The Intracellular Life of Mycobacterium avium subsp. paratuberculosis During Macrophage Stress: A Key Role for the Alternative Sigma Factor, SigH. Mycobacterium avium subsp. paratuberculosis K-10

Mycobacterium avium subspecies paratuberculosis (a.k.a. M. paratuberculosis) causes Johne’s disease, an enteric infection in cattle and other ruminants, greatly afflicting the dairy industry worldwide. Once inside the cell, M. paratuberculosis is known to survive harsh microenvironments, especially those inside activated macrophages. To improve our understanding of M. paratuberculosis pathogenesis, we examined the phagosome maturation associated with transcriptional responses of M. paratuberculosis during macrophage infection. Monitoring cellular markers, only live M. paratuberculosis bacilli were able to prevent phagosome maturation and reduce its acidification. On the transcriptional level, over 300 of M. paratuberculosis genes were significantly, differentially regulated in both naïve and IFN-γ-activated macrophages. These genes include the sigma factor H (sigH) that was shown to be important during persistent infection in M. tuberculosis. Interestingly, a sigH-knockout mutant showed increased sensitivity to a sustained level of thiol-specific oxidative stress. Large-scale RNA sequence analysis revealed that a large number of genes belong to the sigH regulon, especially following diamide stress. Genes involved in oxidative stress and virulence were among the induced genes in the sigH regulon with a putative consensus sequence for SigH binding was recognized in a subset of these genes (N=30), suggesting direct regulation with SigH. Finally, mice infections showed a significant attenuation of the ∆sigH mutant compared to its parental strain suggesting a role for sigH in M. paratuberculosis virulence. Such analysis could identify potential targets for further testing as vaccine candidates against Johne’s disease. Overall design: Examination of role played by alternative sigma factor, SigH during M. paratuberculosis infection.