Detection of Drug Susceptibility and Drug Interactions in <em>Mycobacterium tuberculosis</em> using Resazurin Microtiter and Checkerboard Assay

The emergence of multidrug-resistant tuberculosis (MDR-TB) necessitates rapid and accurate drug susceptibility testing (DST) methods to guide effective treatment. This study introduces a methodology combining the resazurin microtiter assay (REMA) and checkerboard assay to determine minimum inhibitory concentration (MIC) and evaluate drug-drug interactions of anti-tuberculosis drugs against Mycobacterium tuberculosis. The REMA, adapted to a 96-well format, leverages the reduction of resazurin dye by metabolically active M. tuberculosis as a visual indicator of drug susceptibility. Varying concentrations of anti-TB drugs are tested against M. tuberculosis isolates, and color changes are observed to determine the MIC. Subsequently, a checkerboard assay is employed to assess potential synergistic, additive, or antagonistic effects between drug combinations. This simple and inexpensive method yields results within seven days, offering a significant advantage over traditional DST methods. This method provides valuable insights into the DST of M. tuberculosis isolates and facilitates the identification of promising drug combinations for improved treatment outcomes against MDR-TB.