Final Activity Report Summary - MACROPHAGE GENOMICS (Genomic Screens to Identify and Characterize Mechanisms of Mycobacterial Killing by Macrophages.) Mycobacteria are opportunistic microbes that reside and multiply in host macrophages, mainly by inhibition of host pro-inflammatory response. NF-kB is a crucial pro-inflammatory transcription factor that is inhibited by pathogenic mycobacteria along with its down-stream cytokine TNFa. Microarray analysis of cells infected with non-pathogenic m. smegmatis revealed that a number of genes were up-regulated, most strikingly TNFa and Rab proteins involved in membrane fusion, in comparison to control un-infected cells. Quantitative-PCR analysis confirmed these micorarray results. TNFa was released rapidly from cells infected with the non-pathogen m. smegmatis as compared to infection with pathogenic m. avium or control un-infected cells, as observed by enzyme-linked immunosorbent assay (ELISA). Furthermore, the addition of recombinant TNFa led to mycobacteria killing whereas treatment with TNFa -protease inhibitor led to dramatic growth in mycobacterial survival. This growth was also achieved when the cells were incubated with TNF-RI blocking antibodies. However, blocking with TNF-RII had only marginal effect. By immunoflourescence microscopy, we showed that addition of TNFa led to more phago-lysosome fusion, thus to more killing of mycobacteria.