Morten Enersen

Human Cytomegalovirus and Epstein-Barr Virus in Apical and Marginal Periodontitis: A Role in Pathology?

Periodontitis is presumably caused by bacterial infection, but it has been shown recently that affected tissue often contains human cytomegalovirus (HCMV) and Epstein–Barr virus (EBV). The present study was initiated to evaluate the role of these viruses in the pathogenesis of periodontitis. HCMV and EBV were quantified in 40 apical and 25 marginal periodontitis samples using real time PCR. In situ hybridization or immunohistochemistry was carried out on apical samples to detect viral presence within cells. A possible association with relevant bacteria was examined. Of the apical periodontitis samples, 50% contained EBV, while none contained HCMV. Of the marginal periodontitis samples, 40% were positive for EBV and 12% for HCMV. With one exception, however, the amount of virus was close to the detection limits. EBV was only detected in 1 out of 15 healthy periodontium samples. Immunohistochemistry and in situ hybridization were all negative. Significant associations were found between periodontal EBV and the presence of Aggregatibacter actinomycetemcomitans and Porphyromonas gingivalis. Although there was an obvious association of the virus with clinical samples, it seems unlikely that these viruses play a major role in the pathogenesis of periodontitis of the average patient. Their presence may reflect that the clinical samples contain more blood or saliva compared to controls, or an accumulation of lymphoid cells harboring virus in the inflamed tissue. J. Med. Virol. 80:1007–1011, 2008.

Porphyromonas gingivalis fimbriae

Marginal periodontitis is not a homogeneous disease but is rather influenced by an intricate set of host susceptibility differences as well as diversities in virulence among the harbored organisms. It is likely that clonal heterogeneity of subpopulations with both high and low levels of pathogenicity exists among organisms harbored by individuals with negligible, slight, or even severe periodontal destruction. Therefore, specific virulent clones of periodontal pathogens may cause advanced and/or aggressive periodontitis. Porphyromonas gingivalis is a predominant periodontal pathogen that expresses a number of potential virulence factors involved in the pathogenesis of periodontitis, and accumulated evidence shows that its expression of heterogenic virulence properties is dependent on clonal diversity. Fimbriae are considered to be critical factors that mediate bacterial interactions with and invasion of host tissues, with P. gingivalis shown to express two distinct fimbria-molecules, long and short fimbriae, on the cell surface, both of which seem to be involved in development of periodontitis. Long fimbriae are classified into six types (I to V and Ib) based on the diversity of fimA genes encoding FimA (a subunit of long fimbriae). Studies of clones with type II fimA have revealed their significantly greater adhesive and invasive capabilities as compared to other fimA type clones. Long and short fimbriae induce various cytokine expressions such as IL-1α, IL-β, IL-6, and TNF-α, which result in alveolar bone resorption. Although the clonal diversity of short fimbriae is unclear, distinct short fimbria-molecules have been found in different strains. These fimbriae variations likely influence the development of periodontal disease.

FimA Genotypes and Multilocus Sequence Types of Porphyromonas gingivalis from Patients with Periodontitis

ABSTRACT Fimbriae are important virulence factors of pathogenic bacteria, facilitating their attachment to host and bacterial cells. In the periodontal pathogen Porphyromonas gingivalis, the fimA gene is classified into six types (genotypes I, Ib, II, III, IV, and V) on the basis of different nucleotide sequences, with fimA genotypes II and IV being prevalent in isolates from patients with periodontitis. The aims of this study were to examine the distribution of fimA genotypes in a collection of 82 P. gingivalis isolates from adult periodontitis patients of worldwide origin and to investigate the relationship between the fimA genotypes and the sequence types (STs), as determined by multilocus sequence typing (MLST), of the isolates. The fimA gene was amplified by PCR with primer sets specific for each genotype. The STs of all strains were assigned according to the MLST database for P. gingivalis (www.pubmlst.org/pgingivalis ). The 82 strains showed extensive genetic diversity and were assigned to 69 STs. Only isolates with closely related STs harbored the same fimA genotype. Twenty-eight (34.1%) strains harbored fimA genotype II, while only the reference strain for fimA genotype V reacted with the primers specific for this genotype. Twenty-one isolates (25.6%) were positive by more than one of the fimA PCR assays; the most frequent combinations were genotypes I, Ib, and II (eight isolates) and genotypes I and II (four isolates). Sequencing of the fimA gene from selected isolates did not support the observed specific fimA genotype combinations, suggesting that the genotyping method used for the major fimbriae in P. gingivalis should be reevaluated.

