical lesion.     

Periodontitis lesions are the main source of salivary cytomegalovirus

Background:  Herpesviruses play causal or cooperative roles in childhood infections, tumorigenesis, ulcerogenesis, and periodontitis. Saliva is a common vehicle of herpesvirus horizontal transmission, but the source of salivary herpesviruses remains obscure. To evaluate the significance of periodontal disease in shedding of oral herpesviruses, this study determined the genome-copy counts of human cytomegalovirus (HCMV) and Epstein–Barr virus (EBV) in whole saliva of subjects with periodontitis, gingivitis, or no natural teeth.
Methods:  Whole saliva was collected from 14 periodontitis patients, 15 gingivitis patients and 13 complete denture wearers. The study subjects were systemically healthy and had not received periodontal treatment in the past 3 months. Real-time TaqMan polymerase chain reaction was used to determine the salivary load of HCMV and EBV.
Results:  Salivary HCMV was detected in seven (50%) periodontitis patients, but not in any gingivitis or edentulous subjects ( P  < 0.001). Salivary EBV was detected in 11 (79%) periodontitis patients, in five (33%) gingivitis patients, and in seven (54%) edentulous subjects ( P  = 0.076). Salivary samples showed copy counts of HCMV in the range of 3.3 × 10³–4.2 × 10⁴/ml and of EBV in the range of 3.6 × 10²–1.6 × 10⁹/ml.
Conclusions:  HCMV and EBV are commonly present in the saliva of periodontitis patients. Periodontitis lesions of systemically healthy subjects seem to constitute the main origin of salivary HCMV, but do not comprise the sole source of salivary EBV.     

Salivary infectious agents and periodontal disease status

Saygun I, Nizam N, Keskiner I, Bal V, Kubar A, Aç?kel C, Serdar M, Slots J. Salivary infectious agents and periodontal disease status. J Periodont Res 2011; 46: 235–239. © 2011 John Wiley & Sons A/S Background and Objectives: The potential of salivary microorganisms to diagnose periodontal disease and to guide periodontal treatment is a research topic of current interest. This study aimed to determine whether the salivary counts of periodontopathic microbes correlated with the periodontal pocket counts of the same infectious agents, and whether the salivary counts of the test infectious agents could distinguish among individuals with periodontal health and various types of periodontal disease. Material and Methods: The study included 150 systemically healthy adults, of whom 37 were periodontally healthy, 31 had gingivitis, 46 had chronic periodontitis and 36 had aggressive periodontitis. Each study subject contributed microbial samples from the two deepest periodontal pockets of the dentition and from whole saliva. Aggregatibacter actinomycetemcomitans, Campylobacter rectus, Fusobacterium nucleatum, Porphyromonas gingivalis, Prevotella intermedia, Tannerella forsythia and Epstein–Barr virus were identified using the TaqMan real‐time PCR methodology. Statistical analysis was performed using the Mann–Whitney U‐test and the receiver operating characteristic statistics. Results: C. rectus, F. nucleatum, P. gingivalis, P. intermedia and T. forsythia occurred with significantly higher copy‐counts in salivary samples from patients with gingivitis, chronic periodontitis and aggressive periodontitis than from periodontally healthy individuals. A. actinomycetemcomitans only showed higher salivary copy‐counts in subjects with aggressive periodontitis compared with subjects with healthy periodontium, and the salivary copy‐counts of Epstein–Barr virus did not reveal any significant difference among the four subject groups studied. The diagnostic sensitivity for periodontitis was 89.19 for P. gingivalis and for T. forsythia and 86.49 for P. intermedia, with specificities ranging from 83.78 to 94.59. The optimal copy‐counts per mL saliva for identifying periodontitis were 40,000 for P. gingivalis, 700,000 for T. forsythia and 910,000 for P. intermedia. Conclusion: Salivary copy‐counts of P. gingivalis, T. forsythia and P. intermedia appear to have the potential to identify the presence of periodontitis, whereas the salivary level of the other test infectious agents may possess little or no diagnostic utility. Longitudinal studies are warranted to determine the ability of salivary copy‐counts of major periodontopathic bacteria to predict future periodontal breakdown.     

Quantitative analysis of association between herpesviruses and bacterial pathogens in periodontitis

