Journal of Microbiology

The Microbiological Society of Korea (한국미생물학회)
권호 표지

Development of Strain-specific PCR Primers Based on a DNA Probe Fu12 for the Identification of Fusobacterium nucleatum subsp. nucleatum ATCC $25586^T$

  • Kim Hwa-Sook (Department of Oral Biochemistry, Seoul National University) ;
  • Song Soo Keun (Department of Oral Biochemistry, Seoul National University) ;
  • Yoo So Young (Department of Oral Biochemistry, Seoul National University) ;
  • Jin Dong Chun (Department of Dental Pharmacology, Seoul National University) ;
  • Shin Hwan Seon (Department of Oral Biochemistry, Seoul National University) ;
  • Lim Chae Kwang (Department of Oral Biochemistry, Seoul National University) ;
  • Kim Myong Soo (Department of Dental Pharmacology, Seoul National University) ;
  • Kim Jin-Soo (Department of Oral and Maxillofacial Radiology, Seoul National University) ;
  • Choe Son-Jin (Department of Oral Microbiology and Immunology, College of Dentistry, Seoul National University) ;
  • Kook Joong-Ki (Department of Oral Biochemistry, Seoul National University)
  • Published : 2005.08.01
Download PDF
⟨ Previous Next ⟩

Abstract

The objective of this study was to assess the strain-specificity of a DNA probe, Fu12, for Fusobacterium nucleatum subsp. nucleatum ATCC $25586^T$ (F. nucleatum ATCC $25586^T$), and to develop sets of strain-specific polymerase chain reaction (PCR) primers. Strain-specificity was tested against 16 strains of F. nucleatum and 3 strains of distinct Fusobacterium species. Southern blot hybridization revealed that the Fu12 reacted exclusively with the HindIII-digested genomic DNA of F. nucleatum ATCC $25586^T$. The results of PCR revealed that three pairs of PCR primers, based on the nucleotide sequence of Fu12, generated the strain-specific amplicons from F. nucleatum ATCC $25586^T$. These results suggest that the DNA probe Fu12 and the three pairs of PCR primers could be useful in the identification of F. nucleatum ATCC $25586^T$, especially with regard to the determination of the authenticity of the strain.

