title: Luciferase reporter phage phAE85 for rapid detection of rifampicin resistance in clinical isolates of Mycobacterium tuberculosis.
abstract: OBJECTIVE
  To evaluate luciferase reporter phage (LRP) phAE85 in rapid detection of rifampicin resistance in a region where TB is endemic.
  
  
  METHODS
  One hundred and ninety primary isolates on Lowenstein-Jensen medium were tested. Middlebrook 7H9 complete medium with and without rifampicin at 2 μg/mL was inoculated with standard inoculum from suspensions of the clinical isolate. After incubation for 72 h, LRP was added. Following 4 h of further incubation, light output from both control and test was measured as relative light units. Strains exhibiting a reduction of less than 50% relative light units in the drug containing vial compared to control were classified as resistant. Results were compared with the conventional minimum inhibitory concentration method (MIC) of drug susceptibility testing.
  
  
  RESULTS
  The two methods showed high level of agreement of 97% (CI 0.94, 0.99) and P value was 0.000 1. The sensitivity and specificity of LRP assay for detection of rifampicin resistance were 91% (CI 0.75, 0.98) and 99% (CI 0.95, 1.00) respectively. Time to detection of resistance by LRP assay was 3 d in comparison with 28 d by the minimum inhibitory concentration method.
  
  
  CONCLUSIONS
  LRP assay with phAE85 is 99% specific, 91% sensitive and is highly reproducible. Thus the assay offers a simple procedure for drug sensitivity testing, within the scope of semi-automation.
authors.affiliation: Department of Bacteriology, National Institute for Research in Tuberculosis (Formerly Tuberculosis Research Centre), Chennai, 600031, India.
authors.forename: Gomathi
authors.initials: G
authors.lastname: Sivaramakrishnan
authors.forename: Balaji
authors.initials: B
authors.lastname: Subramanyam
authors.forename: Ponnuraja
authors.initials: P
authors.lastname: C
authors.forename: Vanaja
authors.initials: V
authors.lastname: Kumar
mesh_terms: D000904:Antibiotics, Antitubercular / Q000494:pharmacology*; D024881:Drug Resistance, Bacterial; D017930:Genes, Reporter; D006801:Humans; D008156:Luciferases / Q000235:genetics / Q000378:metabolism; D008826:Microbial Sensitivity Tests; D009160:Mycobacteriophages / Q000235:genetics / Q000502:physiology*; D009169:Mycobacterium tuberculosis / Q000187:drug effects* / Q000302:isolation & purification / Q000821:virology*; D012293:Rifampin / Q000494:pharmacology*; D012680:Sensitivity and Specificity; D018088:Tuberculosis, Multidrug-Resistant / Q000382:microbiology*
chemical_list: D000904:Antibiotics, Antitubercular; D008156:Luciferases; D012293:Rifampin
keywords: Mycobacterium tuberculosis; Rapid detection; Rifampicin resistance