title: Rapid real-time PCR for detection of Mycobacterium tuberculosis complex DNA in formalin-fixed paraffin embedded tissues: 16% of histological 'sarcoid' may contain such DNA.
abstract: OBJECTIVE
  To investigate the diagnostic accuracy of IS6110 real-time PCR for detection of Mycobacterium tuberculosis complex (MTBC) in DNA extracted from formalin-fixed paraffin embedded (FFPE) tissues using two different methods. In the absence of material submitted for tuberculosis (TB) culture, MTBC detection in FFPE tissue can be an important aid to diagnosis.
  
  
  METHODS
  We collected 144 FFPE tissue blocks (lung and lymph node) for IS6110 real-time PCR. Two DNA extraction methods (QIAamp FFPE tissue kit and NucliSENS easyMAG) were assessed within a general laboratory setting. PCR results were compared with histology and culture.
  
  
  RESULTS
  In the histological MTBC and culture MTBC (TB-positive) groups, 72.4% were IS6110-positive and 27.6% negative. IS6110-negative results were obtained from 98%, 61.5% and 84% of the histologically MTBC-negative (TB-negative) group, histologically TB/no culture group and sarcoidosis group, respectively. Review of 19 IS6110-positive patients in the latter three groups showed that 15 had clinical TB. Thirteen of 15 (86.7%) IS6110-positive patients in the histological TB/no culture group and 2 of 4 (50%) IS6110-positive patients in the sarcoidosis group were clinically diagnosed with TB which highlights the difficulty of a pathological diagnosis.
  
  
  CONCLUSIONS
  IS6110 real-time PCR using easyMAG extracted DNA is a moderately sensitive, specific and rapid method for MTBC detection in FFPE material, but must be interpreted in the overall clinical context. PCR results can be available in around 5 h from FFPE specimen receipt, with minimal hands-on time.
authors.affiliation: Regional Infectious Diseases Unit, Western General Hospital, Edinburgh, UK.
authors.forename: Güzin
authors.initials: G
authors.lastname: Surat
authors.affiliation: Department of Pathology, Royal Infirmary of Edinburgh, Edinburgh, UK.
authors.forename: William A
authors.initials: WA
authors.lastname: Wallace
authors.affiliation: Scottish Mycobacteria Reference Laboratory (SMRL), Royal Infirmary of Edinburgh, Edinburgh, UK.
authors.forename: Ian F
authors.initials: IF
authors.lastname: Laurenson
authors.affiliation: Scottish Mycobacteria Reference Laboratory (SMRL), Royal Infirmary of Edinburgh, Edinburgh, UK.
authors.forename: Amie-Louise
authors.initials: AL
authors.lastname: Seagar
mesh_terms: D004251:DNA Transposable Elements; D004269:DNA, Bacterial / Q000032:analysis*; D005557:Formaldehyde; D006801:Humans; D009169:Mycobacterium tuberculosis; D016612:Paraffin Embedding; D060888:Real-Time Polymerase Chain Reaction / Q000379:methods*; D012189:Retrospective Studies; D017565:Sarcoidosis, Pulmonary / Q000382:microbiology*; D016707:Tissue Fixation; D014376:Tuberculosis / Q000175:diagnosis*
chemical_list: D004251:DNA Transposable Elements; D004269:DNA, Bacterial; D005557:Formaldehyde
keywords: Histopathology; Mycobacteria; PCR