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Background  Bacteria of the suborder Corynebacterineae include significant human pathogens such as  Mycobacterium tuberculosis  and  M. leprae . Drug resistance in mycobacteria is increasingly common making identification of new antimicrobials a priority. Mycobacteria replicate intracellularly, most commonly within the phagosomes of macrophages, and bacterial proteins essential for intracellular survival and persistence are particularly attractive targets for intervention with new generations of anti-mycobacterial drugs.    Methodology/Principal Findings  We have identified a novel gene that, when inactivated, leads to accelerated death of  M. smegmatis  within a macrophage cell line in the first eight hours following infection. Complementation of the mutant with an intact copy of the gene restored survival to near wild type levels. Gene disruption did not affect growth compared to wild type  M. smegmatis  in axenic culture or in the presence of low pH or reactive oxygen intermediates, suggesting the growth defect is not related to increased susceptibility to these stresses. The disrupted gene,  MSMEG_5817 , is conserved in all mycobacteria for which genome sequence information is available, and designated  Rv0807  in  M. tuberculosis . Although homology searches suggest that MSMEG_5817 is similar to the serine:pyruvate aminotransferase of  Brevibacterium linens  suggesting a possible role in glyoxylate metabolism, enzymatic assays comparing activity in wild type and mutant strains demonstrated no differences in the capacity to metabolize glyoxylate.    Conclusions/Significance    MSMEG_5817  is a previously uncharacterized gene that facilitates intracellular survival of mycobacteria. Interference with the function of MSMEG_5817 may provide a novel therapeutic approach for control of mycobacterial pathogens by assisting the host immune system in clearance of persistent intracellular bacteria.

Identification of a Novel Gene Product That Promotes Survival of Mycobacterium smegmatis in Macrophages