This subproject is one of many research subprojects utilizing the resources provided by a Center grant funded by NIH/NCRR. Primary support for the subproject and the subproject's principal investigator may have been provided by other sources, including other NIH sources. The Total Cost listed for the subproject likely represents the estimated amount of Center infrastructure utilized by the subproject, not direct funding provided by the NCRR grant to the subproject or subproject staff. The basic mechanisms employed by M. tuberculosis (Mtb) to successfully infect and persist in mammalian lungs are poorly understood. DeADMAn (Designer Arrays for Defined Mutant Analysis) represents a high-throughput approach for subtractive identification of mutants attenuated for survival in animal tissues. Monkeys infected with Mtb represent an excellent model of TB - they exhibit pathology remarkably similar to humans and develop human-like latent TB. We conducted experiments with the DeADMAn technology in monkeys. This is the first report of transposon mutagenesis being utilized to study virulence in a bacterial pathogen using the macaque model. We identified over 100 genes, mutants which were attenuated for in-vivo growth in macaque lungs, in this model of acute TB. One of the major results of our study was that almost 33% of all mutants were attenuated in NHP lungs relative to mice, where only 6-8% of the tested mutants were attenuated. Some of these genes have previously been shown to be implicated in having a role during growth in macrophage or mouse studies. We also identified numerous novel genes with no previous report of being important for in-vivo growth. These include the cadI, cobL, drrA, lpqP, fadD21, cycA, yrbE3B, lipU, lprO, lppV, mce1, mce3 and mce4 genes. We have now been funded to study these differences using the identical mutant screen, but in a model of latent disease. These studies will shed light on the differences in virulence-mechanisms employed by tubercle bacilli for infecting various animal models.
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