Visualising the subcellular distribution of antibiotics against tuberculosis

Tuberculosis (TB), caused by the intracellular pathogen Mycobacterium tuberculosis (Mtb), remains the world's deadliest infectious disease. Although treatable, effective chemotherapy requires at least six months of treatment with a minimum of four antibiotics. Novel antibiotics are needed to quell the pandemic. However, we do not fully understand why current treatments take so long to work in patients. Mtb has a dynamic intracellular lifestyle, and this thesis tests the hypothesis that not all antibiotics penetrate into, or are effective within, all compartments containing Mtb during infection. Our understanding of the intracellular pharmacokinetics of drugs against TB has been limited by a lack of technologies for studying the subcellular distribution of antibiotics. This work developed a correlative imaging workflow incorporating fluorescence, electron and nanoscale ion microscopy (CLEIM) to map the subcellular distribution of two antibiotics, bedaquiline (BDQ) and pyrazinamide (PZA), at sub-micrometre resolution in Mtb-infected human macrophages. This workflow was complemented with orthogonal methods, including high-content live-cell imaging, to study the dynamic processes that contribute to antibiotic ... (continues)