title: Microflora in teeth associated with apical periodontitis: a methodological observational study comparing two protocols and three microscopy techniques.
abstract: OBJECTIVE
  The aim of this study was to compare two protocols to examine bacterial colonization in teeth associated with chronic apical periodontitis with acute episodes (ap), using light microscopy (LM), transmission electron microscopy (TEM) and scanning electron microscopy (SEM).
  
  
  METHODS
  Nine root samples (seven teeth) were processed using either Eastman Dental Institute (EDI) (n = 4 teeth/4 roots) or Zurich (n = 3 teeth/5 roots) protocols. The roots were sectioned longitudinally; one root portion was viewed with SEM, descriptively dividing its length into apical, middle and coronal; semi-thin and ultra-thin transverse sections were viewed under LM and TEM from each third of the other root portion. Each root was therefore examined using all microscopy techniques. Observations of bacterial presence, description and distribution within the root canal lumen and root dentine were systematically recorded using pre-determined criteria.
  
  
  RESULTS
  The Zurich technique gave a more predictable division of the root, but the surface was slightly smeared and demineralization was incomplete. The Eastman Dental Institute (EDI) approach appeared to provide better ultrastructural detail. Bacteria were detected in eight of the nine roots. Bacterial biofilms were commonly seen adhering to the root canal surface, containing various cellular morphotypes: rods, cocci, filaments and spirochaetes. Bacteria were more evident apically than coronally, associated with the canal wall but were more commonly evident coronally than apically within the dentinal tubules. Polymorphs (PMNs) were found in all the root thirds, especially apically, often numerous and walling off the bacterial biofilm from the remaining canal lumen.
  
  
  CONCLUSIONS
  Both protocols had merits and de-merits. The combination of microscopy techniques offered complementary views of intra-radicular bacterial colonization. The perception of confinement of the host/microbial interface at the apical foramen is not entirely correct; PMNs may be found even in the coronal third of root canals containing necrotic pulp tissue.
authors: Unit of Endodontology, Division of Restorative Dental Science, UCL Eastman Dental Institute, University College London, London WC1X 8LD, UK.
authors: N
authors: N
authors: Richardson
authors: N J
authors: NJ
authors: Mordan
authors: J A P
authors: JA
authors: Figueiredo
authors: Y-L
authors: YL
authors: Ng
authors: K
authors: K
authors: Gulabivala
mesh_terms: D001419:Bacteria / Q000145:classification / Q000648:ultrastructure*; D018441:Biofilms; D003648:Decalcification Technique; D003786:Dental Pulp Cavity / Q000382:microbiology* / Q000648:ultrastructure; D003790:Dental Pulp Necrosis / Q000382:microbiology / Q000473:pathology; D003804:Dentin / Q000382:microbiology / Q000648:ultrastructure; D004912:Erythrocytes / Q000382:microbiology / Q000648:ultrastructure; D006652:Histological Techniques; D054884:Host-Pathogen Interactions; D006801:Humans; D008853:Microscopy; D008855:Microscopy, Electron, Scanning; D046529:Microscopy, Electron, Transmission; D009504:Neutrophils / Q000382:microbiology / Q000648:ultrastructure; D010485:Periapical Periodontitis / Q000382:microbiology* / Q000473:pathology; D016610:Tissue Embedding; D019227:Tooth Apex / Q000382:microbiology / Q000648:ultrastructure