Assessment of intraradicular bacterial composition by terminal restriction fragment length polymorphism analysis

Oral Microbiol Immunol. 2009 Oct;24(5):369-76. doi: 10.1111/j.1399-302X.2009.00525.x.

Abstract

Background: The aim of the study was to assess the bacterial community structures associated with endodontic infections using terminal restriction fragment length polymorphism (T-RFLP), and to investigate the correlation of whole community profiles with the manifestation of particular clinical features.

Methods: Intraradicular samples were collected from 34 subjects and classified into three study groups based on the observed clinical symptoms: acute (n = 16), sub-acute (n = 8), and asymptomatic (n = 10). Genomic DNA was extracted from each sample, submitted to polymerase chain reaction using a fluorescently labeled 16S ribosomal DNA forward primer, and digested with two tetrameric endonucleases (HhaI and MspI). The terminal restriction fragments (T-RFs) were subsequently discriminated in an automated DNA sequencer, and the results were filtered using a statistics-based criterion.

Results: Totals of 138 (HhaI) and 145 (MspI) unique T-RFs were detected (means 13.1 and 11.9) and there was high inter-subject variability in the bacterial assemblages. Odds-ratio analysis unveiled the existence of higher order groups of positively associated T-RFs, restating the concept that intricate ecological relationships may take place in the root canal space. A significantly greater T-RF prevalence was detected in acute cases, suggesting a straight correlation between species richness and spontaneous pain.

Conclusion: Overall, no T-RFLP profile representing a specific bacterial consortium could be associated with the manifestation of symptoms of endodontic origin.

Publication types

  • Comparative Study
  • Research Support, Non-U.S. Gov't

MeSH terms

  • Actinomyces / classification
  • Adolescent
  • Adult
  • Bacteria / classification*
  • Bacteria / genetics
  • Bacteroides / classification
  • Campylobacter sputorum / classification
  • Capnocytophaga / classification
  • DNA, Bacterial / genetics
  • Dental Pulp Cavity / microbiology*
  • Dental Pulp Necrosis / microbiology*
  • Deoxyribonuclease HpaII
  • Deoxyribonucleases, Type II Site-Specific
  • Eubacterium / classification
  • Female
  • Flavobacterium / classification
  • Fusobacterium nucleatum / classification
  • Humans
  • Lactobacillus / classification
  • Male
  • Middle Aged
  • Peptostreptococcus / classification
  • Periapical Diseases / microbiology
  • Polymorphism, Restriction Fragment Length / genetics*
  • Prevotella / classification
  • Selenomonas / classification
  • Sequence Analysis, DNA
  • Veillonella / classification
  • Young Adult

Substances

  • DNA, Bacterial
  • Deoxyribonuclease HpaII
  • Deoxyribonucleases, Type II Site-Specific
  • GCGC-specific type II deoxyribonucleases