Background and Objective: The development of human periodontitis may depend upon cooperative interactions among herpesviruses, specific pathogenic bacteria and tissue‐destructive inflammatory mediators. This study sought to identify associations among human cytomegalovirus, Epstein–Barr virus and six putative periodontopathic bacteria in periodontitis lesions. Material and Methods: Fifteen periodontitis patients (nine with aggressive periodontitis and six with chronic periodontitis) and 15 periodontally normal subjects were included in the study. In each study subject, a microbiological sample was collected, using a curette, from the deepest periodontal probing depth of the dentition. A real‐time TaqMan® polymerase chain reaction assay was employed to determine the subgingival counts of human cytomegalovirus, Epstein–Barr virus, Porphyromonas gingivalis, Tannerella forsythia, Prevotella intermedia, Aggregatibacter actinomycetemcomitans, Fusobacterium nucleatum and Campylobacter rectus. Statistical analysis was performed using the Student's t‐test, the Pearson correlation coefficient test and the single variable logistic regression test for odds ratio‐based risk calculation. Results: Human cytomegalovirus was detected in eight periodontitis lesions and in one normal periodontal site, Epstein–Barr virus was detected in nine periodontitis lesions and in two normal periodontal sites, and the study bacteria were detected in 6–15 periodontitis lesions and in 1–11 normal periodontal sites. Correlations were found between counts of human cytomegalovirus and Epstein–Barr virus, between counts of human cytomegalovirus and P. gingivalis, T. forsythia and C. rectus, and between counts of Epstein–Barr virus and P. gingivalis and T. forsythia. Human cytomegalovirus and Epstein–Barr virus counts were also positively associated with the level of periodontal attachment loss, probing pocket depth and gingival bleeding on probing. Conclusion: This study confirmed that periodontal human cytomegalovirus and Epstein–Barr virus are associated with major periodontopathic bacteria and with the severity of periodontal disease. The finding of abundant herpesviruses in periodontitis lesions redefines the pathogenic paradigm of the disease. Understanding the interplay between herpesviruses and specific bacterial species in the pathogenesis of periodontitis may form the basis for new approaches to preventing, reducing or delaying tissue breakdown from periodontal infections.     

Incidence of periodontitis recurrence in treated patients with and without cultivable Actinobacillus actinomycetemcomitans, Prevotella intermedia, and Porphyromonas gingivalis: a prospective study

A total of 98 adults previously treated for moderate to advanced periodontitis and on a trimonthly recall schedule were screened for the presence of critical levels of Actinobacillus actinomycetemcomitans, Prevotella (Bacteroides) intermedia, and Porphyromonas (Bacteroides) gingivalis. Patients with at least 2 positive sites were placed in a positive group and patients without or with low levels of these bacteria in a negative group. During the 30-month study the incidence of disease recurrence was greater in the positive group, but did not reach statistical significance. Positive patients with deeper pockets tended to be at greater risk of developing recurrent disease than those with shallower pockets. In the positive group only, both A. actinomycetemcomitans recovery and antibody levels to A. actinomycetemcomitans strain NCTC 9710 (serotype c) were inversely correlated with disease recurrence. The presence of A. actinomycetemcomitans and P. intermedia above critical levels did not reliably predict future episodes of disease recurrence in this population. The sparse recovery of P. gingivalis did not permit us to assess its diagnostic value. With the exception of P. gingivalis, for which insufficient data were available, the results indicate that the presence or absence of the above bacterial species cannot of itself serve as a reliable predictor of future episodes of recurrent disease in a population of treated patients on a regular trimonthly recall schedule.     

Campylobacter rectus in human periodontitis

Campylobacter rectus (formerly Wolinella recta ) in periodontitis lesions was studied relative to age and sex distribution, relationship to disease-active periodontitis, response to periodontal debridement and in vitro antimicrobial susceptibility. Subgingival C. rectus was collected with paper points, transported in VMGA III and plated onto nonselective enriched brucella blood agar and Hammond's selective medium for C. rectus , both incubated anaerobically. C. rectus was recovered from 80% of 1654 periodontitis patients. Although the organism showed similar age and sex occurrence, its proportional recovery in culture-positive adults was inversely related to increasing age ( r = 0.999, P <0.001). The organism was positively associated (summary odds ratio = 2.95) with disease activity in a 24-month longitudinal study of 93 adult periodontitis patients on maintenance therapy. C. rectus decreased from 8.2% to 0.7% following local periodontal debridement of 20 culture-positive adult periodontitis patients. The organism exhibited high in vitro susceptibility to therapeutic levels of tetracycline hydrochloride, metronidazole, penicillin G and ciprofloxacin. These findings further delineate the epidemiology and potential pathogenic role of C. rectus in human periodontitis.     

Herpesviruses in HIV-periodontitis

BACKGROUND/AIMS: Human herpesvirus-associated diseases exhibit elevated morbidity and mortality in patients infected with human immunodeficiency virus (HIV).This study aimed to investigate the occurrence of herpesviruses in HIV-periodontitis. METHOD: Gingival biopsies from periodontitis lesions of 21 HIV-patients and 14 non HIV-patients were studied. Nested-polymerase chain reaction methods were employed to detect human cytomegalovirus, Epstein-Barr virus type 1 and 2 (EBV-1, EBV-2), herpes simplex virus, human herpes virus (HHV)-6, HHV-7 and HHV-8. RESULTS: Gingival biopsies from HIV-periodontitis lesions showed on average 4.0 herpesvirus species and gingival biopsies from HIV periodontitis lesions of non-HIV patients revealed an average of 1.9 herpesvirus species (p<0.001). Occurrence of 4 to 6 different herpesviruses was more common in HIV- than in non HIV-gingival biopsies (71% vs. 7%) (p<0.001). EVB-2 was detected in 12 (57%) biopsies from HIV-periodontitis but was absent in non HIV-periodontitis biopsies (p= 0.002). HHV-6 also occurred in significantly higher frequency in HIV-periodontitis (71%) than in non HIV-periodontitis (21%) (p=0.01). HHV-8 was detected only in biopsies from HIV-periodontitis lesions.. CONCLUSION: HIV-periodontitis seems to be associated with elevated occurrence of EBV-2, HHV-6 and herpesvirus co-infections compared to periodontitis in non-HIV-patients. The periodontopathic significance of herpesviruses in HIV-periodontitis constitutes a research topic of considerable interest.     

