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

Gomathi, S.; Balaji, S.; Ponnuraja, C.; Kumar, V.

Asian Pacific Journal of Tropical Medicine; 2013; 6; 728-731.

Abstract: Objective: To evaluate luciferase reporter phage (LRP) phAE85 in rapid detection of rifampicin 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 m g/ml was inoculated with standard inoculum from suspensions of the clinical isolate. After incubation for 72 h, LRP was added. Folowiong 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 concentraton method (MIC) of drug susceptibility testing.

Results : The two methods showed high level agreement of 97% ( CI 0.94, 0.99) and P value was 0.0001. The sensitivity and specificity of LRP assay for detection of rifampicin 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 cmparison 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.

Keywords: Mycobacterium tuberculosis ; Rifampicin resistance; Rapid detection

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