Patient with severe periodontitis and subgingival Epstein-Barr virus treated with antiviral therapy

It has been suggested that the presence of Epstein-Barr virus (EBV) can increase the severity of marginal periodontitis. We administered antiviral treatment (Valtrex for a period of 10 days) to a patient with recurrent periodontal disease and a high EBV load subgingivally. The antiviral treatment decreased the presence of EBV to the detection limit and the periodontal condition improved dramatically. One year after treatment, the periodontal condition was still stable and the virus barely detectable. The case suggests that virus screening and subsequent antiviral therapy may be useful as an adjunct to conventional periodontal therapy.

Multilocus Sequence Typing of Porphyromonas gingivalis Strains from Different Geographic Origins

ABSTRACT Porphyromonas gingivalis is an important periodontal pathogen that can be isolated from both active and inactive periodontal lesions. Apparently, differences in virulence between P. gingivalis strains exist, but the mechanisms underlying these differences are not yet fully understood. To obtain more information about pathogenicity and virulence of P. gingivalis, it is relevant to assess the genetic population structure of the species and to examine the occurrence of putative virulence factors against the genetic background. Presently, multilocus sequence typing (MLST) is the best method for analyzing bacterial population structures. Forty P. gingivalis strains from worldwide sources were analyzed by MLST. Internal 310- to 420-bp DNA fragments of the eight ubiquitous chromosomal genes, ftsQ, hagB, gdpxJ, pepO, mcmA, recA, pga, and nah, were amplified by PCR and then sequenced. The number of alleles at individual loci ranged from 2 to 19, and a total of 33 allelic profiles, or sequence types (STs), were identified. Nucleotide variation between alleles was located at one or a few sites. Identical or similar STs were found in isolates from different geographic regions. Our results showed signs of a clonal population structure with a level of recombination not as high as that previously suggested for the species. We also found that P. gingivalis isolates from individual patients were genetically heterogeneous.

Er:YAG laser in the treatment of periodontal sites with recurring chronic inflammation: a 12‐month randomized, controlled clinical trial

AIM The objective of this randomized, controlled clinical trial was to compare the clinical and microbiological effects of pocket debridement using erbium-doped: yttrium, aluminium and garnet (Er:YAG) laser with conventional debridement in maintenance patients. MATERIAL & METHODS Fifteen patients, all smokers, having at least four teeth with residual probing depth (PD) ≥ 5 mm were recruited. Two pockets in two jaw quadrants were randomly assigned to subgingival debridement using an Er:YAG laser (test) or ultrasonic scaler/curette (control) at 3-month intervals. Relative attachment level (RAL), PD, bleeding on probing and dental plaque were recorded at baseline and at 6 and 12 months. Microbiological subgingival samples were taken at the same time points and analysed using a checkerboard DNA-DNA hybridization technique. RESULTS A significant decrease in PD took place in both treatments from baseline to 12 months (p < 0.01). In the control, the mean initial PD decreased from 5.4 to 4.0 mm at 12 months. For the test, a similar decrease occurred. No significant between-treatment differences were shown at any time point. The mean RAL showed no overall significant inter- or intra-treatment differences (p > 0.05). No significant between-treatment differences were observed in subgingival microbiological composition or total pathogens. CONCLUSION The results failed to support that an Er:YAG laser may be superior to conventional debridement in the treatment of smokers with recurring chronic inflammation. This appears to be the first time that repeated Er-YAG laser instrumentation has been compared with mechanical instrumentation of periodontal sites with recurring chronic inflammation over a clinically relevant time period.

Porphyromonas gingivalis: a clonal pathogen?: Diversities in housekeeping genes and the major fimbriae gene.

Abstract The introduction of multilocus sequence typing (MLST) in infectious disease research has allowed standardized typing of bacterial clones. Through multiple markers around the genome, it is possible to determine the sequence type (ST) of bacterial isolates to establish the population structure of a species. For the periodontal pathogen, Porphyromonas gingivalis, the MLST scheme has been established at www.pubmlst.org/pgingivalis, and data from the database indicate a high degree of genetic diversity and a weakly clonal population structure comparable with Neisseria menigitidis. The major fimbriae (FimA) have been held responsible for the adhesive properties of P. gingivalis and represent an important virulence factor. The fimA genotyping method (PCR based) indicate that fimA genotype II, IV and Ib are associated with diseased sites in periodontitis and tissue specimens from cardiovascular disease. fimA genotyping of the isolates in the MLST database supports the association of genotypes II and IV with periodontitis. As a result of multiple positive PCR reactions in the fimA genotyping, sequencing of the fimA gene revealed only minor nucleotide variation between isolates of the same and different genotypes, suggesting that the method should be redesigned or re-evaluated. Results from several investigations indicate a higher intraindividual heterogeneity of P. gingivalis than found earlier. Detection of multiple STs from one site in several patients with “refractory” periodontitis, showed allelic variation in two housekeeping genes indicating recombination between different clones within the periodontal pocket.

