DIFFERENTIAL TRAFFICKING OF LIVE AND DEAD MYCOBACTERIUM-MARINUM ORGANISMS IN MACROPHAGES

We characterized the Mycobacterium marinum phagosome by using a variet y of endocytic markers to follow the path of the bacteria through a mo use macrophage cell line. Using a laser confocal microscope, we found that the majority of viable M. marinum cells were in nonacidic vacuole s that did not colocalize with the vacuolar proton ATPase (V-ATPase), the calcium-independent mannose-6-phosphate receptor (CI-M6PR), or cat hepsin D. In contrast, heat-killed organisms and latex beads were in a cidic vacuoles which contained the V-ATPase, the CI-M6PR, and cathepsi n D. A population of vesicles that contained live M. marinum labeled w ith the lysosomal glycoprotein LAMP-1, but the percentage of vacuoles that labeled was lower than for heat-killed organisms or latex beads. When testing live and heat-killed Mycobacterium tuberculosis, we found levels of colocalization with LAMP-1 and cathepsin D comparable to th ose for the M. marinum isolate. We conclude that M. marinum, like M. t uberculosis, can circumvent the host endocytic pathway and reside in a n intracellular compartment which is not acidic and does not fuse,vith lysosomes. In addition, we describe a system for sampling a large pop ulation of intracellular organisms by using a laser confocal microscop e.