title: Novel rapid PCR for the detection of Ile491Phe rpoB mutation of Mycobacterium tuberculosis, a rifampicin-resistance-conferring mutation undetected by commercial assays.
abstract: OBJECTIVE
  Neither the liquid medium-based Bactec MGIT, nor commercial molecular assays such as the Xpert MTB/RIF and the MTBDRplus V2.0 assays are capable of detecting up to 30% of rifampicin-resistant Mycobacterium tuberculosis strains in Swaziland because of the large proportion of the rpoB Ile491Phe mutations. In other countries, the frequency of this mutation is thought to be low.
  
  
  METHODS
  We designed a real-time multiplex allele-specific PCR assay to identify the rpoB Ile491Phe mutation responsible for these undetected resistant M. tuberculosis strains.
  
  
  RESULTS
  The technique showed 100% similarity with rpoB sequencing on a panel of 78 strains from Swaziland.
  
  
  CONCLUSIONS
  We propose that the detection of the rpoB Ile491Phe rpoB mutation should complement commercial assays for the diagnosis of rifampicin-resistant M. tuberculosis in routine conditions, particularly in countries where this specific mutation is frequent. The technique proposed in this paper is adapted for most reference laboratories.
authors.affiliation: Pôle de Microbiologie Médicale, Institut de Recherche Expérimentale et Clinique, Université Catholique de Louvain, Brussels, Belgium; Service de Microbiologie, Cliniques Universitaires Saint-Luc, Brussels, Belgium. Electronic address: emmanuel.andre@uclouvain.be.
authors.forename: E
authors.initials: E
authors.lastname: André
authors.affiliation: Pôle de Microbiologie Médicale, Institut de Recherche Expérimentale et Clinique, Université Catholique de Louvain, Brussels, Belgium.
authors.forename: L
authors.initials: L
authors.lastname: Goeminne
authors.affiliation: Service de Microbiologie, Cliniques Universitaires Saint-Luc, Brussels, Belgium.
authors.forename: A
authors.initials: A
authors.lastname: Colmant
authors.affiliation: Molecular and Experimental Mycobacteriology Group, Research Centre Borstel, Borstel, Germany; German Centre for Infection Research, Borstel Site, Borstel, Germany.
authors.forename: P
authors.initials: P
authors.lastname: Beckert
authors.affiliation: Molecular and Experimental Mycobacteriology Group, Research Centre Borstel, Borstel, Germany; German Centre for Infection Research, Borstel Site, Borstel, Germany.
authors.forename: S
authors.initials: S
authors.lastname: Niemann
authors.affiliation: Pôle de Microbiologie Médicale, Institut de Recherche Expérimentale et Clinique, Université Catholique de Louvain, Brussels, Belgium; Service de Microbiologie, Cliniques Universitaires Saint-Luc, Brussels, Belgium.
authors.forename: M
authors.initials: M
authors.lastname: Delmee
mesh_terms: D019943:Amino Acid Substitution; D000904:Antibiotics, Antitubercular / Q000494:pharmacology*; D001426:Bacterial Proteins / Q000235:genetics*; D003062:Codon; D012321:DNA-Directed RNA Polymerases / Q000235:genetics*; D024881:Drug Resistance, Bacterial; D006801:Humans; D060885:Multiplex Polymerase Chain Reaction; D009154:Mutation; D009169:Mycobacterium tuberculosis / Q000187:drug effects* / Q000235:genetics*; D060888:Real-Time Polymerase Chain Reaction; D012293:Rifampin / Q000494:pharmacology*; D014376:Tuberculosis / Q000175:diagnosis* / Q000382:microbiology*
chemical_list: D000904:Antibiotics, Antitubercular; D001426:Bacterial Proteins; D003062:Codon; C505805:rpoB protein, Mycobacterium tuberculosis; D012321:DNA-Directed RNA Polymerases; D012293:Rifampin
keywords: I491F; I572F; Ile491Phe; Ile572Phe; Multidrug-resistant tuberculosis; Multiplex allele-specific-PCR; Tuberculosis; Xpert; rpoB