| Project/Area Number |
07557136
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| Research Category |
Grant-in-Aid for Scientific Research (A)
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| Allocation Type | Single-year Grants |
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| Section | 展開研究 |
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| Research Field |
矯正・小児・社会系歯学
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| Research Institution | KYUSHU UNIVERSITY |
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Principal Investigator |
YAMASHITA Yoshihisa KYUSHU UNIVERSITY,Faculty of Dentistry, Associate Professor, 歯学部, 助教授 (20192403)
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| Co-Investigator(Kenkyū-buntansha) |
YAMAGUCHI Noboru KYUSHU UNIVERSITY,Faculty of Dentistry, Research Associate, 歯学部, 助手 (00230368)
OHO Takahiko KYUSHU UNIVERSITY,Faculty of Dentistry, Assistant Professor, 歯学部, 講師 (50160940)
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| Project Period (FY) |
1995 – 1997
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| Project Status |
Completed (Fiscal Year 1997)
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| Budget Amount *help |
¥7,000,000 (Direct Cost: ¥7,000,000)
Fiscal Year 1997: ¥2,000,000 (Direct Cost: ¥2,000,000)
Fiscal Year 1996: ¥2,000,000 (Direct Cost: ¥2,000,000)
Fiscal Year 1995: ¥3,000,000 (Direct Cost: ¥3,000,000)
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| Keywords | Oral Diagnosis / Dental Diseases / Oral Pathogens / Gene Amplification / 遺伝子増幅法 / リスクファクター / 口膣病原性細菌 |
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| Research Abstract |
By using a colorimetric assay utilizing polymerase chain reaction (PCR), Actinobacillus actinomycetemcomitans, Porphyromonas gingivalis and Treponema denticola in heterogeneous mixed eubacterial suspension were quantitatively identified and also total cell number in the mixed suspension was quantified. The sets of specific primers for A.actinomycetemcomitans, P.gingivalis, and T.denticola were prepared on the basis of nucleotid sequences of the lktA,fim and atc genes, respectively. The set of universal primers for eubacteria was designed from nucleotide sequences of highly conserved region in eubacterial 16S rRNA sequences. A known amount of non-bacterial DNA fragment which was artificially synthesized and the set of the corresponding primers were added to the PCR mixtures the amplified DNA fragment was used as an internal standard. The colorimetric enhacement from each target DNA was compensated as compared with that from internal standard. The bacterial cell numbers between 10^2 and 10^6 cells were able to be quantitatively estimated by this colorimetric PCR assay. Clinical plaque samples from patients with periodontal diseases were evaluated with this assay. More than 2*10^6 cells of total bacteria were estimated to be present in subgingival plaque samples from all diseased sites. On the contrary, less than 2*10^6 cells of total bacteria were estimated in those from more than half of healthy sites, suggesting that subgingival plaque in diseased site consists of relatively large number of bacteria compared with that in healthy site. While A.actinomycetemcomitans was detected in both the healthy and diseased sites, P.gingivalis and T.denticola were observed only in diseased site. These periodontopathic bacteria were estimated to occupy the minor portin (less than 0.01%) of total subgingival plaque bacteria. Now we developing the same kind of diagnosis method for dental caries evaluating Streptococcus mutans and Streptococcus sobrims.
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