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Infection and Immunity, October 2002, p. 5800-5807, Vol. 70, No. 10
0019-9567/02/$04.00+0     DOI: 10.1128/IAI.70.10.5800-5807.2002
Copyright © 2002, American Society for Microbiology. All Rights Reserved.

The Mycobacterium tuberculosis Phagosome in Human Macrophages Is Isolated from the Host Cell Cytoplasm

Daniel L. Clemens,* Bai-Yu Lee, and Marcus A. Horwitz

Division of Infectious Diseases, Department of Medicine, UCLA School of Medicine, Center for Health Sciences, Los Angeles, California 90095

Received 25 March 2002/ Returned for modification 31 May 2002/ Accepted 5 July 2002

Knowledge of whether Mycobacterium tuberculosis resides within a relatively impermeable membrane-bound vacuole or is free within the cytoplasm within its host cell is central to an understanding of the immunobiology of this intracellular parasite but is a matter of controversy. To explore this issue, we assessed the accessibility of medium-size protein molecules (Fab fragments of 50,000 Da) to M. tuberculosis within human macrophages. We infected the macrophages with wild-type or green fluorescent protein-expressing M. tuberculosis, microinjected Fab fragments directed against a major surface antigen of M. tuberculosis into the host cell, and assayed the accessibility of the bacteria to the Fab fragments by both immunofluorescence microscopy and immunogold electron microscopy. Whereas microinjected intact immunoglobulin G molecules against cytoplasmic early endosomal antigen 1 readily stained this antigen, microinjected Fab fragments against M. tuberculosis did not stain the bacterium within its phagosome. In contrast, microinjected Fab fragments against Listeria monocytogenes, an intracellular bacterium known to permeabilize its phagosomal membrane, strongly stained this bacterium. Our study shows that M. tuberculosis resides in an isolated phagosome that is relatively impermeable to cytoplasmic constituents.

* Corresponding author. Mailing address: Division of Infectious Diseases, Dept. of Medicine, UCLA School of Medicine, CHS 37-121, Los Angeles, CA 90095. Phone: (310) 825-9324. Fax: (310) 794-7156. E-mail: dclemens{at}mednet.ucla.edu.

Editor: S. H. E. Kaufmann

Infection and Immunity, October 2002, p. 5800-5807, Vol. 70, No. 10
0019-9567/02/$04.00+0     DOI: 10.1128/IAI.70.10.5800-5807.2002
Copyright © 2002, American Society for Microbiology. All Rights Reserved.



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