Poster Presentation HUPO 2019 - 18th Human Proteome Organization World Congress

Associating H2O2 and NO related changes in the proteome of mycobacterium smegmatis with enhanced survival in macrophages (#530)

Naadir Ganief 1 , Jessica Sjoureman 1 , Claudia Albeldas 1 , Kehilwe C Nakedi 1 , Clemens Hermann 1 , Bridget Calder 1 , Jonathan M Blackburn 1 2 , Nelson C Soares 1
  1. Division of Chemical and Systems Biology, Department of Integrative Biomedical Sciences, University of Cape Town, Cape Town, Western Cape, South Africa
  2. Institute of Infectious Disease and Molecular Medicine, University of Cape Town, Cape Town, Western Cape, South Africa


Mycobacterium tuberculosis is typically spread through aerosolised droplets containing the bacilli1. These bacilli are thought to originate from ruptured granulomas in the lung where they may multiply for extended periods, while experiencing sub-lethal stresses including oxidative and nitrosative insult1,2,3. Transmitted bacilli may, therefore, exist in a biochemical state typically not observed during routine culture conditions. This work4 aims to understand the potential role of these stresses on the infectivity and survival of Mycobacteria by correlating the proteomic response and survival of Mycobacteria following infection in-vitro.


Mycobacterium smegmatis was cultured with sub-lethal doses of H2O2, DETA-NO or without stress and the proteome was measured using label-free shotgun LC-MS/MS, at three time points corresponding to: onset of, recovery from, and post recovery from stress. Similarly, bacteria from these time points were used to infect Raw264.7 murine macrophage like cells; uptake and survival were quantified 3 hours and 24 hours post infection, respectively. Proteomic data was processed and analysed using MaxQuant, Perseus, StringDB, KEGG and Cytoscape.


We observed that sub-lethal doses of H2O2 or DETA-NO increased Mycobacterial survival in macrophages by up to 9-fold with DETA-NO able to induce the most resistant phenotype. Proteomic analysis identified and quantified 3336 protein. Pathway analysis revealed, that sub-lethal challenge with DETA-NO or H2O2, induced expression of the DevR regulon, and other known virulence factors. Furthermore, DETA-NO altered regulation in lipid metabolism, which may implicate the cell wall in increased survival.


These findings indicate a large overlap between the response to sub-lethal doses of H2O2 or DETA-NO, which may account for the increased survival observed. Additionally, we correlated an altered lipid metabolism with the higher rates of survival observed for bacteria exposed to DETA-NO challenge. This work highlights the need to study Mycobacteria under physiological conditions and may highlight potential therapies.

  1. Gengenbacher, M., & Kaufmann, S. H. E. (2012). Mycobacterium tuberculosis: Success through dormancy. FEMS Microbiology Reviews, 36, 514–532.
  2. (Ehrt & Schnappinger, 2009; Ganief et al., 2018; Gengenbacher & Kaufmann, 2012; Gouzy, Poquet, & Neyrolles, 2014)Ehrt, S., & Schnappinger, D. (2009). Mycobacterial survival strategies in the phagosome: Defence against host stresses. Cellular Microbiology, 11(May), 1170–1178.
  3. Gouzy, A., Poquet, Y., & Neyrolles, O. (2014). Amino acid capture and utilization within the Mycobacterium tuberculosis phagosome. Future Microbiology, 9(MAY 2014), 631–637.
  4. Ganief, N., Sjouerman, J., Albeldas, C., Nakedi, K. C., Hermann, C., Calder, B., … Soares, N. C. (2018). Associating H 2 O 2- and NO-related changes in the proteome of Mycobacterium smegmatis with enhanced survival in macrophage. Emerging Microbes & Infections.