Tuberculosis (TB) kills nearly 2 million people each year, and, together with HIV, Mycobacterium tuberculosis top the WHO list of deaths caused by infectious agents. In HIV infected individuals, Mycobacterium avium complex (MAC) is a major cause of second ary lethal infections. The only vaccine currently available, BCG, provides protection against childhood TB but not against adult pulmonary TB. One hypothesis is that BCG-vaccination is inefficient in people already sensitized to mycobacteria. Two types of vaccines may therefore be warranted: 1) An improved live mycobacterial vaccine to replace BCG for neonate vaccination (non-sensitized group), and 2) a booster subunit vaccine aimed at restoring protective immunity to sensitized people. Our proposal addre sses these aspects:The intracellular life of the mycobacterium encompasses two aspects. It resides inside macrophages in early phagosomes where it avoids detection, and while it lives there it needs nutrition. We believe that these two processes are coup led. The sensible strategy is thus to study both simultaneously. By rationally manipulating the genome of the mycobacterium, we can achieve insight in both how they escape destruction and how they survive. The block of phagolysosome maturation results in evasion from Toll-like receptors and the antigen presentation machinery. Using antigen presentation as read-out we can identify the most important mycobacterial genes responsible for the immune evasion and thus rationally design vaccine variants. Understa nding the role of TLRs in recognition and antigen-presentation of mycobacterial adjuvant antigens will create a basis for subunit booster vaccines. With this application we seek to expand our group with knowledge in adaptive immunity and vaccine efficienc y, and to support continuing work on mycobacterial genetics, innate immunity, infection immunology and imaging.