Quantitative detection of periodontal pathogens using real‐time polymerase chain reaction with TaqMan probes

Kuboniwa, Masae; Amano, Atsuo; Kimura, K. R.; Sekine, Shinichi; Kato, Shingo; Yamamoto, Yumiko; Okahashi, Nobuo; Iida, Tetsuya; Shizukuishi, Satoshi

doi:10.1111/j.0902-0055.2004.00135.x

Cited by 160 publications

(139 citation statements)

References 68 publications

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“…Numerous methods applying molecular biology techniques which can overcome this culture bias have been developed and used for bacterial detection. A number of these allow the qualitative or quantitative detection of single bacterial species (13), while others can detect and profile whole heterogeneous populations (25). In order to more fully study the complex oral microbiota, techniques such as denaturing gradient gel electrophoresis (DGGE) can be used (8,9,15,16,26).…”

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confidence: 99%

Drinking Habits Are Associated with Changes in the Dental Plaque Microbial Community

et al. 2010

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Caries and gingivitis are the most prevalent oral infectious diseases of humans and are due to the accumulation of dental plaque (a microbial biofilm) on the tooth surface and at the gingival margin, respectively. Several in vitro and in vivo studies have shown that many natural components of foods and beverages inhibit the adhesion of and/or exert activity against oral bacteria. These biological activities have mainly been attributed to the polyphenol fraction. In order to explore the possibility that diet can alter the dental plaque community, in this study we evaluated the composition of the microbiota of supra-and subgingival plaque samples collected from 75 adult subjects with different drinking habits (drinkers of coffee, red wine, or water for at least 2 years) by analyzing the microbial population through the separation of PCR-amplified fragments using the denaturing gradient gel electrophoresis (DGGE) technique. The mean numbers of bands of the DGGE profiles from all three categories were evaluated. There were no significant differences between the two kinds of plaque collected from the control group (water drinkers), and this group showed the highest number of bands (supragingival plaque, 18.98 ؎ 3.16 bands; subgingival plaque, 18.7 ؎ 3.23 bands). The coffee and wine drinker groups generated the lowest numbers of bands for both supragingival plaque (coffee drinkers, 8.25 ؎ 3.53 bands; wine drinkers, 7.93 ؎ 2.55 bands) and subgingival plaque (coffee drinkers, 8.3 ؎ 3.03 bands; wine drinkers, 7.65 ؎ 1.68 bands). The differences between coffee drinkers or wine drinkers and the control group (water drinkers) were statistically significant. A total of 34 microorganisms were identified, and the frequency of their distribution in the three subject categories was analyzed. A greater percentage of subjects were positive for facultative aerobes when supragingival plaque was analyzed, while anaerobes were more frequent in subgingival plaque samples. It is noteworthy that the frequency of identification of anaerobes was significantly reduced when the frequencies for coffee and wine drinkers were compared with the frequencies for subjects in the control group. The DGGE profiles of the organisms in both plaque samples from all groups were generated and were used to construct dendrograms. A number of distinct clusters of organisms from water, coffee, and wine drinkers were formed. The clustering of some of the DGGE results into cohort-specific clusters implies similarities in the microbiotas within these groups and relevant differences in the microbiotas between cohorts. This supports the notion that the drinking habits of the subjects may influence the microbiota at both the supragingival and the subgingival levels.

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confidence: 99%

Drinking Habits Are Associated with Changes in the Dental Plaque Microbial Community

et al. 2010

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“…Modern molecular techniques, particularly those based on amplifying 16S rRNA genes, include cloning and sequencing, quantitative DNA-DNA hybridization (checkerboard), denaturing gradient gel electrophoresis (DGGE), and real-time PCR. These methods have superseded conventional culture-based methods for quantifying the microbiota of healthy and diseased oral cavity (9,12,13,28). But to date, the implementation of these molecular techniques is still restricted to central laboratories because of the high equipment cost and the requirement for highly trained personnel.…”

