Evaluation of Rpoβ Gene and Its Various Mutants in Multidrug-Resistant Tuberculosis Cases by GeneXpert Method

Abstract

Introduction: India is one of the countries in the world which contribute to the global burden of multidrugresistant tuberculosis (MDR-TB). GeneXpert is a method recommended by the World Health Organization (WHO) that uses five overlapping probes (Probe A to E) to detect mutation in the beta subunit of ribonucleic acid (RNA) polymerase gene (Rpoβ) responsible for rifampicin resistance in Mycobacterium tuberculosis (MTB).Method: All the pulmonary and extrapulmonary samples received in tuberculosis (TB) laboratory for testing of MTB from January 2018 to December 2020 were analyzed for bacillary load and rifampicin resistance was identified by analyzing the number of missing probes.Results: During the study period, a total of 10,021 samples were tested for MDR-TB. Out of those samples, 2674 samples were positive for MTB of which 2321 were pulmonary samples and 353 were extra-pulmonary samples. Rifampicin resistance was detected in 385 pulmonary samples and 63 extrapulmonary samples. These samples were further differentiated according to the bacillary load. The highest number of mutations was observed in Probe E followed by Probe B, Probe A, and Probe D, and the lowest was observed in Probe C. Also, mutations were associated when all probes were present or a few combinations of probes were missing.Conclusion: GeneXpert assay is a rapid molecular method that detected MTB and rifampicin resistance with a two-hour turnaround. It uses molecular beacons to detect mutation in the Rpoβ gene. This study can be useful in analyzing the prevalence and epidemiology of MTB in a particular demographic area and also the addition of a few more probes can enhance the identification of the mutation in other codons and therefore better therapeutic interventions can be developed accordingly.

Evaluating the Sensitivity of Different Molecular Techniques for Detecting Mycobacterium tuberculosis Complex in Patients with Pulmonary Infection

“…In comparison, GeneXpert MTB/RIF and phenotypic DST tests confirmed only four samples to be drug-resistant MTBC isolates. A hypothesis supported by several studies regarding the inability of GeneXpert MTB/RIF to detect RIF resistance-conferring mutations in sputum samples with heterologous DNA flora was due to the small number of mutant rpo B gene alleles, which should be present in at least 65% of the sample DNA to be detected by the GeneXpert assay ( Miotto et al 2018 ; S J, Kar et al 2022 ).…”