Human papillomavirus subtypes in oral lesions compared to healthy oral mucosa

BACKGROUND Human papillomaviruses (HPV) are involved in the etiology of cervix cancer, but it is still unclear whether they play a role in related oral lesions. OBJECTIVES The presence of HPV in oral leukoplakia biopsies (n=50) and oral squamous carcinoma biopsies (n=50) was compared to normal oral mucosa swabs (n=50) for the purpose of indicating a possible etiological role for the virus. STUDY DESIGN DNA was extracted from tissue biopsies and from mucosa swabs of control samples. Nested PCR was performed with primers targeting conserved sequences within the capsid gene L1. PCR products were sequenced to identify the HPV genotype. RESULT The results reveal a profile of low-risk HPV genotypes in oral leukoplakia similar to that in healthy controls, while HPV was less frequently observed in oral squamous carcinoma. CONCLUSIONS HPV does not seem to represent an important causal factor for the development of oral leukoplakia or oral squamous carcinoma.

Site-specific treatment outcome in smokers following non-surgical and surgical periodontal therapy.

Abstract Aim To evaluate the effect of smoking at patient, tooth, and site level following non‐surgical and surgical periodontal therapy. Material and Methods Eighty chronic periodontitis patients, 40 smokers and 40 non‐smokers, were recruited to this single‐arm clinical trial. Smoking status was validated by measuring serum cotinine levels. Periodontal examinations were performed at baseline (T0) and 3 months following non‐surgical and surgical periodontal therapy (T1). At T0 and T1, subgingival plaque samples were collected from the deepest periodontal pocket in each patient and analysed using checkerboard DNA–DNA hybridization. Probing depth (PD) ≥ 5 mm with bleeding on probing (BoP) was defined as the primary outcome. Unadjusted and adjusted logistic regression analyses, corrected for clustered observations within patients and teeth, were conducted comparing smokers with non‐smokers. Results Clinical parameters significantly improved in both groups (p < 0.001). An association was revealed between smoking and PD ≥ 5 mm with BoP (OR= 1.90, CI: 1.14, 3.15, p = 0.013), especially for plaque‐positive sites (OR= 4.14, CI: 2.16, 7.96, p < 0.001). A significant reduction of red complex microbiota was observed for non‐smokers only (p = 0.010). Conclusion Smokers respond less favourably to non‐surgical and surgical periodontal therapy compared with non‐smokers, in particular at plaque‐positive sites.

Influence of the Apical Preparation Size and the Irrigant Type on Bacterial Reduction in Root Canal–treated Teeth with Apical Periodontitis

Introduction: This clinical study evaluated the influence of the apical preparation size using nickel‐titanium rotary instrumentation and the effect of a disinfectant on bacterial reduction in root canal–treated teeth with apical periodontitis. Methods: Forty‐three teeth with posttreatment apical periodontitis were selected for retreatment. Teeth were randomly divided into 2 groups according to the irrigant used (2.5% sodium hypochlorite [NaOCl], n = 22; saline, n = 21). Canals were prepared with the Twisted File Adaptive (TFA) system (SybronEndo, Orange, CA). Bacteriological samples were taken before preparation (S1), after using the first instrument (S2), and then after the third instrument of the TFA system (S3). In the saline group, an additional sample was taken after final irrigation with 1% NaOCl (S4). DNA was extracted from the clinical samples and subjected to quantitative real‐time polymerase chain reaction to evaluate the levels of total bacteria and streptococci. Results: S1 from all teeth were positive for bacteria. Preparation to the first and third instruments from the TFA system showed a highly significant intracanal bacterial reduction regardless of the irrigant (P < .01). Apical enlargement to the third instrument caused a significantly higher decrease in bacterial counts than the first instrument (P < .01). Intergroup comparison revealed no significant difference between NaOCl and saline after the first instrument (P > .05). NaOCl was significantly better than saline after using the largest instrument in the series (P < .01). Conclusions: Irrespective of the type of irrigant, an increase in the apical preparation size significantly enhanced root canal disinfection. The disinfecting benefit of NaOCl over saline was significant at large apical preparation sizes. HighlightsThe influence of apical enlargement on bacterial reduction was evaluated.Irrigation was done with either sodium hypochlorite (NaOCl) or saline.The larger the apical preparation size, the greater the bacterial reduction.NaOCl was superior to saline only at large apical preparation sizes.