Section: Methodsmentioning

confidence: 99%

<b>PCR-dipstick DNA chromatography for profiling of a subgroup of cariesassociated bacterial species in plaque from healthy coronal surfaces and periodontal </b><b>pockets </b>

et al. 2016

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The onset of plaque-mediated disease, including dental caries and periodontal diseases, is highly associated with compositional change of the resident microflora from the ecological perspective. As specific bacterial profiles have been linked to different disease stages, microbial compositional measurements might therefore have great value for clinical diagnosis. Previously we have reported a dry-reagent strip biosensor-PCR-dipstick DNA chromatography, which utilized molecular recognition of oligonucleotides and biotin-streptavidin, and the optical property of colored microspheres, for semiquantifying a five-membered subgroup of caries-associated bacterial species in supragingival plaque from healthy coronal surfaces of teeth. The present study aimed to evaluate this technique's ability to differentiate microflora by comparing the subset profiles. Sixteen subgingival plaque specimens were pooled from periodontal pockets and analyzed for the composition of Streptococcus mutans, Streptococcus sobrinus, Scardovia wiggsiae, Actinomyces sp. and Veillonella parvula. Detection frequencies, relative abundance of each bacterial species, and the five-membered bacterial profiles were compared between supra-and subgingival groups. The supragingival plaque harbored significantly more of the tested species and higher amount of Actinomyces sp. and V. parvula. In subgingival plaque, the predominance was obscured, since several highly overlapped profiles were found at comparable frequencies. Thus, PCR-dipstick DNA chromatography using the same plaque sample enabled simultaneous profiling of multiple species at species level and facilitated discrimination between anticipated different microflora, making this technique a promising chair-side microbiota profiling method.The ecological plaque hypothesis (18) has revolutionized our view of the etiology of the most prevalent oral diseases, including caries and periodontal diseases. Contrary to the classical medical model, which ascribes an infection to a single microbial pathogene, the new hypothesis suggests that a dysbiosis of the resident oral microflora, driven by an ecological perturbation, leads to plaque-mediated disease. During the transition from oral health to disease state, a dynamic interaction between the resident microflora and the habitat has been evidenced, with numerous bacterial species being implicated (2,3,22,31). A substantial change of the environmental determinants could disrupt the homeostasis of the resident microflora, thereby inducing the prolifera-

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“…24,25 In combination with such specimens, we normally use saliva as a PCR template that reflects the overall condition of the oral cavity. In fact, bacteria causing dental caries and periodontitis are more frequently detected in saliva than in the dental plaque.…”

Section: Development Of Taqman Real-time Pcr For the Detection Of Oramentioning

confidence: 99%

Quantitative Analysis of Multi-Species Oral Biofilms by TaqMan Real-Time PCR

Suzuki

Yoshida²,

Nakano³

2005

Clinical Medicine & Research

109

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Oral infectious diseases, including dental caries, various forms of periodontitis and oral malodor, are not caused by a single pathogen.The etiology of these diseases is known to be associated with bacterial accumulation and plaque composition on the hard and soft tissues of the oral cavity. Therefore, the quantitative, as well as qualitative, analysis of the microorganisms present in oral biofilms, namely dental plaque, subgingival plaque and tongue debris, is important for diagnosis and rational treatment decisions.The quantitative microbial analysis of oral multi-species biofilms also provides useful information for establishing the etiology of oral infectious diseases. Recently, a 5 ′ fluorogenic, nuclease-based, real-time polymerase chain reaction (PCR) technique has been increasingly employed for the quantitative microbial assessment of the human oral cavity. We review the development and use of TaqMan real-time PCR for quantifying oral bacteria, its role in the diagnosis of oral infectious diseases and their microbial etiology.

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Quantitative detection of periodontal pathogens using real‐time polymerase chain reaction with TaqMan probes

Cited by 160 publications

References 68 publications

Drinking Habits Are Associated with Changes in the Dental Plaque Microbial Community

Drinking Habits Are Associated with Changes in the Dental Plaque Microbial Community

<b>PCR-dipstick DNA chromatography for profiling of a subgroup of cariesassociated bacterial species in plaque from healthy coronal surfaces and periodontal </b><b>pockets </b>

Quantitative Analysis of Multi-Species Oral Biofilms by TaqMan Real-Time PCR

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