Quantitative PCR analysis of salivary pathogen burden in periodontitis

Abstract

Our aim was to investigate the value of salivary concentrations of four major periodontal pathogens and their combination in diagnostics of periodontitis. The Parogene study included 462 dentate subjects (mean age 62.9 ± 9.2 years) with coronary artery disease (CAD) diagnosis who underwent an extensive clinical and radiographic oral examination. Salivary levels of four major periodontal bacteria were measured by quantitative real-time PCR (qPCR). Median salivary concentrations of Porphyromonas gingivalis, Tannerella forsythia, and Prevotella intermedia, as well as the sum of the concentrations of the four bacteria, were higher in subjects with moderate to severe periodontitis compared to subjects with no to mild periodontitis. Median salivary Aggregatibacter actinomycetemcomitans concentrations did not differ significantly between the subjects with no to mild periodontitis and subjects with moderate to severe periodontitis. In logistic regression analysis adjusted for age, gender, diabetes, and the number of teeth and implants, high salivary concentrations of P. gingivalis, T. forsythia, and P. intermedia were significantly associated with moderate to severe periodontitis. When looking at different clinical and radiographic parameters of periodontitis, high concentrations of P. gingivalis and T. forsythia were significantly associated with the number of 4–5 mm periodontal pockets, ≥6 mm pockets, and alveolar bone loss (ABL). High level of T. forsythia was associated also with bleeding on probing (BOP). The combination of the four bacteria, i.e., the bacterial burden index, was associated with moderate to severe periodontitis with an odds ratio (OR) of 2.40 (95% CI 1.39–4.13). When A. actinomycetemcomitans was excluded from the combination of the bacteria, the OR was improved to 2.61 (95% CI 1.51–4.52). The highest OR 3.59 (95% CI 1.94–6.63) was achieved when P. intermedia was further excluded from the combination and only the levels of P. gingivalis and T. forsythia were used. Salivary diagnostics of periodontitis has potential especially in large-scale population studies and health promotion. The cumulative strategy appears to be useful in the analysis of salivary bacteria as markers of periodontitis.

Comparison of quantitative detection of periodontal pathogens before and after scaling by real-time polymerase chain reaction

“…In this study, we used real-time PCR assays, which are rapid and sensitive techniques for detecting or quantifying microorganisms and for identifying genes or mutations in pathogens associated with antimicrobial resistance. New advances in real-time PCR in clinics will insure better management of patients, and could impact the spread of antibiotic resistance as reported in several studies [22][23][24]26) . The presence of systemic humoral and cellular reactivity to a variety of oral microorganisms has been found in subjects with different forms of periodontal disease [6][7][8][9] .…”

“…New advances in real-time PCR in clinics will insure better management of patients, and could impact the spread of antibiotic resistance as reported in several studies [22][23][24]26) . The presence of systemic humoral and cellular reactivity to a variety of oral microorganisms has been found in subjects with different forms of periodontal disease [6][7][8][9] . Some investigations have shown the efficacy of scaling in the treatment of gingivities and periodontitis [17][18][19][20][21] .…”

“…P. gingivalis and T. forsythia are strong markers of periodontitis in adults, and these species have been linked to the progression of the disease 8,9) . The dental plaque presence of members of bacterial species that constitute the red complex, such as P. gingivalis, T. denticola, and T. forsythia, has been associated with advanced periodontitis and perimplantitis.…”

“…Quantification of bacteria is the preferred technique to achieve this task nowadays, and real-time polymerase chain reaction(PCR) that relies on real-time detection of the levels of fluorescence generated within the reaction, is probably the most accurate technology 22,23) . Kim et al 24) reported the prevalence of periodontopathic bacteria in 8 to 18 year old subjects using real-time PCR.…”

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“…Over the age of 30 approximately 50% of people has some form of gingivitis 19 . The periodontal progression continues by the transformation of gram positive followed by dominance of gram negative bacteria 20 . The widely known periodontal pathogens present in plaque includes aerobic and anaerobic bacteria like Streptococci species, P.gingivalis, Prevotella intermedia, Treponema denticola, T. forsythia, Campylobacter rectus, Selenomonas spp, A.a, Eubacterium timidum, Fusobacterium nucleatum and Parvimonas micra 2 .…”