Large nonionic surfactants lack the properties of common detergents, but modulate lipids on the surface of M tuberculosis (MTB) and produce diverse effects on infection. Some surfactants injected into mice suppress infection while others enhance infection. Similarly, some surfactants added to culture media reduce the virulence of MTB while others enhance virulence. These changes persist through several cell divisions after removal of the surfactants. This project will exploit situations induced by these agents to investigate the host-MTB relationship. Our approach is based on several novel findings: 1) We discovered a surfactant, CRL1072, that is bactericidal for MTB in vivo even though it is only bacteriostatic in vitro. Its mechanisms include modulation of the permeability of organisms and enhancement of NO production, 2) We found CRL1072 and live MTB induce NO production by murine and human macrophages, 3) We found that a mycobacterial glycolipid, TDM, that can enhance MTB infection is active only when configured as a crystalline surface monolayer and demonstrated that TDM can be a target of surfactants, 4) We established murine models of acute, subacute and chronic MTB infection. The project will proceed via a series of targeted investigations. First, we will investigate whether CRL1072 enhances cell wall permeability of MTB and increases their susceptibility to NO. Next, using CRL1072 and ineffective surfactants as controls, we will investigate steps leading to the production of NO by MTB infected human cells by measuring NO, iNOS and iNOS mRNA. Electron microscopy will be used to assess lysosome-phagosomes fusion and related phenomena. Studies with lung macrophages from normal individuals and people with AIDS will be conducted to determine if findings in cell lines are applicable to normal or diseased human cells. In concurrent experiments, we will investigate the effects of CRL1072 on the host response during acute, chronic and dormant MTB infection concentrating correlations between host reactions and the permeability, lipid composition and - ultrastructure of the organisms. In continuing studies, surfactants that enhance MTB growth in mice and in macrophage culture will be produced and evaluated similarly. MTB will be cultured in media containing selected surfactants in an effort to induce organisms with enhanced as well as reduced virulence. These organisms will be used to test the hypothesis that changes in virulence are related to modifications of lipids on the surface of the organisms. Finally, we will recover and characterize organisms from animals with chronic and dormant infection and at intervals during reactivation of infection in order to__develop an improved understanding of dormancy and reactivation of tuberculosis.
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