Efficacy of TrueNat cartridge extracted DNA for detecting drug resistant tuberculosis by line probe assay

Abstract

BackgroundCurrently for diagnosing Mycobacterium tuberculosis and its drug resistance, two sputum samples are required. One of them is subjected to TrueNat™ and if positive the other sample is subjected to line probe assay (LPA). This study was done to evaluate whether TrueNat extracted DNA can be directly used for performing LPA in a diagnostic laboratory setting to decrease patient turn-around time. MethodsTotal 45 smear positive sputum samples were subjected to TrueNat™ MTB detection and first and second line (FL and SL) LPA testing in parallel. DNA extracted by Trueprep® Cartridge was also tested by LPA and results were compared. Further, TrueNat extracted DNA from 20 samples was divided into 6 aliquots each, two of which were stored at 4 °C, 37 °C and 55 °C (under humidification) each. One aliquot from stored DNA at each temperature was used for FL & SL LPA on day three and the other on day eight. The blots thus obtained were compared with those of conventional LPA at day 1. ResultsFor FL-LPA, TrueNat extracted DNA gave valid results for all 45 (100%) samples but conventionally extracted DNA could give results for 44 (97.8%) samples. Likewise, for SL-LPA, valid results were obtained for 40 (88.9%) and 35 (77.8%) samples respectively using TrueNat extracted DNA and conventionally extracted DNA respectively. All samples with invalid LPA results had Ct values ≥ 28 by TrueNat PCR. LPA results were obtained for all the 20 samples using stored DNA at all temperatures and duration. ConclusionsTrueNat extracted DNA can be used for performing LPA under field conditions for selected samples.