A quantitative single-cell investigation of cell growth and antibiotic tolerance in Mycobacterium smegmatis

Tuberculosis is a disease which has afflicted humans for millennia and currently holds the dubious title of the greatest cause of mortality by any single infectious agent. Mycobacterium tuberculosis-the causative agent of tuberculosis-is exquisitely adapted to the infection of humans, with the ability to persist within the hostile intracellular environment of the macrophage. To enable this, M. tuberculosis enacts a dormancy programme that permits long-term survival. This programme has been linked to the requirement for six months treatment to cure tuberculosis since it may enhance drug tolerance-the ability for bacteria to survive antibiotics without genetic resistance. In this thesis, I extensively utilise single-cell time-lapse microscopy coupled with microfluidics to observe Mycobacterium smegmatis cells over many generations. I describe in detail a robust methodology for long-term time-lapse and subsequent image analysis workflow, including its application to a challenging dataset. I then investigate how single cells maintain their cell size under diverse environmental conditions, and determine that birth length and division length are intimately linked by a fixed average extension when grown in standard ... (continues)