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Development of Oral Diagnosis Using Rapid and Simple Quantitative Gene Amplification of Oral Pathogens

Research Project

Project/Area Number 07557136
Research Category

Grant-in-Aid for Scientific Research (A)

Allocation TypeSingle-year Grants
Section展開研究
Research Field 矯正・小児・社会系歯学
Research InstitutionKYUSHU UNIVERSITY

Principal Investigator

YAMASHITA Yoshihisa  KYUSHU UNIVERSITY,Faculty of Dentistry, Associate Professor, 歯学部, 助教授 (20192403)

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)
Project Period (FY) 1995 – 1997
Project Status Completed (Fiscal Year 1997)
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)
KeywordsOral 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
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.

Report

(4 results)
  • 1997 Annual Research Report   Final Research Report Summary
  • 1996 Annual Research Report
  • 1995 Annual Research Report
  • Research Products

    (23 results)

All Other

All Publications (23 results)

  • [Publications] Y.Sumi et al.: "Semiquantitative analysis of periodontopathogens by gene amplification" Japanese Journal of Medical Science and Biology. 48. 177-191 (1995)

    • Description
      「研究成果報告書概要(和文)」より
    • Related Report
      1997 Final Research Report Summary
  • [Publications] N.Yamaguchi et al.: "Role of capsular-like scrotype-specific polysaccharide antigen in resistance of Actinobacillus actinomycetemcomitans to phagocytosis by human polymorphonuclear leukocytes" Infection and Immunity. 63. 4589-4594 (1995)

    • Description
      「研究成果報告書概要(和文)」より
    • Related Report
      1997 Final Research Report Summary
  • [Publications] N.Takamatsu-Matsushita et al.: "Immunogenicity of Actinobacillus actinomycetemcomitans serotype b-specific polysaccharide-protein conjugate" Oral Microbiology and Immunology. 11. 220-225 (1996)

    • Description
      「研究成果報告書概要(和文)」より
    • Related Report
      1997 Final Research Report Summary
  • [Publications] Y.Nakano et al.: "New shuttle vectors for Actinobacillus actinomycetemcomitans and Escherichia coli" Gene. 169. 139-140 (1996)

    • Description
      「研究成果報告書概要(和文)」より
    • Related Report
      1997 Final Research Report Summary
  • [Publications] N.Yamaguchi et al.: "Actinobacillus actinomycetemcomitans serotype b-specific polysaccharide antigen stimulates production of chemotactic factors and inflammatory cytokines by human monocytes" Infection and Immunity. 64. 2563-2570 (1996)

    • Description
      「研究成果報告書概要(和文)」より
    • Related Report
      1997 Final Research Report Summary
  • [Publications] Y.Yoshida et al.: "The gnd gene encoding a novel 6-phosphogluconate dehydrogenase and its adjacent region of Actinobacillus actinomycetemcomitans chromosomal DNA" Biochemical and Biophysical Research Communications. 230. 220-225 (1997)

    • Description
      「研究成果報告書概要(和文)」より
    • Related Report
      1997 Final Research Report Summary
  • [Publications] Y.Yoshida et al.: "Identification of a gene locus essential for serotype b-specific antigen synthesis in Actinobacillus actinomycetemcomitans" Infection and Immunity. 66. 107-114 (1998)

    • Description
      「研究成果報告書概要(和文)」より
    • Related Report
      1997 Final Research Report Summary
  • [Publications] 山下 喜久: "う蝕細菌の分子生物学「グルコシルトランスファーゼのう蝕誘発への寄与」" 岩波書店, 360 (1997)

    • Description
      「研究成果報告書概要(和文)」より
    • Related Report
      1997 Final Research Report Summary
  • [Publications] Sumi, Y., Yamashita, Y., Nakano, Y., and Koga, T.: "Semiquantitative analysis of periodontopathogens by gene amplification" Jpn.J.Med.Sci.Biol.48. 177-191 (1995)

    • Description
      「研究成果報告書概要(欧文)」より
    • Related Report
      1997 Final Research Report Summary
  • [Publications] Yamaguchi, N., Kawashaki, M., Yamashita, Y., Nakashima, K., and Koga, T.: "Role of capsular-like serotype-specific polysaccharide antigen in resistance to of Actinobacillus actinomycetemcomitans to phagocytosis by human polymorphonuclear leukocytes" Infect.Immun.63. 4589-4594 (1995)

