Polymerase chain reaction for the identification of Porphyromonas gingivalis collagenase genes

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Sequence variations in rgpA and rgpB of Porphyromonas gingivalis in periodontitis

Objective: The aim of the present study was to determine sequence variations in the active centre of the Arg-X-specific protease encoding genes rgpA and rgpB of clinical Porphyromonas gingivalis isolates and to analyse their prevalence in periodontitis patients before and 3 months after mechanical periodontal therapy. Background: Genetic diversity at nucleotides 281, 283, 286 and 331 has been shown to result in amino acid substitutions in the catalytic domain of RgpA and RgpB that affect the substrate specificity and thus may influence the efficacy of Arg-X-protease specific inhibitors. Methods: Sequence analysis of rgpA and rgpB genes in clinical P. gingivalis strains isolated from subgingival plaque samples of 82 periodontitis patients before and 3 months after mechanical supra- and subgingival debridement was performed. Results: No specific variation within the rgpA sequence was observed. However, the rgpB sequence in the region of the active centre showed five different rgpB genotypes, which were named NYPN, NSSN, NSSK, NYPK and DYPN according to the derived amino acid substitution. Porphyromonas gingivalis genotype NYPN was detected in 27 patients (32.9%) before and in 8 patients (9.8%) after therapy, NSSN in 26 (31.7%) and 10 (12.2%), NSSK in 22 (26.8%) and 2 (2.4%), NYPK in 5 (6.2%) and 1 (1.2%), and DYPN in 1 patient (1.2%) and 0 patients (0%), respectively. Only one patient (1.2%) harboured two P. gingivalis rgpB genotypes (NSSK/NYPN) before treatment; these were no longer detected after therapy. Conclusion: The results indicate that five rgpB genotypes are maintained in natural populations of P. gingivalis. These data may be of importance with regard to the development of specific rgpB inhibitors. © Blackwell Munksgaard 2005.link_to_subscribed_fulltex

Differential clinical treatment outcome after systemic metronidazole and amoxicillin in patients harboring Actinobacillus actinomycetemcomitans and/or Porphyromonas gingivalis

48 adult patients with untreated periodontitis harboring subgingival Actinobacillus actinomycetemcomitans and/or Porphyromonas gingivalis as assessed by PCR were randomly assigned to receive full-mouth scaling alone (control) or scaling with systemic metronidazole plus amoxicillin and supragingival irrigation with chlorhexidine digluconate (test). In patients harboring A. actinomycetemcomitans intraorally at baseline, the adjunctive antimicrobial therapy resulted in a significantly higher incidence of probing attachment level (PAL) gain of 2 mm or more compared to scaling alone over 12 months (p<0.05). In addition, suppression of A. actinomycetemcomitans in subgingival plaque below detectable levels was associated with an increased incidence of PAL gain. In contrast, patients initially harboring P. gingivalis but not A. actinomycetemcomitans in the oral cavity showed a significantly higher incidence of PAL loss following adjunctive antimicrobial therapy compared to scaling alone (p<0.05). When the presence of pathogens at baseline was disregarded in the analysis, adjunctive antimicrobial therapy did not significantly enhance clinical treatment outcome. The results indicated that adults with untreated periodontitis harboring A. actinomycetemcomitans may benefit from the adjunctive antimicrobial therapy for a minimum of 12 months, whereas, the regimen may adversely affect the clinical treatment outcome of patients harboring P. gingivalis but not A. actinomycetemcomitans. © Munksgaard, 1998.link_to_subscribed_fulltex

Association between infection of different strains of Porphyromonas gingivalis and Actinobacillus actinomycetemcomitans in subgingival plaque and clinical parameters in chronic periodontitis

Objective: The aim of this study was to investigate subgingival infection frequencies of Porphyromonas gingivalis and Actinobacillus actinomycetemcomitans strains with genetic variation in Chinese chronic periodontitis (CP) patients and to evaluate its correlation with clinical parameters. Methods: Two multiplex polymerase chain reaction (PCR) assays were developed to detect the 16SrDNA, collagenase (prtC) and fimbria (fimA) genes of P. gingivalis and the 16SrDNA, leukotoxin (lktA) and fimbria-associated protein (fap) genes of A. actinomycetemcomitans in 60 sulcus samples from 30 periodontal healthy subjects and in 122 subgingival plaque samples from 61 patients with CP. The PCR products were further T-A cloned and sent for nucleotide sequence analysis. Results: The 16SrDNA, prtC and fimA genes of P. gingivalis were detected in 92.6%, 85.2% and 80.3% of the subgingival plaque samples respectively, while the 16SrDNA, lktA and fap genes of A. actinomycetemcomitans were in 84.4%, 75.4% and 50.0% respectively. Nucleotide sequence analysis showed 98.62%~100% homology of the PCR products in these genes with the reported sequences. P. gingivalis strains with prtC+/fimA+ and A. actinomycetemcomitans with lktA+ were predominant in deep pockets (>6 mm) or in sites with attachment loss ≥5 mm than in shallow pockets (3~4 mm) or in sites with attachment loss ≤2 mm (P<0.05). P. gingivalis strains with prtC+/fimA+ also showed higher frequency in gingival index (GI)=3 than in GI=1 group (P<0.05). Conclusion: Infection of P. gingivalis with prtC+/fimA+ and A. actinomycetemcomitans with lktA+ correlates with periodontal destruction of CP in Chinese. Nonetheless P. gingivalis fimA, prtC genes and A. actinomycetemcomitans lktA gene are closely associated with periodontal destruction, while A. actinomycetemcomitans fap gene is not