Periodontal ligament and gingival fibroblasts from periodontitis patients are more active in interaction with Porphyromonas gingivalis.

BACKGROUND AND OBJECTIVE: Inflammatory responses of host cells to oral pathogenic bacteria, such as Porphyromonas gingivalis, are crucial in the development of periodontitis. Host cells, such as periodontal ligament and gingival fibroblasts, from periodontitis patients may respond to P. gingivalis in a different manner compared with cells from healthy persons. The aim of this study was to investigate inflammatory responses to viable P. gingivalis by periodontal ligament and gingival fibroblasts from periodontitis patients and healthy control subjects. MATERIAL AND METHODS: Primary periodontal ligament and gingival fibroblasts from periodontitis patients (n=14) and healthy control subjects (n=8) were challenged in vitro with viable P. gingivalis. Gene expression of Toll-like receptors (TLRs) 1, 2, 4, 6, 7 and 9, CD14, nuclear factor-κB1 and its putative inhibitor NF-κB inhibitor-like protein1, and of interleukin-1ß, interleukin-6, interleukin-8, tumour necrosis factor-α, monocyte chemotactic protein-1 and regulated upon activation, normal T-cel expressed, and secreted, were assessed by real-time PCR. RESULTS: Periodontal ligament fibroblasts from periodontitis patients had a higher mRNA expression of TLR1, TLR4, TLR7 and CD14, and a lower expression of NFKBIL1, both before and after P. gingivalis challenge. In contrast, gingival fibroblasts from periodontitis patients had stronger induction of TLR1, TLR2 and TLR7 by P. gingivalis. Cytokine responses were not different between patients and control subjects. Interestingly, periodontal ligament, but not gingival, fibroblasts from P. gingivalis culture-positive persons responded more strongly to P. gingivalis than periodontal ligament fibroblasts from P. gingivalis-negative persons. CONCLUSION: Periodontal ligament and gingival fibroblasts respond to P. gingivalis in a different manner and may play different roles in periodontitis. Both subsets of fibroblasts from patients appear more active in interaction with P. gingivalis. Moreover, periodontal ligament fibroblasts from P. gingivalis-positive donors are more responsive to an in vitro P. gingivalis challenge.

Comparison of periodontal pathogens between cats and their owners.

The periodontal pathogens Porphyromonas gingivalis and Tannerella forsythia are strongly associated with periodontal disease and are highly prevalent in humans with periodontitis. Porphyromonas and Tannerella spp. have also been isolated from the oral cavity of cats. The oral microflora in animals was compared with those in humans in earlier studies, but no studies are available on the comparison of the oral microflora from pets and their respective owners. The aim of this study was to determine the presence of these bacteria in the oral microflora of cats and their owners, since animal to human transmission, or vice versa, of oral pathogens could have public health implications. This study investigated the prevalence of Porphyromonas gulae, P. gingivalis, and T. forsythia in the oral microflora of cats and their owners, using culture and polymerase chain reaction (PCR). All Porphyromonas isolates from cats (n=64) were catalase positive, whereas the Porphyromonas isolates from owners (n=7) were catalase negative, suggesting that the isolates from cats were P. gulae whereas those from the owners were P. gingivalis. T. forsythia was recovered from both cats (n=63) and owners (n=31); the proportion of T. forsythia relative to the total CFU was higher in cats with periodontitis than in cats without periodontal disease. Genotyping of T. forsythia isolates (n=54) in six cat/owner couples showed that in one cat/owner couple the T. forsythia isolates (n=6) were identical. These T. forsythia isolates were all catalase positive, which led us to hypothesize that transmission from cats to owners had occurred and that cats may be a reservoir of T. forsythia.

Smoking affects the subgingival microflora in periodontitis.

BACKGROUND: Tobacco smoking has been identified as one major risk factor for destructive periodontal disease. Scaling and root planing have been shown to be less effective in smokers with periodontitis. The aim of the present study was to compare the subgingival microbial flora of treated and untreated smokers and non-smokers. METHODS: Four independent adult patient groups with periodontitis were included in this investigation: 88 untreated smokers (U-S); 90 untreated non-smokers (U-NS); 119 treated non-smokers (T-NS); and 171 treated smokers (T-S). Clinical variables included cumulative plaque index (CPI), probing depth (PD), clinical attachment level (CAL), cumulative bleeding index (CBI), and cumulative suppuration index (CSI). Paper point samples from the deepest bleeding pocket in each quadrant of the dentition were analyzed for the presence and levels of 6 periodontal bacterial pathogens using anaerobic culture techniques. RESULTS: U-S showed a higher mean cumulative plaque index than U-NS (3.5 versus 2.7). Mean PD and mean CAL were higher in the T-S in comparison to the T-NS group (7.0 versus 6.6 mm and 5.6 versus 4.7 mm, respectively). Microbiological characteristics of U-S were a higher prevalence of Prevotella intermedia/nigrescens and higher mean levels of Peptostreptococcus micros (Pm) and Fusobacterium nucleatum (Fn). T-S patients were characterized by higher prevalence of Bacteroides forsythus (Bf), Pm, and Campylobacter rectus (Cr) and higher mean levels of Pm and Fn. The mean percentage of B. forsythus tended to be higher in the T-S group than in the T-NS group (6.9% versus 5.6%). The relative risk to be infected with Bf, Pm, and Cr was statistically higher in smokers (odds ratios: 1.9, 1.9, and 1.6, respectively). The chance to find > or =10% of Bf, Pm, and/or Fn was 3.3 higher in smokers when A. actinomycetemcomitans and P gingivalis were absent. Detection of > or =20% Pm/Fn in treated patients was strongly associated with smoking (odds ratio 13.8, P= 0.002). CONCLUSIONS: Smoking is a determining factor for the composition of the subgingival microflora in adult patients with periodontitis and may select for a specific cluster of periodontal pathogens, notably Bf, Pm, Fn, and Cr. On the basis of these observations, smoking, among other criteria, may be one parameter to use in deciding to treat refractory periodontitis in smokers with a systemic antibiotic therapy directed against smoking-associated periodontal bacteria.