Subgingival microflora and periodontal disease

Abstract This article describes the subgingival microflora of the healthy periodontium, gingivitis, advanced adult periodontitis, and juvenile periodontitis. A total of seven to nine subjects were examined in each of the four periodontal clinical entities listed. The individual bacteriological samples included material from the base of a single periodontal pocket. The sampling, the treatment of the samples, and the bacteriological cultivations were carried out using continuous anaerobic techniques. Briefly, the healthy gingival sulcus harbored a scant microflora dominated by Gram-positive organisms (85%), usually Streptococcus and facultative Actinomyces species. The development of gingivitis was accompanied by a marked increase in the total number of Gram-negative organisms. Fusobacterium nucleatum, Bacteroides melaninogenicus ss. intermedius, Haemophilus species, and other Gram-negative organisms comprised about 45% of the total gingivitis isolates. Streptococcus and facultative and anaerobic Actinomyces species constituted the majority of the Gram-positive gingivitis isolates. The micro-flora of advanced adult periodontitis was comprised mainly of Gram-negative anaerobic rods (about 75%), B. melaninogenicus ss. asaccharolyticus and F. nucleatum being the most predominant isolates. The deep pocket microflora in juvenile periodontitis was also made up mainly of Gram-negative organisms (about 65%), but was of a nature different from that of adult periodontitis, being predominated by isolates of Bacteroides species and other organisms of unknown species. The present article also concerns factors of importance for the colonization of Gram-negative anaerobic rods in the oral cavity and periodontal pockets. In vitro experiments showed that cells of B. melaninogenicus ss. asaccharolyticus and other Gram-negative organisms attached in high numbers to epithelial cells, hydrosyapatite (HA) surface, and Gram-positive bacteria when suspended in phosphate-buffered saline; however, the bacterial attachment to epithelial cells and HA was strongly inhibited in the presence of human saliva and serum. In contrast, saliva and serum had little effect upon the attachment of Gram-negative bacteria to Gram-positive bacterial cells. These findings agreed well with data from an in vivo study, in which streptomycin-labeled cells of B. melaninogenicus ss. asaccharolyticus were introduced into the mouth of two volunteers. A significantly higher number of B. melaninogenicus cells was recovered from dental plaque than from the other oral surfaces studied. The present series of studies has pointed to certain Gram-negative organisms as potential pathogens in rapidly progressing periodontal lesions. The available data on oral microbial ecology suggest that the presence of dental plaque containing Gram-positive organisms may be essential for the attachment and colonization of several Gram-negative species after their initial introduction into the mouth and the periodontal pocket area. The clinical relevance of these findings is discussed.     

Herpesviruses 6, 7 and 8 in HIV- and non-HIV-associated periodontitis

Human herpesviruses, especially cytomegalovirus and Epstein Barr virus type-1, occur with higher frequency in subgingival specimens from periodontitis lesions than from healthy/gingivitis sites. Little or no information is available on the relationship between herpesvirus 6 (HHV-6), herpesvirus 7 (HHV-7) and herpesvirus 8 (HHV-8) and periodontal disease. This study determined the periodontal occurrence of HHV-6, HHV-7 and HHV-8 in 21 HIV-seropositive and 14 HIV-negative adults affected by periodontitis. Gingival biopsy specimens and paper-point samples of subgingival plaque were collected from sites showing 5 mm or more in probing depth. Nested polymerase chain reaction methodology was employed in herpesvirus identification. In the HIV-seropositive periodontitis group, 90% of gingival biopsies and 62% of subgingival plaque samples revealed at least one of the test viruses. HHV-6 occurred in 71%, HHV-7 in 67% and HHV-8 in 24% of gingival biopsies. In the HIV-negative adult periodontitis group, 43% of gingival biopsies showed at least 1 of the test viruses, with HHV-6 present in 21% and H HV-7 in 29% of gingival biopsies and with no detection of HHV-8. The combined occurrence of the 3 test herpesviruses was significantly higher in HIV-seropositive than in HIV-negative adult periodontitis patients (p = 0.008). The human periodontium might constitute a site of infection or reservoir for HHV-6, -7, -8.