Keywords

References

  1. Albandar, J.M., L.J. Brown, and H. Loe. 1997. Putative periodontal pathogens in subgingival plaque of young adults with and without early-onset periodontitis. J. Periodontol. 68, 973-981 https://doi.org/10.1902/jop.1997.68.10.973
  2. Baek, D.-H., M.-S. Kim, D.-K Kim, and J.-K. Kook. 2003. Identification of Fusobacterium nucleatum ATCC 25586 using potentially strain-specific DNA probe Fu12. Int. J. Oral Biol. 28, 1-4
  3. Diaz, P.I., P.S. Zilm, and A.H. Rogers. 2002. Fusobacterium nucleatum supports the growth of Porphyromonas gingivalis in oxygenated and carbon-dioxide-depleted environments. Microbiol. 148, 467-472 https://doi.org/10.1099/00221287-148-2-467
  4. DiRienzo, J.M., S. Cornell, and H. Boehringer. 1991. Use of randomly cloned DNA fragments for the identification of oral spirochetes. Oral Microbiol. Immunol. 6, 88-96 https://doi.org/10.1111/j.1399-302X.1991.tb00457.x
  5. Dzink, J.L., M.T. Sheenan, and S.S. Socransky. 1990. Proposal of three subspecies of Fusobacterium nucleatum Knorr 1922: Fusobacterium nucleatum subsp. nucleatum subsp. nov., comb. nov.; Fusobacterium nucleatum subsp. polymorphum subsp. nov., nom. rev., comb. nov.; and Fusobacterium nucleatum subsp. vincentii subsp. nov., nom. rev., comb. nov. Int. J. Syst. Bacteriol. 40, 74-78 https://doi.org/10.1099/00207713-40-1-74
  6. Gharbia, S.E. and H.N. Shah. 1992. Fusobacterium nucleatum subsp. fusiforme subsp. nov. and Fusobacterium nucleatum subsp. animalis subsp. nov. as additional subspecies within Fusobacterium nucleatum. Int. J. Syst. Bacteriol. 42, 296-298 https://doi.org/10.1099/00207713-42-2-296
  7. Kim, M.-K., H.-S. Kim, B.-O. Kim, S.Y. Yoo, J.-H. Seong, D.-K. Kim, S.E. Lee, S.-J. Choe, J.-C. Park, B.-M. Min, M.-J. Jeong, D.K. Kim, Y.K. Shin, and J.-K. Kook. 2004. Multiplex PCR Using Conserved and Species-Specific 16S rDNA Primers for Simultaneous Detection of Fusobacterium nucleatum and Actinobacillus actinomycetemcomitans. J. Microbiol. Biotechnol. 14, 110-115
  8. Komiya, A., T. Kato, T. Nakagawa, A. Saito, J. Takahashi, S. Yamada, and K. Okuda. 2000. A rapid DNA probe method for detection of Porphyromonas gingivalis and Actinobacillus actinomycetemcomitans. J. Periodontol. 71, 760-767 https://doi.org/10.1902/jop.2000.71.5.760
  9. Kook, J.-K., J.J. Han, H.-S. Kim, J.-H. Seong, D.-K. Kim, D.-H. Baek, and S.-J. Choe. 2003a. Cloning of a potentially strainspecific DNA probe of Prevotella intermedia ATCC 25611 by inverted dot blot hybridization screening method. J. Microbiol. Biotechnol. 13, 282-286
  10. Kook, J.-K., M.-K. Kim, J.-H. Seong, D.-K. Kim, B.-O. Kim, J.-C. Park, K.-K. Kim, S.-J. Choe, and B.-M. Min. 2003b. A new method for rapid screening of bacterial species- or subspeciesspecific DNA probes. FEMS Microbiol. Lett. 219, 121-127 https://doi.org/10.1016/S0378-1097(03)00021-1
  11. Krieg, N.R. 2001. Identification of Procaryotes, p.33-38. In G. Garrity (ed.), Bergey's Manual of Systematic Bacteriology, Vol. 1, 2nd ed. Springer Verlag, New York
  12. Lee, S.-A., S.Y. Yoo, K.-S. Kay, and J.-K. Kook. 2004. Detection of Hepatitis B virus and Mycobacterium tuberculosis in Korean dental patients. J. Microbiol. 42, 239-242
  13. Lippke, J.A., M.N. Strzempko, F.F. Raia, S.L. Simon, and C.K. French. 1987. Isolation of intact high-molecular-weight DNA by using guanidine isothiocyanate. Appl. Environ. Microbiol. 53, 2588-2589
  14. Matheson, V.G., J. Munakata-Marr, G.D. Hopkins, P.L. McCarty, J.M. Tiedje, and L.J. Forney. 1997. A novel means to develop strain-specific DNA probes for detecting bacteria in the envi ronment. Appl. Environ. Microbiol. 63, 2863-2869
  15. Roberts, G.L. 2000. Fusobacterial infections: an underestimated threat. Br. J. Biomed. Sci. 57, 156-162
  16. Slots, J., A. Ashimoto, M.J. Flynn, G. Li, and C. Chen. 1995. Detection of putative periodontal pathogens in subgingival by 16S ribosomal DNA amplification with the polymerase chain reaction. Clin. Infect. Dis. 20(suppl. 2), 304-307 https://doi.org/10.1093/clinids/20.Supplement_2.S304
  17. Tew, J.G., S.S. Thomas, and R.R. Ranney. 1987. Fusobacterium nucleatum-mediated immunomodulation of the in vitro secondary antibody response to tetanus toxoid and Actinobacillus actinomycetemcomitans. J. Periodontal. Res. 22, 506-512 https://doi.org/10.1111/j.1600-0765.1987.tb02062.x
  18. Tran, S.D. and J.D. Rudney. 1999. Improved multiplex PCR using conserved and species-specific 16S rRNA gene primers for simultaneous detection of Actinobacillus actinomycetemcomitans, Bacteroides forsythus, and Porphyromonas gingivalis. J. Clin. Microbiol. 37, 3504-3508
  19. Wang, H.-Y. and G.-H. Lee. 2003. Rapid Identification of Vibrio vulnificus in Seawater by Real-Time Quantitative TaqMan PCR. J. Microbiol. 41, 320-326