Characterization of intracellular survival of Mycobacterium avium subspecies paratuberculosis in J774 murine macrophages

Mycobacterium avium subspecies paratuberculosis (M. a. ptb) is the causative agent of Johne's disease, a progressive diarrheal disease of wild and domestic ruminants. Like other pathogenic species of mycobacteria, M. a. ptb is an intracellular pathogen of monocytes and monocyte-derived macrophages. In this dissertation we tested the hypothesis that M. a. ptb survives within macrophages by inhibition of acidification and maturation of the phagosomal compartment in which it resides.;Chapter 1 consists of a general introduction and a literature review of recent reports concerning normal phagosomal maturation and maturation of the mycobacterial phagosome. Chapter 2 characterizes acidification of the M. a. ptb phagosomal compartment in infected J774 cells. In this study we determined that the phagosomes of M. a. ptb have diminished ability to become acidified. In Chapter 3 we evaluated maturation of the M. a. ptb phagosome by identifying two stage specific phagosomal markers, the transferrin receptor (TFR) and lysosome associated membrane protein-1 (Lamp-1). The phagosomal compartment containing M. a. ptb had reduced amounts of Lamp-1, and increased amounts of TRF, which indicates that the M. a. ptb phagosome retains characteristics of an early phagosome and does not mature into later phagosomal stages. In Chapter 4 we characterized the effects of interferon gamma (IFN-gamma) and lipopolysaccharide (LPS) on development of the M. a. ptb phagosome in infected J774 cells. We determined that IFN-gamma/LPS treated J774 cells have increased ability to cause maturation and acidification of the M. a. ptb phagosomal compartment.;In this dissertation we determined that M. a. ptb survives within J774 cells by residing within a phagosomal compartment that has diminished ability to become acidified and resists normal phagosomal maturation. In addition, treatment of J774 cells with IFN-gamma/LPS increases their ability to cause acidification and maturation of the M. a. ptb phagosome.