Band Pattern Analysis of the rpoB, inhA, and katG Gene in Mycobacterium tuberculosis through Line Probe Assay in Patients from Western Rajasthan, India

Abstract

Tuberculosis is globally a leading cause of death with increasing rates of drug resistance which has become a serious topic of concern. Early diagnosis and properly administered treatment regimen are the key elements to combat Tuberculosis and its transmission. Rapid molecular methods have been recommended for early detection of drug resistant Tuberculosis. This study aimed to analyze the mutation band pattern in rpoB, inhA, katG gene of drug resistant pulmonary tuberculosis through Hybridization technique using Genotype MTBDR plus V.2 assay in Western Rajasthan. Positive sputum samples were collected from MDR/ Mono DR Patients. Sputum decontamination was performed after the quality assurance then DNA extraction, amplification, hybridization and band pattern analysis of line probe assay strips was performed as per manufacturer’s instructions by GenoType MTBDRplus kit from Hain Lifescience. Out of 215 study participants, 174 (80.9%) were male and 41 (19%) were female, with a mean age group of 41 years, where (146/215; 67.9%) were rural dwellers. 56.2% of the population was sensitive to both the drug (Rifampicin and Isoniazid). Of the 215 isolates, 12 (5.58%) were hetero-resistant to both RIF and INH i.e. the overall proportion of MDR-TB was 12 (5.58%) There were 6 isolates (2.79%) mono-resistant to RIF and 16 isolates (7.44%) mono-resistant to INH. In rpoB gene, WT 7 (2%) and WT 8 (7.2%) band were the most common band to be absent. In katG gene, MUT 1 band showed maximum mutation (14.9%). In inhA gene, WT 1 band was most common band to be absent (4.6%) and MUT 1 band showed 2.5% of mutation. In the present study analysis of MUT 2 band of rpoB, katG and inhA genes did not show any presence. High frequencies of mutations in rpoB, inhA, and katG gene by LPA were observed. The most common resistance-conferring mutations to RIF occurred at rpoB gene with absence of band WT8 and presence of MUT3 band. In katG gene absence of WT1 and presence of MUT1 band was most common pattern of mutation and in inhA gene, WT1 band was prominently absent.