Denaturing gradient gel electrophoresis analysis to study bacterial community structure in pockets of periodontitis patients

Abstract

Bacteria are involved in the onset and progression of periodontitis. A promising molecular technique, denaturing gradient gel electrophoresis (DGGE), to study microbial population dynamics in the subgingival pocket is presented. Twenty-three samples were taken from the subgingival pockets of nine patients and six healthy family members. From four periodontitis patients, 12 samples were evaluated before, 1 day after and 3 months after treatment. Part of the 16S rRNA gene of all bacteria was amplified by PCR and separated by DGGE, creating banding patterns representative of the community structure. Shifts in composition and diversity of the microbial population could be determined semiquantitatively, and this showed that treatment resulted in a decrease in the diversity of the population. After 3 months a microbial population 33-47% different from the population before treatment had re-established. Intense bands representing Exiguobacterium aurantiacum were present in 13 out of 25 samples, indicating that this species may play a role in periodontal disease.

Interspecies Interactions within Oral Microbial Communities

“…It was used to conduct surveys of microbial diversity in human saliva (112,165) and to study oral bacterial community structure associated with severe dental caries (114), endodontic infections (124,182,183), and periodontitis (55,217), as well as to evaluate the oral microbial changes after prophylactic treatments (113). Due to its broad-range nature, this method facilitates the detection of previously unknown fastidious or uncultivable microbial species (177), and it is less labor-intensive than the traditional cloning methods.…”

“…DGGE has also proven to be a useful tool for analyzing the microbiota in the human oral cavity (112,113,165,217). It was used to conduct surveys of microbial diversity in human saliva (112,165) and to study oral bacterial community structure associated with severe dental caries (114), endodontic infections (124,182,183), and periodontitis (55,217), as well as to evaluate the oral microbial changes after prophylactic treatments (113).…”

Biofilm formation of salivary microbiota on dental restorative materials analyzed by denaturing gradient gel electrophoresis and sequencing

“…DGGE has been used in many fields to analyze the composition and diversity of microbial communities in different niches 21,22) . PCR-DGGE is also extensively employed to analyze oral microbial communities [9][10][11][12][13]16,23) . However, there are no studies in published literature that have used DGGE to examine microbial compositions detected on dental materials.…”

“…When doublestranded DNA molecules are run in polyacrylamide gel containing a linear gradient of DNA denaturants (urea and formamide), DNA fragments of the same length but with different base-pair sequences will stop migrating at different positions in the gel 8) . DGGE has been widely employed to examine the microbial populations found in subgingival plaque [9][10][11] , endodontic infections 12) , and dental plaque microbiome. It has been proposed that DGGE could be a useful tool for identifying the profiles of microbial pathogens implicated in periodontal diseases and caries, which then aids in the diagnosis and risk assessment of these patients 13) .…”

Use of denaturing gradient gel electrophoresis for the identification of mixed oral yeasts in human saliva

“…The use of PCR together with denaturing gradient gel electrophoresis (DGGE) as described here enables detection of the presence of such species and frequently their presumptive identification, even if they are present as minor populations. DGGE based on 16S and 26-28S rRNA gene sequences has been used to characterize complex microbial communities, including those in saliva, dental plaque and plaque microcosms (Ledder et al, 2006;Li et al, 2006;Rasiah et al, 2005;Zijnge et al, 2003). It has also been applied to monitor fungal communities in complex ecosystems such as soil, and in wine and food fermentations (Cocolin et al, 2000;Meroth et al, 2003;Muccilli et al, 2011;Oros-Sichler et al, 2006;Vilela et al, 2010).…”