title: Clinical utility of RD1, RD9 and hsp65 based PCR assay for the identification of BCG in vaccinated children.
abstract: BACKGROUND
  Mycobacterium bovis Bacille Calmette-Guérin (BCG) vaccine is widely administered to prevent tuberculosis. Vaccine complications are rare. However, when BCG-related adverse reactions arise there is a need to rapidly and reliably identify BCG from other members of the Mycobacterium tuberculosis complex (TBC). PCR assays based on the detection of the regions of difference (RD), in particular RD1 and RD9, have been invaluable in the identification of BCG. Prior to this study, specimens were identified through HPLC analysis at a local reference laboratory taking up to 2 weeks for a result. We sought to expedite the identification process by validating a RD1, RD9 and hsp65 PCR assay for the identification and differentiation of BCG from TBC.
  
  
  RESULTS
  In last past 3 years, we validated the RD1, RD9 and hsp65 PCR assay for 16 mycobacterial isolates obtained from children who had experienced adverse reactions to BCG vaccination. In these cases, the clinician required a definitive identification of the isolate. The RD1 and RD9 PCR profiles indicated that all 16 isolates were BCG whilst amplification of the hsp65 target functioned as a PCR positive control. When tested against clinical M. tuberculosis (MTB), reference and non-tuberculous mycobacteria the PCR assay demonstrated 100% sensitivity and specificity.
  
  
  CONCLUSIONS
  The RD1, RD9 and hsp65 PCR assay is a useful tool for the rapid and reliable identification of BCG. Its ease of use has allowed it to be implemented in our clinical microbiology laboratory.
authors.affiliation: Department of Laboratory Medicine, Microbiology Unit, National University Hospital, Singapore 119074, Republic of Singapore. Jeanette_Teo@nuhs.edu.sg.
authors.forename: Jeanette W P
authors.initials: JW
authors.lastname: Teo
authors.forename: Janet W S
authors.initials: JW
authors.lastname: Cheng
authors.forename: Roland
authors.initials: R
authors.lastname: Jureen
authors.forename: Raymond T P
authors.initials: RT
authors.lastname: Lin
mesh_terms: D001500:BCG Vaccine / Q000008:administration & dosage / Q000009:adverse effects; D001426:Bacterial Proteins / Q000235:genetics* / Q000302:isolation & purification; D018834:Chaperonin 60 / Q000235:genetics* / Q000302:isolation & purification; D017931:DNA Primers / Q000737:chemistry; D004269:DNA, Bacterial / Q000235:genetics* / Q000302:isolation & purification; D005260:Female; D006801:Humans; D007223:Infant; D008297:Male; D009163:Mycobacterium bovis / Q000235:genetics / Q000302:isolation & purification; D009169:Mycobacterium tuberculosis / Q000235:genetics* / Q000302:isolation & purification; D016133:Polymerase Chain Reaction / Q000379:methods*; D012680:Sensitivity and Specificity; D014397:Tuberculosis, Pulmonary / Q000517:prevention & control
chemical_list: D001500:BCG Vaccine; D001426:Bacterial Proteins; D018834:Chaperonin 60; D017931:DNA Primers; D004269:DNA, Bacterial; C074119:heat-shock protein 65, Mycobacterium