No abstract

Mycobacterium tuberculosis is an important pathogen of mammals that relies on 2-hydroxyphenyloxazoline-containing siderophore molecules called mycobactins for the acquisition of iron in the restrictive environment of the mammalian macrophage. These compounds have been proposed to be biosynthesized through the action of a cluster of genes that include both nonribosomal peptide synthase and polyketide synthase components. One of these genes encodes a protein, MbtB, that putatively couples activated salicylic acid with serine or threonine and then cyclizes this precursor to the phenyloxazoline ring system. We have used gene replacement through homologous recombination to delete the mbtB gene and replace this with a hygromycin-resistance cassette in the virulent strain of M. tuberculosis H37Rv. The resulting mutant is restricted for growth in iron-limited media but grows normally in ironreplete media. Analysis of siderophore production by this organism revealed that the biosynthesis of all salicylate-derived siderophores was interrupted. The mutant was found to be impaired for growth in macrophage-like THP-1 cells, suggesting that siderophore production is required for virulence of M. tuberculosis. These results provide conclusive evidence linking this genetic locus to siderophore production.

The pathogenesis of tuberculosis involves multiple phases and is believed to involve both a carefully deployed series of adaptive bacterial virulence factors and inappropriate host immune responses that lead to tissue damage. A defined Mycobacterium tuberculosis mutant strain lacking the sigH-encoded transcription factor showed a distinctive infection phenotype. In resistant C57BL͞6 mice, the mutant achieved high bacterial counts in lung and spleen that persisted in tissues in a pattern identical to those of wild-type bacteria. Despite a high bacterial burden, the mutant produced a blunted, delayed pulmonary inflammatory response, and recruited fewer CD4 ؉ and CD8 ؉ T cells to the lung in the early stages of infection. In susceptible C3H mice, the mutant again showed diminished immunopathology and was nonlethal at over 170 days after intravenous infection, in contrast to isogenic wild-type bacilli, which killed with a median time to death of 52 days. Complete genomic microarray analysis revealed that M. tuberculosis sigH may mediate the transcription of at least 31 genes directly and that it modulates the expression of about 150 others; the SigH regulon governs thioredoxin recycling and may be involved in the maintenance of intrabacterial reducing capacity. These data show that the M. tuberculosis sigH gene is dispensable for bacterial growth and survival within the host, but is required for the production of immunopathology and lethality. This phenotype demonstrates that beyond an ability to grow and persist within the host, M. tuberculosis has distinct virulence mechanisms that elicit deleterious host responses and progressive pulmonary disease.T uberculosis is one of the leading infectious causes of death and claims Ϸ2 million lives annually (1). There is controversy over whether the disease is primarily a dysfunctional immunologic reaction to a persistent microbe or whether the bacteria themselves produce tissue damage; there is evidence that both host and bacterial factors play key roles in disease severity. In susceptible mouse strains, such as C3H, the pathogen elicits a dysregulated, necrotizing host immune response leading to tissue destruction and further bacterial replication. In resistant mice such as C57BL͞6, Mycobacterium tuberculosis survives in high numbers for many months and is contained in organized granulomatous lesions in the lung without progressive lung damage. Thus whereas mycobacteria survive in both genetic backgrounds, disease progression is delayed in resistant animals (2-4).On the bacterial side, M. tuberculosis virulence has been associated with its initial survival within macrophages and resistance to reactive oxygen and nitrogen intermediates (ROIs and RNIs) (5-8). Tubercle bacilli demonstrate inducible responses to oxidative stresses, and several M. tuberculosis genes, including katG (catalase peroxidase), ahpC (alkylhydroperoxide reductase), and sodA and sodC (superoxide dismutases) have been implicated in protection from the macrophage oxidative burst (9-11). Another potential ...

AN OVERVIEW OF IRON METABOLISM Iron is a prerequisite for in vitro growth of mycobacteria.