Porphyromonas gingivalis in an edentulous proband. A case-report.

In a previous study, an edentulous subject showed the presence of P. gingivalis, only in saliva. The present case report is a longitudinal follow-up on this proband (female, 63 years) and her spouse with periodontitis (67 years). Samples were taken from the proband and the spouse for microbiological analysis at several occasions during a 3-year period and a 1-year period respectively. In the edentulous proband, P. gingivalis was present in saliva only, at all times. In the spouse, P. gingivalis was present in the periodontal pocket and on the mucous membranes, but the saliva was culture-negative for this species at both sampling times. The restriction enzyme analysis (REA) patterns of isolates (n=7) of P. gingivalis from the proband recovered over 3 years showed no differences. This indicated that the same clonal type of P. gingivalis persisted for 3 years in the proband. The P. gingivalis isolates (n=13) from the spouse showed no differences when REA patterns were compared. Comparison of REA patterns from the proband and the spouse also showed no differences. Thus the same clonal type of P. gingivalis can persist over several years in an edentulous subject and at least for 1 year in a subject with periodontal disease.

Intra-familial transmission and distribution of Prevotella intermedia and Prevotella nigrescens.

The periodontal bacteria Prevotella intermedia and Prevotella nigrescens have been recently separated from each other. The purpose of this study was to investigate the distribution and routes of transmission of these bacteria among family members. Seven patients with moderate to severe periodontitis were selected. These probands, their spouses and 14 of their children were investigated. The presence of Pr. intermedia and Pr. nigrescens was determined by culture techniques in pooled subgingival plaque samples, in the saliva, on the tongue, tonsils and buccal mucosa. Differentiation of Pr. intermedia and Pr. nigrescens was performed by enzyme electrophoretic mobility. From all 7 patients, as well as 4 spouses and 3 of the children, Pr. intermedia could be isolated. Pr. nigrescens was found in 2 of the 7 patients, in 5 of the spouses and in 5 of the 6 children aged 5-10 yr. In the 8 children aged 0-4 yr both species were seldom isolated. These data are in accordance with earlier findings that Pr. intermedia is associated with periodontitis and Pr. nigrescens with a relatively healthy periodontal condition. Ribotyping of bacteria was performed by hybridization of HindIII restriction endonuclease digests of chromosomal DNA with ribosomal DNA. Isolates from unrelated individuals always had distinct ribotypes. Indistinguishable ribotypes of Pr. intermedia and Pr. nigrescens were found both among married couples and among parents and children. This indicates that intrafamilial transmission of Pr. intermedia and Pr. nigrescens is possible both between adults and between parents and children.

Effect of antibodies on chemiluminescence and on killing of Streptococcus sobrinus by polymorphonuclear leukocytes.

The effect of a polyclonal antiserum and OMVU10, a monoclonal antibody reactive with Antigen B of Streptococcus sobrinus, on the interaction of polymorphonuclear leukocytes with S. sobrinus was studied, using chemiluminescence and bacterial killing assays. Increased stimulation of neutrophils as measured in the chemiluminescence assays was established when S. sobrinus was preincubated with polyclonal antiserum or when polyclonal antiserum was added to the reaction mixture. Higher counts were measured in comparison to preimmune serum. After 90 min, 52% of S. sobrinus preincubated with polyclonal antiserum was killed. Killing was also increased when polyclonal antiserum was added to the reaction mixture in comparison to the controls. No killing was found when bacteria were preincubated with OMVU10 or when OMVU10 was added to the reaction mixture in comparison to Clone 24, a control antibody.

Comparison of six typing methods for Actinobacillus actinomycetemcomitans.

Actinobacillus actinomycetemcomitans is an important pathogen in the etiology of severe periodontitis. For epidemiological studies on the prevalence of certain pathogenic clones and transmission of this bacterium, adequate typing methods are necessary. The purpose of this study was to compare six different typing methods for A. actinomycetemcomitans. Five reference strains and 27 fresh clinical isolates from periodontitis patients were used. Serotyping showed 12 serotype a strains, 13 type b strains, 6 type c strains, and 1 nontypeable strain. Biotyping on the basis of the fermentation of mannose, mannitol, and xylose resulted in six biotypes. Antibiogram typing was evaluated by measuring the inhibition zones of seven antibiotics in agar diffusion tests. With this method eight main types which could be further differentiated into 15 subtypes were found. Sodium dodecyl sulfate-polyacrylamide gel electrophoresis patterns of outer membrane proteins were similar among all isolates tested. Restriction endonuclease analysis (REA) of whole chromosomal DNA resulted in five main types. These five main types were further differentiated into 24 subtypes on the basis of DNA fragment differences in the high-molecular-weight region. Hybridization of DNA fragments with ribosomal DNA (ribotyping) resulted in 22 to 24 different types, depending on the restriction endonuclease used. Ribotype patterns were easy to interpret and provided an univocal distinction between different strains compared with REA results. When applied to epidemiologically related isolates, all methods were able to discriminate two clonal types among five isolates from five children from one family. We conclude that serotyping, biotyping, and outer membrane patterns were reproducible but had a low discriminatory potential. REA and ribotyping were reproducible and gave the highest number of distinct types. When the DNA typing methodis were compared, all strains tested could be distinguished. These findings confirm the heterogeneity found within the species A. actinomycetemcomitans.