Early events in phagosome establishment are required for intracellular survival of Legionella pneumophila

Lawrence A. Wiater, Kenneth Dunn, Frederick R. Maxfield, Howard A. Shuman

Research output: Contribution to journalArticle

93 Citations (Scopus)

Abstract

During infection, the Legionnaires' disease bacterium, Legionella pneumophila, survives and multiplies within a specialized phagosome that is near neutral pH and does not fuse with host lysosomes. In order to understand the molecular basis of this organism's ability to control its intracellular fate, we have isolated and characterized a group of transposon-generated mutants which were unable to kill macrophages and were subsequently found to be defective in intracellular multiplication. These mutations define a set of 20 genes (19 icm [for intracellular multiplication] genes and dotA [for defect in organelle trafficking]). In this report, we describe a quantitative assay for phagosome-lysosome fusion (PLF) and its use to measure the levels of PLF in cells that have been infected with either wild-type L. pneumophila or one of several mutants defective in different icm genes or dotA. By using quantitative confocal fluorescence microscopy, PLF could be scored on a per- bacterium basis by determining the extent to which fluorescein-labeled L. pneumophila colocalized with host lysosomes prelabeled with rhodamine- dextran. Remarkably, mutations in the six genes that were studied resulted in maximal levels of PLF as quickly as 30 min following infection. These results indicate that several, and possibly all, of the icm and dotA gene products act at an early step during phagosome establishment to determine whether L. pneumophila-containing phagosomes will fuse with lysosomes. Although not ruled out, subsequent activity of these gene products may not be necessary for successful intracellular replication.

Original languageEnglish
Pages (from-to)4450-4460
Number of pages11
JournalInfection and Immunity
Volume66
Issue number9
StatePublished - 1998

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Legionella pneumophila
Phagosomes
Lysosomes
Organelles
Genes
Bacteria
Legionnaires' Disease
Mutation
Cell Fusion
Infection
Fluorescein
Fluorescence Microscopy
Confocal Microscopy
Macrophages

ASJC Scopus subject areas

  • Immunology

Cite this

Early events in phagosome establishment are required for intracellular survival of Legionella pneumophila. / Wiater, Lawrence A.; Dunn, Kenneth; Maxfield, Frederick R.; Shuman, Howard A.

In: Infection and Immunity, Vol. 66, No. 9, 1998, p. 4450-4460.

Research output: Contribution to journalArticle

Wiater, Lawrence A. ; Dunn, Kenneth ; Maxfield, Frederick R. ; Shuman, Howard A. / Early events in phagosome establishment are required for intracellular survival of Legionella pneumophila. In: Infection and Immunity. 1998 ; Vol. 66, No. 9. pp. 4450-4460.
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