Luciferase reporter phage phAE85 for rapid detection of rifampicin resistance in clinical isolates of Mycobacterium tuberculosis

Abstract

AbstractObjectiveTo evaluate luciferase reporter phage (LRP) phAE85 in rapid detection of rifampicin resistance in a region where TB is endemic.MethodsOne 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.ResultsThe 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.ConclusionsLRP 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