Recent studies have shown that matrix metalloproteinases (MMPs) are induced by Mycobacterium tuberculosis during pulmonary infection. Here, expression of MMP-9 during pulmonary M. tuberculosis infection was characterized to determine whether its production correlated with disease resistance in vivo and to determine what role, if any, MMP-9 might have in granuloma formation. Following aerosol infection with M. tuberculosis, dissemination of bacilli occurred earlier in the C57BL/6 resistant mouse strain than in the susceptible CBA/J strain, as was evident from an increased number of bacteria in the blood, spleen, and liver at day 14 after infection. In addition, early dissemination of the bacilli was associated with early induction of protective immunity as assessed from gamma interferon levels. Nonspecific blocking of MMPs in C57BL/6 mice early during infection reduced hematogenous spread of the bacilli, suggesting that MMPs indeed play a role in facilitating dissemination, likely via extracellular matrix degradation. The concentration of active MMP-9, specifically, was greater in the lungs of C57BL/6 mice than in those of the CBA/J mice at day 28, thereby suggesting that MMP-9 is not one of the MMPs directly involved in promoting early dissemination of M. tuberculosis. Instead, however, histological lung sections and flow cytometric analysis of lung cells from MMP-9-knockout mice showed that MMP-9 is involved in macrophage recruitment and granuloma development. These combined data support the idea that early MMP activity is an essential component of resistance to pulmonary mycobacterial infection and that MMP-9, specifically, is required for recruitment of macrophages and tissue remodeling to allow for the formation of tight, well-organized granulomas.