Abstract

Mycobacterium tuberculosis infects billions of people worldwide and kills more than 1.5 million per year. TB remains extremely difficult to treat with antibiotics, requiring months to years of therapy for cure. The variable course of disease and treatment response suggests that functionally heterogeneous populations of mycobacteria respond differently stress. Using a quantitative single-cell approach, we show that mycobacteria deterministically generate diversity in their growth characteristics through an asymmetric growth pattern. Coupled with a cell cycle regulated by time and not size, this asymmetry creates subpopulations of cells with distinct growth rates and cell sizes that are differentially susceptible to clinically relevant classes of antibiotics. Thus, the unusual growth pattern intrinsic to mycobacteria deterministically creates a diverse population structure that may underlie phenotypes previously thought to be controlled by external stressors. We have also observed variation among microcolonies in antibiotic susceptibility that cannot be explained by growth pole age. We have established markers of cell cycle state to test the hypothesis that cell cycle state is another determinant of drug susceptibility. Armed with new image analysis algorithms, reporters of cell state, and mathematical models, we seek to quantitatively characterize and pharmacologically target drug tolerant mycobacterial subpopulations.