Transcription profiling of genes encoding components of the respiratory chain and the ATP synthesizing apparatus of Mycobacterium tuberculosis was conducted in vivo in the infected mouse lung, and in vitro in bacterial cultures subjected to gradual oxygen depletion and to nitric oxide treatment. Transcript levels changed dramatically as infection progressed from bacterial exponential multiplication (acute infection) to cessation of bacterial growth (chronic infection) in response to host immunity. The protonpumping type-I NADH dehydrogenase and the aa3-type cytochrome c oxidase were strongly down-regulated. Concurrently, the less energy-efficient cytochrome bd oxidase was transiently upregulated. The nitrate transporter NarK2 was also up-regulated, indicative of increased nitrate respiration. The reduced efficiency of the respiratory chain was accompanied by decreased expression of ATP synthesis genes. Thus, adaptation of M. tuberculosis to host immunity involves three successive respiratory states leading to decreased energy production. Decreased bacterial counts in mice infected with a cydC mutant (defective in the cytochrome bd oxidase-associated transporter) at the transition to chronic infection provided initial evidence that the bd oxidase pathway is required for M. tuberculosis adaptation to host immunity. In vitro, NO treatment and hypoxia caused a switch from transcription of type I to type II NADH dehydrogenase. Moreover, cytochrome bd oxidase expression increased, but cytochrome c oxidase expression decreased slightly (nitric oxide) or not at all (hypoxia). These specific differences in respiratory metabolism during M. tuberculosis growth arrest in vitro and in vivo will guide manipulation of in vitro conditions to model bacterial adaptation to host immunity. nitric oxide treatment ͉ transcriptional profiling ͉ dormancy ͉ hypoxia M ycobacterium tuberculosis is an airborne bacterial pathogen causing a chronic lung infection that passes through several stages. In most infected persons, host defenses either clear infection or drive it into a chronic latent state that is potentially long-lasting. Weakening of host immunity can result in release from latency and reactivation of disease. It has been suggested that various stages of M. tuberculosis infection are associated with different physiological states of the pathogen (1-5). Work with murine infection models shows that adaptation of M. tuberculosis to host immunity involves replacement of sugars by fatty acids as a carbon and energy source (6, 7). Respiration may also be an important aspect of energy metabolism involved in M. tuberculosis adaptation to host immunity, because respiration in bacteria is a flexible process that changes as microorganisms respond to environmental stresses (reviewed in refs. 8 and 9). However it is not known whether bacterial pathogens, including M. tuberculosis, reroute electron flow during the infection of a host animal.One model for studying M. tuberculosis adaptation to host immunity involves infecting mice with tubercle...

Mycobacterium tuberculosis is predicted to subsist on alternative carbon sources during persistence within the human host. Catabolism of odd-and branched-chain fatty acids, branched-chain amino acids, and cholesterol generates propionyl-coenzyme A (CoA) as a terminal, three-carbon (C 3 ) product. Propionate constitutes a key precursor in lipid biosynthesis but is toxic if accumulated, potentially implicating its metabolism in M. tuberculosis pathogenesis. In addition to the well-characterized methylcitrate cycle, the M. tuberculosis genome contains a complete methylmalonyl pathway, including a mutAB-encoded methylmalonyl-CoA mutase (MCM) that requires a vitamin B 12 -derived cofactor for activity. Here, we demonstrate the ability of M. tuberculosis to utilize propionate as the sole carbon source in the absence of a functional methylcitrate cycle, provided that vitamin B 12 is supplied exogenously. We show that this ability is dependent on mutAB and, furthermore, that an active methylmalonyl pathway allows the bypass of the glyoxylate cycle during growth on propionate in vitro. Mycobacterium tuberculosis is an obligate human pathogen that is expected to adapt metabolically to conditions that are often hostile and nutrient poor during successive cycles of infection, replication, persistence, and transmission. In particular, glucose deficiency and an abundance of fatty acids are thought to dictate mycobacterial metabolism during infection (3, 35), consistent with the complex repertoire of genes involved in lipid metabolism in the M. tuberculosis genome (10). Subsistence on fatty acids requires the sequential action of the catabolic ␤-oxidation cycle and, where glycolytic substrates are limiting, the anaplerotic glyoxylate cycle, which enables the assimilation of derivative two-carbon (C 2 ) acetyl-coenzyme A (CoA) subunits (37). In addition to producing acetyl-CoA, ␤-oxidation of odd-and branched-chain fatty acids yields the C 3 subunit propionyl-CoA. This metabolite can also be generated by the catabolism of branched-chain amino acids (24) and cholesterol. Recently, a cassette of genes involved in the catabolism of the A and B rings of cholesterol to propionyl-CoA, pyruvate, and other metabolites was identified in actinomycetes, including members of the M. tuberculosis complex (27,52). Although the relevance of cholesterol as a carbon source for M. tuberculosis in vivo has yet to be established, the likely action of this catabolic pathway during intracellular growth and survival of M. tuberculosis (52) suggests that it may constitute an additional, and potentially significant, source of propionylCoA in this pathogen.Propionyl-CoA is a key precursor in several lipid biosynthetic pathways in M. tuberculosis (28); however, while providing a high-energy metabolite, the accumulation of propionate is toxic to the cell, and as such, efficient mechanisms are required for its disposal (5). This dual nature implies a central role for propionate metabolism in the growth and persistence of M. tuberculosis in vivo (18,37). Evi...