SummaryThe expression of protective immunity to Mycobacterium tuberculosis in mice is mediated by T lymphocytes that secrete cytokines . These molecules then mediate a variety of roles, including the activation ofparasitized host macrophages, and the recruitment of other mononuclear phagocytes to the site of the infection in order to initiate granuloma formation . Among these cytokines, interferon y (IFN-y) is believed to play a key role is these events. In confirmation ofthis hypothesis, we show in this study that mice in which the IFN-y gene has been disrupted were unable to contain or control a normally sublethal dose of M. tuberculosis, delivered either intravenously or aerogenically. In such mice, a progressive and widespread tissue destruction and necrosis, associated with very high numbers ofacid-fast bacilli, was observed. In contrast, despite the lack of protective immunity, some DTH-like reactivity could still be elicited. These data, therefore, indicate that although IFN-y may not be needed for DTH expression, it plays a pivotal and essential role in protective cellular immunity to tuberculosis infection .C urrent murine models of experimental Mycobacterium tuberculosis infection indicate that the emergence of acquired immunity to this organism is mediated by populations of both class I and II MHC-restricted T lymphocytes, which secrete cytokines that result in the activation of parasitized macrophages and promotion of the granulomatous response (1-4) . The cytokine IFN-y, which is an effective inducer of antimicrobial mechanisms in several systems (5, 6) has been shown to inhibit the growth of mycobacteria in vitro (7-9), but its role in vivo has yet to be precisely defined. In this regard, we report here that mice in which the IFN-y gene has been disrupted (IFN-y gene knockout mice [GKO mice]), develop a fatal, disseminated form of disease when inoculated with a normally sublethal inoculum of M. tuberculosis . These results indicate, therefore, that IFN-'Y plays a pivotal role in the expression ofprotective immunity to this infection in the mouse. Moreover, this new immunodeficient mouse model may prove highly useful in the evaluation of therapeutic intervention strategies in the severely immunocompromised host.Mice. GKO mice were generated as described previously (10). Briefly, a normal IFN-y allele in mouse embryonic stem cells was replaced with a defective gene using a targeted vector which introduced a termination codon after the first 30 amino acids of the mature IFN-y protein . The altered stem cells were injected into C57BL/6J blastocysts and transmitted via the germline. Heterozygous offspring ofthe chimeras were intercrossed to generate mice homozygous for the altered (GKO) and wild type (WT) allele . The GKO mice were previously characterized as normal in terms of spleen and thymus cell number and expression of CD3, B220, CD4, and CD8 surface markers, and were shown to be incapable of IFN-y secretion (10).Experimental Infections. The Erdman strain ofM. tuberculosis was grown in Proskauer B...

Mycobacterium tuberculosis claims more human lives each year than any other bacterial pathogen. Infection is maintained in spite of acquired immunity and resists eradication by antimicrobials. Despite an urgent need for new therapies targeting persistent bacteria, our knowledge of bacterial metabolism throughout the course of infection remains rudimentary. Here we report that persistence of M. tuberculosis in mice is facilitated by isocitrate lyase (ICL), an enzyme essential for the metabolism of fatty acids. Disruption of the icl gene attenuated bacterial persistence and virulence in immune-competent mice without affecting bacterial growth during the acute phase of infection. A link between the requirement for ICL and the immune status of the host was established by the restored virulence of delta icl bacteria in interferon-gamma knockout mice. This link was apparent at the level of the infected macrophage: Activation of infected macrophages increased expression of ICL, and the delta icl mutant was markedly attenuated for survival in activated but not resting macrophages. These data suggest that the metabolism of M. tuberculosis in vivo is profoundly influenced by the host response to infection, an observation with important implications for the treatment of chronic tuberculosis.

The success of Mycobacterium species as pathogens depends on their ability to maintain an infection inside the phagocytic vacuole of the macrophage. Although the bacteria are reported to modulate maturation of their intracellular vacuoles, the nature of such modifications is unknown. In this study, vacuoles formed around Mycobacterium avium failed to acidify below pH 6.3 to 6.5. Immunoelectron microscopy of infected macrophages and immunoblotting of isolated phagosomes showed that Mycobacterium vacuoles acquire the lysosomal membrane protein LAMP-1, but not the vesicular proton-adenosine triphosphatase (ATPase) responsible for phagosomal acidification. This suggests either a selective inhibition of fusion with proton-ATPase-containing vesicles or a rapid removal of the complex from Mycobacterium phagosomes.

The progression of tuberculosis from a latent, sub-clinical infection to active disease that culminates in transmission of infectious bacilli is determined locally at the level of the granuloma. This progression takes place even in the face of a robust immune response that, while it contains infection, is unable to eliminate the bacterium. The factors or environmental conditions that influence this progression remain to be determined. Recent advances have indicated that pathogen-induced dysregulation of host lipid synthesis and sequestration plays a critical role in this transition. The foamy macrophage appears to be a key player in both sustaining persistent bacteria and contributing to the tissue pathology that leads to cavitation and release of infectious bacilli.