Recent studies in addition to studies based on whole genome sequencing (WGS) of clinical isolates of Mycobacterium tuberculosis (MTB) from diverse geographical regions have provided useful insights into the mechanisms of drug resistance. Of importance, are some of the findings pertaining to mechanisms of resistance to two of the first-line anti-tuberculosis (TB) drugs, namely, rifampicin (RIF) and pyrazinamide (PZA). For example, the implication of mutations in rpoA and rpoC genes that act as compensatory mutations for those in the rpoB gene with respect to RIF resistance is noteworthy. Similarly, in the case of PZA resistance, the role of rpsA and panD genes has recently been noted. This highlights the evolving knowledge of resistance to these drugs. The present article provides a detailed account of the molecular mechanisms of resistance against two sterilizing first line anti-TB drugs (RIF and PZA) as well as an overview of sequence-based methods for detection of resistance to the drugs.