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Infection and Immunity, January 2001, p. 486-493, Vol. 69, No. 1
0019-9567/01/$04.00+0   DOI: 10.1128/IAI.69.1.486-493.2001
Copyright © 2001, American Society for Microbiology. All rights reserved.

Brucella suis-Impaired Specific Recognition of Phagosomes by Lysosomes due to Phagosomal Membrane Modifications

Aroem Naroeni,1 Nicolas Jouy,2 Safia Ouahrani-Bettache,1 Jean-Pierre Liautard,1 and Françoise Porte1,*

Institut National de la Santé et de la Recherche Médicale U-4311 and Laboratoire Génie Biologique Science des Aliments-Unité Physiologie et Technologie des Végétaux,2 Université Montpellier II, Montpellier, France

Received 7 July 2000/Returned for modification 25 September 2000/Accepted 17 October 2000

Brucella species are gram-negative, facultatively intracellular bacteria that infect humans and animals. These organisms can survive and replicate within a membrane-bound compartment in phagocytic and nonprofessional phagocytic cells. Inhibition of phagosome-lysosome fusion has been proposed as a mechanism for intracellular survival in both types of cells. However, the biochemical mechanisms and microbial factors implicated in Brucella maturation are still completely unknown. We developed two different approaches in an attempt to gain further insight into these mechanisms: (i) a fluorescence microscopy analysis of general intracellular trafficking on whole cells in the presence of Brucella and (ii) a flow cytometry analysis of in vitro reconstitution assays showing the interaction between Brucella suis-containing phagosomes and lysosomes. The fluorescence microscopy results revealed that fusion properties of latex bead-containing phagosomes with lysosomes were not modified in the presence of live Brucella suis in the cells. We concluded that fusion inhibition was restricted to the pathogen phagosome and that the host cell fusion machinery was not altered by the presence of live Brucella in the cell. By in vitro reconstitution experiments, we observed a specific association between killed B. suis-containing phagosomes and lysosomes, which was dependent on exogenously supplied cytosol, energy, and temperature. This association was observed with killed bacteria but not with live bacteria. Hence, this specific recognition inhibition seemed to be restricted to the pathogen phagosomal membrane, as noted in the in vivo experiments.

* Corresponding author. Mailing address: INSERM U-431, Université Montpellier II, C.P. 100, Pl. E. Bataillon, 34095 Montpellier, France. Phone: (33) 4 67 14 42 38. Fax: (33) 4 67 14 33 38. E-mail: porte{at}crit.univ-montp2.fr.

Infection and Immunity, January 2001, p. 486-493, Vol. 69, No. 1
0019-9567/01/$04.00+0   DOI: 10.1128/IAI.69.1.486-493.2001
Copyright © 2001, American Society for Microbiology. All rights reserved.


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