Construction and Characterization of aMycobacterium tuberculosisMutant Lacking the Alternate Sigma Factor Gene,sigF

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Abstract
The alternate RNA polymerase sigma factor gene,sigF, which is expressed in stationary phase and under stress conditions in vitro, has been deleted in the virulent CDC1551 strain ofMycobacterium tuberculosis. The growth rate of the ΔsigFmutant was identical to that of the isogenic wild-type strain in exponential phase, although in stationary phase the mutant achieved a higher density than the wild type. The mutant showed increased susceptibility to rifampin and rifapentine. Additionally, the ΔsigFmutant displayed diminished uptake of chenodeoxycholate, and this effect was reversed by complementation with a wild-typesigFgene. No differences in short-term intracellular growth between mutant and wild-type organisms within human monocytes were observed. Similarly, the organisms did not differ in their susceptibilities to lymphocyte-mediated inhibition of intracellular growth. However, mice infected with the ΔsigFmutant showed a median time to death of 246 days compared with 161 days for wild-type strain-infected animals (P< 0.001). These data indicate thatM. tuberculosis sigFis a nonessential alternate sigma factor both in axenic culture and for survival in macrophages in vitro. While the ΔsigFmutant produces a lethal infection of mice, it is less virulent than its wild-type counterpart by time-to-death analysis.