Methods to Improve Confidence in the Accuracy of Molecular Testing for Multidrug-Resistant Tuberculosis

Abstract Background Diagnosis of tuberculosis (TB) and multidrug-resistant tuberculosis (MDR-TB) is increasingly performed using molecular tools that detect Mycobacterium tuberculosis DNA. To ensure accurate and reliable results from the molecular tests, appropriate quality assessment is required. This involves implementing reference measurement procedures (RMPs) to characterize material standards that are representative of the clinical specimen. These material standards should address drug resistance and mixtures of drug-resistant and -susceptible bacteria. However, currently these RMPs and materials standards do not exist, which can hamper the accuracy and precision of routine clinical testing. To address this, we applied digital PCR (dPCR) as a RMP to MDR-TB material standards. Methods Four standards were prepared and characterized using dPCR to quantify drug-resistant and -susceptible genotypes. We investigated the performance of existing molecular tests via an interlaboratory study including 9 laboratories from Africa and Europe, assessing 3 methods for MDR-TB detection and 2 methods for TB-only detection. Results All tests correctly identified M. tuberculosis, and 2 out of 3 tests identified the associated drug resistance (one test failed to identify drug resistance in one of the materials). Generally, discrepancies occurred with the more challenging samples bearing lower concentrations and mixed genotypes. Conclusions The approaches used in this study will enhance the quality assessment of MDR-TB and can be applied to afford test manufacturers and clinical laboratories more accurate results to guide test development, selection, and regulation. Such an approach can improve confidence in MDR-TB testing, enabling physicians to guide treatment, potentially leading to better patient outcomes.