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Mycobacterium abscessus is an emerging human pathogen responsible for lung infections, skin and soft-tissue infections and disseminated infections in immunocompromised patients. It may exist either as a smooth S or rough R morphotype, the latter being associated with increased pathogenicity in various models. Genetic tools for homologous recombination and conditional gene expression are desperately needed to allow the study of M. abscessus virulence. However, descriptions of knock-out KO mutants in M. abscessus are rare, with only one KO mutant from an S strain described so far. Moreover, of the three major tools developed for homologous recombination in mycobacteria, only the one based on expression of phage recombinases is working. Several conditional gene expression tools have recently been engineered for Mycobacterium tuberculosis and Mycobacterium smegmatis, but none have been tested yet in M. abscessus. Based on previous experience with genetic tools allowing homologous recombination and their failure in M. abscessus, we evaluated the potential interest of a conditional gene expression approach using a system derived from the two repressors system, TetR-PipOFF. After several steps necessary to adapt TetR-PipOFF for M. abscessus, we have shown the efficiency of this system for conditional expression of an essential mycobacterial gene, fadD32. Inhibition of fadD32 was demonstrated for both the S and R isotypes, with marginally better efficiency for the R isotype. Conditional gene expression using the dedicated TetR-PipOFF system vectors developed here is effective in S and R M. abscessus, and may constitute an interesting approach for future genetic studies in this pathogen.



Autor: Mélanie Cortes, Anil Kumar Singh, Jean-Marc Reyrat †, Jean-Louis Gaillard, Xavier Nassif, Jean-Louis Herrmann

Fuente: http://plos.srce.hr/



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