    • Description
      「研究成果報告書概要(欧文)」より
    • Related Report
      1997 Final Research Report Summary
  • [Publications] Takamatsu-Matsushita, N., Yamaguchi, N., Kawasaki, M., Yamashita, Y., Takehara, T., and Koga, T.: "Immunogenicity of Actinobacillus actinomycetemcomitans serotype b-specific polysaccharide-protein conjugate" Oral Microbiol.Immunol.11. 220-225 (1996)

    • Description
      「研究成果報告書概要(欧文)」より
    • Related Report
      1997 Final Research Report Summary
  • [Publications] Nakano, Y., Yoshida.Y., Yamashita, Y., and Koga, T.: "New shuttle vectors for Actinobacillus actinomycetemcomitans and Escherichia coli" Gene. 169. 139-140 (1996)

    • Description
      「研究成果報告書概要(欧文)」より
    • Related Report
      1997 Final Research Report Summary
  • [Publications] Yamaguchi, N., Yamashita, Y., Ikeda.D., and Koga, T: "Actinobacillus actinomycetemcomitansserotype b-specific polysaccharide antigen stimulates production of chemotactic factors and inflammatory cytokines by human monocytes" Infect.Immun.64. 2563-2570 (1996)

    • Description
      「研究成果報告書概要(欧文)」より
    • Related Report
      1997 Final Research Report Summary
  • [Publications] Yoshida, Y., Nakano, Y., Yamashita, Y., and Koga, T.: "The gnd gene encoding a novel 6-phosphogluconate dehydrogenase and its adjacent region of Actinobacillus actinomycetemcomitans chromosomal DNA" Biochem.Biophys.Res Commun.230. 220-225 (1997)

    • Description
      「研究成果報告書概要(欧文)」より
    • Related Report
      1997 Final Research Report Summary
  • [Publications] Yoshida, Y., Nakano, Y., Yamashita, Y., and Koga, T: "Identification of a gene locus cssential for serotype b-specific antigen synthesis in Actinobacillus actinomycetemcomitans" Infect.Immun.66. 107-114 (1998)

    • Description
      「研究成果報告書概要(欧文)」より
    • Related Report
      1997 Final Research Report Summary
  • [Publications] Yasuo Yoshida et al.: "Identication of a genetic locus essential for serotype b-specific antigen synthesis in Actinobacillus actinomycetemcomitans" Infenction and Inmmunity. 66・1. 107-114 (1998)

    • Related Report
      1997 Annual Research Report
  • [Publications] Noboru Yamaguchi et al.: "Binding of the capusle-like serotype-specific polysaccharide antigen and the lipopolysaccharide from Actinobacilllus actinomycetemcomitans to human complement C36" Oral Microbiology and Immunology. (印刷中).

    • Related Report
      1997 Annual Research Report
  • [Publications] Nobumi Takamatsu-Matsushita: "Immunogenicity of Actiobacillus actinomycetemcomitans serotype b-specific polysaccharide-protein conjugate21GC01:Oral Microbiology and Immunology" 11. 220-225 (1996)

    • Related Report
      1996 Annual Research Report
  • [Publications] Yoshio Nakano et al.: "New shutlle vectors for Actinobacillus actinomycetemcomitans and Escherichia coli" Gene. 169. 139-140 (1996)

    • Related Report
      1996 Annual Research Report
  • [Publications] Noboru Yamaguchi et al.: "Actinobacillus actinomycetemcomitans serotype b-specific polysaccharide antigen stimulates production of chemotactic factors and inflammatory..." Infection and Immunity. 64・7. 2563-2570 (1997)

    • Related Report
      1996 Annual Research Report
  • [Publications] Yasuo Yoshida et al.: "The gnd gene encoding a novel 6-phosphogluconate dehydrogenase and its a digacent region of Actinobacillus actinomycetemcomitans chromosomal DNA" Biochemical and Biophysical Research Communication. 230. 220-225 (1997)

    • Related Report
      1996 Annual Research Report
  • [Publications] Yoshihisa SUMI: "Semiquantitative analysis of periodontopathogens by gene amplification" Japanese Journal of Medical Science & Biology. 48. 177-191 (1995)

    • Related Report
      1995 Annual Research Report
  • [Publications] Noboru YAMAGUCHI: "Role of the capsular polysaccharide-like serotype-specific antigen in resistance of Actinobacillus actinomycetemcomitans to phagocytosis by human pdymorphonuclear" Infection and Immunity. 63. 4589-4594 (1995)

    • Related Report
      1995 Annual Research Report

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Published: 1995-04-01   Modified: 2016-04-21  

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