Background Assays enabling efficient high throughput drug screening are necessary for the discovery of new anti-mycobacterial drugs. The purpose of our work was to develop and validate an assay based on live-cell imaging which can monitor growth of two distinct phenotypes of Mycobacterium tuberculosis and to test their susceptibility to commonly used TB drugs. Results Both planktonic and cording phenotypes were successfully monitored as fluorescent objects using the live-cell imaging system Incucyte S3, allowing collection of data describing distinct characteristics of aggregate size and growth. The quantification of changes in total area of aggregates was used to define IC50 and MIC values of selected TB drugs which revealed that the cording phenotype grew more rapidly and displayed a higher susceptibility to rifampicin. A checkerboard approach, testing pair-wise combinations of sub-inhibitory concentrations of drugs, revealed rifampicin, linezolid and pretomanid as superior in inhibiting growth of cording phenotype. Conclusion Our results emphasize the efficiency of using automated live-cell imaging and its potential in high-through put whole-cell screening to evaluate existing and search for novel antimycobacterial drugs.
