Avaliação da ocorrência de periodontopatógenos em sítios periodontais de pacientes com e sem doença periodontal por cultura microbiológica e por reação de polimerização em cadeia

Abstract

Black-pigmented anaerobic rods, Fusobacterium nucleatum, Actinobacillus actinomycetemcomitans, Eikenella corrodens, Tannerella forsythensis and Treponema denticola are putative periodontopathogens indigenous to the oral cavity, which frequency and proportion in healthy and diseased periodontal sites may exhibit regional differences. The purpose of this study was to evaluate, by employing microbiological culture and PCR, the presence of these microorganisms in periodontal sites of 30 patients with chronic periodontitis, 30 patients with aggressive periodontitis and 30 subjects with healthy periodontium. The samples were collected with paper points and transferred to Ringer PRAS solution, under CO2 flux. The material was homogenized, introduced into an anaerobic chamber and, following serial 10-fold dilutions up to 10-4, it was cultured onto selective media and incubated at 37ºC for up to 15 days. Bacterial identification was based on macroscopic, microscopic and biochemical-physiological characteristics. Also, part of the clinical specimen was centrifuged and the sediment used for DNA extraction by the phenol-chloroform method. Specific primers that targeted the 16S rDNA were employed for PCR amplification. The results of the microbiological culture have shown associations between P. intermedia/nigrescens, F. nucleatum, F. varium and chronic periodontitis, and between P. intermedia/nigrescens, A. actinomycetemcomitans and aggressive periodontitis. The PCR results have indicated that P. intermedia, P. nigrescens, Fusobacterium spp., T. forsythensis and T. denticola were statistically associated to the chronic and the aggressive forms of periodontitis, whereas E. corrodens was shown to be associated only to chronic periodontitis. PCR has been shown to be more sensitive for the detection of most of the investigated microorganisms. The antagonistic or synergistic associations observed among the bacterial samples isolated from patients with aggressive periodontitis, differ from those found in patients with chronic periodontitis, which reinforces the importance of those associations in periodontal disease expression.