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Identification of genetic loci implicated in the survival of Mycobacterium smegmatis in human mononuclear phagocytes
Author(s) -
Lagier Béatrice,
Pelicic Vladimir,
Lecossier Denise,
Prod'hom Guy,
Rauzier Jean,
Guilhot Christophe,
Gicquel Brigitte,
Hance Allan J.
Publication year - 1998
Publication title -
molecular microbiology
Language(s) - English
Resource type - Journals
SCImago Journal Rank - 1.857
H-Index - 247
eISSN - 1365-2958
pISSN - 0950-382X
DOI - 10.1046/j.1365-2958.1998.00933.x
Subject(s) - biology , mutant , complementation , mycobacterium smegmatis , gene , intracellular , mycobacterium tuberculosis , genetics , mycobacterium , peripheral blood mononuclear cell , microbiology and biotechnology , tuberculosis , bacteria , in vitro , medicine , pathology
A luminescence‐based procedure that permits the rapid evaluation of the survival of mycobacteria within mononuclear phagocytes was developed and used to screen insertional mutants of Mycobacterium smegmatis for their ability to survive in human monocyte‐derived macrophages. Among the 5000 mutants tested, eight mutants were identified that demonstrated impaired intracellular survival in human macrophages but that grew normally in the absence of cells. For each mutant, a portion of the gene interrupted by the transposition event was amplified by ligand‐mediated PCR and sequenced. In all cases, the existence of homologous genes of as yet unknown function were identified in the Mycobacteium tuberculosis genome. Complementation of the mutant mycobacterial strains with cosmids containing the homologous loci from M. tuberculosis restored normal intracellular growth in three of the four mutants tested, supporting the idea that these loci contain genes that are important for intracellular survival. This study demonstrates the feasibility of directly screening mutant mycobacterial strains to identify genes coding for activities necessary for the intracellular survival in human mononuclear phagocytes, an important initial step in the identification of potential targets for new therapeutic agents.