Primary Multidrug Resistant Tuberculosis and Utility of Line Probe Assay for Its Detection in Smear-Positive Sputum Samples in a Tertiary Care Hospital in South India

Abstract

In a high tuberculosis burdened country like India, rapid, cost-effective, and reliable diagnostic tools for tuberculosis are an urgent need of the hour to prevent inappropriate treatment strategies and further spread of resistance. This study aimed to estimate the proportion of new smear-positive tuberculosis cases with primary resistance to rifampicin and/or isoniazid as well as identify the common mutations associated with it. Sputum of 200 newly diagnosed smear-positive cases of 1+ score and above was directly subjected to Line Probe Assay using the GenoType MTBDRplus assay kit. All samples were inoculated onto solid media and 61 samples were inoculated in automated liquid culture also. The Line Probe Assay gave hundred percent interpretable results with 2.5% of the study population showing resistant pattern. Only 1% of the cases were primary multidrug resistant tuberculosis and 1.5% showed isoniazid monoresistance. S531L and C15T were the most common genetic mutations seen for rifampicin and isoniazid resistance, respectively. 40% had absent rpoB wild type 8 band indicating probable silent mutation after clinical correlation. The average turnaround time for Line Probe Assay was far less (3.8 days) as compared to solid and liquid cultures (35.6 days and 13.5 days, resp.).

Cost minimization analysis of line probe assay for detection of multidrug-resistant tuberculosis in Arkhangelsk region of Russian Federation

“…Clinical outcomes of LPA implementation in Russia proved its high effectiveness [15]. These findings were line with other recent studies that Line Probe Assay is a rapid tool for screening TB and DST [7, 8, 10, 15, 34–41]. The economic evaluation of LPA proved that it was a cost minimizing strategy for both Healthcare system (with cost minimization observed in costs of hospital days, laboratory costs, medications) and for patients.…”

“…First-line LPAs were first recommended by WHO in 2008; the second-line LPA was first recommended in May 2016 [1]. However, some research still show that the LPA cannot completely replace phenotypic culture methods [34]. High costs of diagnostic equipment, its maintenance and consumables need extra investments and make it extremely important to assess cost minimization of innovative methods for TB diagnostics based on full societal costs (including overall health service-related and patient’s cost).…”

Molecular Detection of Rifampicin and Isoniazid Resistant Mycobacterium Tuberculosis and Their Associated Mutation Pattern From Smear Positive Sputum Samples From a Tertiary Care Centre of West U.P.

“…This mixture is used within 24 hours of preparation. [5] DNA extraction-500μl of the decontaminated specimen is taken in a microcentrifuge tube and centrifuged at 13000rpm for 15 minutes then supernatant was discarded and pellet is re-suspended in 100μl of molecular grade water and incubated in a heating bath at 95°C for 20 minutes. After this, it was kept in sonicator for 15 minutes.…”

“…After this, it was kept in sonicator for 15 minutes. [5] Mastermix Preparation-Master mix was prepared by combining the 5μl extracted DNA with the specific primers (AM-A and AM-B) as per instruction in kit manual and then this mixture was kept in thermal cycler for amplification. Amplification-Amplification was done in Veriti® 96 well thermocycler by Thermofisher™.…”

“…Hybridization steps were performed using reagents provided with the kit as per manufacturer's instruction. [5]…”