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Infection and Immunity, January 2002, p. 69-78, Vol. 70, No. 1
0019-9567/01/$04.00+0     DOI: 10.1128/IAI.70.1.69-78.2002
Copyright © 2002, American Society for Microbiology. All Rights Reserved.

icmT Is Essential for Pore Formation-Mediated Egress of Legionella pneumophila from Mammalian and Protozoan Cells

Maelle Molmeret,1 O. A. Terry Alli,1 Steven Zink,1 Antje Flieger,2 Nicholas P. Cianciotto,2 and Yousef Abu Kwaik1*

Department of Microbiology and Immunology, The University of Kentucky College of Medicine, Lexington, Kentucky 40536,1 Department of Microbiology and Immunology, Northwestern University Medical School, Chicago, Illinois 606112

Received 2 July 2001/ Returned for modification 19 September 2001/ Accepted 2 October 2001

The final step of the intracellular life cycle of Legionella pneumophila and other intracellular pathogens is their egress from the host cell after termination of intracellular replication. We have previously isolated five spontaneous mutants of L. pneumophila that replicate intracellularly similar to the wild-type strain but are defective in pore formation-mediated cytolysis and egress from mammalian and protozoan cells, and the mutants have been designated rib (release of intracellular bacteria). Here, we show that the rib mutants are not defective in the activity of enzymes secreted through the type II secretion system, including phospholipase A, lysophospholipase A, and monoacylglycerol lipase, although they are potential candidates for factors that lyse host cell membranes. In addition, the pilD and lspG mutants, which are defective in the type II secretion system, are not defective in the pore-forming toxin. We show that all five rib mutants have an identical point mutation (deletion) following a stretch of poly(T) in the icmT gene. Spontaneous revertants of the rib mutants, due to an insertion of a nucleotide following the poly(T) stretch in icmT, have been isolated and shown to have regained the wild-type phenotype. We constructed an icmT insertion mutant (AA100kmT) in the chromosome of the wild-type strain by allelic exchange. The AA100kmT mutant was as defective as the rib mutant in pore formation-mediated cytolysis and egress from mammalian and protozoan cells. Both the rib mutant and the AA100kmT mutant were complemented by the icmT gene for their phenotypic defect. rtxA, a gene that is thought to have a minor role in pore formation, was not involved in pore formation-mediated cytolysis and egress from mammalian and protozoan cells. We conclude that the icmT gene is essential for pore formation-mediated lysis of mammalian and protozoan cells and the subsequent bacterial egress.

* Corresponding author. Mailing address: Department of Microbiology and Immunology, University of Kentucky Chandler Medical Center, Lexington, KY 40536-0084. Phone: (859) 323-3873. Fax: (859) 257-8994. E-mail: yabukw{at}pop.uky.edu.

Editor: J. T. Barbieri

Infection and Immunity, January 2002, p. 69-78, Vol. 70, No. 1
0019-9567/01/$04.00+0     DOI: 10.1128/IAI.70.1.69-78.2002
Copyright © 2002, American Society for Microbiology. All Rights Reserved.



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