Characterization and intracellular trafficking pattern of vacuoles containing Chlamydia pneumoniae in human epithelial cells

Chlamydiae are obligate intracellular pathogens that reside within a membrane-bound vacuole throughout their developmental cycle. In this study, the intraphagosomal pH of Chlamydia pneumoniae (Cpn) was qualitatively assessed, and the intracellular fate of the pathogen-containing vacuole and its interaction with endocytic organelles in human epithelial cells were analysed using conventional immunofluorescence and confocal microscopy. The pH-sensitive probes acridine orange (AO), LysoTracker (LyT) and DAMP did not accumulate in the bacterial inclusion. In addition, exposure of cells to bafilomycin A1 (BafA1), a potent acidification inhibitor, did not inhibit or delay chlamydial growth. The chlamydial compartment was not accessible to the fluid-phase tracer Texas Red (TR)-dextran and did not exhibit any level of staining for the late endosomal marker cation-independent mannose-6-phosphate receptor (Ci-M6PR) or for the lysosomal-associated membrane proteins (LAMP-1 and -2) and CD63. In addition, transferrin receptor (TfR)-enriched vesicles were observed close to Cpn vacuoles, potentially indicating a specific translocation of these organelles through the cytoplasm to the vicinity of the vacuole. We conclude that Cpn, like other chlamydial spp., circumvents the host endocytic pathway and inhabits a non-acidic vacuole, which is dissociated from late endosomes and lysosomes, but selectively accumulates early endosomes.