Dr Nastassja Kriel, Stellenbosch University - VALIDATE Fellow
Identifying persister Mycobacterium tuberculosis biosignatures
Project Aims
Mycobacterium tuberculosis causes tuberculosis (TB) in people worldwide. The only TB vaccine available is only effective in preventing disease in children with diminishing effects over time. Infection and treatment of TB can result in the formation of bacterial sleeper cells, known as persisters. Persisters can survive antibiotic treatment and resume growth, resulting in the reoccurrence of disease after treatment. Persister bacteria are unique because the resistance they have to treatments is not caused by DNA changes. However, very little is known about how these bacteria influence the cells which protect the body against infections, allowing them to survive.
Bacteria export proteins out of the cell to re-program the body's defence systems and allow survival of persister bacteria. I will investigate which proteins are secreted by persister TB bacteria to re-program human defence cells for their long-term survival. I will go on to investigate which proteins are produced by human defence cells in response to infection with persister TB bacteria. I will do this by making use of a special form of TB, which when forming persister TB, makes red proteins. We will isolate these red TB bacteria and use state of the art molecular techniques and sophisticated computational analyses of data to provide new insights on how persister TB interact with human defence cells and how these cells respond to infection. Understanding the host-pathogen interactions between human defence cells and TB will provide new insights into how we can prevent persister formation. The identification of persister TB exported proteins will provide new anti-TB vaccine candidates, which target dormant infections and could provide long-term protection against TB.
Project Outcomes
Prolonged treatment is required to sterilize tuberculosis (TB) infections. Persister Mycobacterium tuberculosis (Mtb), a sub-population of viable but non-replicating (VBNR) Mtb that is phenotypically resistant to anti-TB treatment, may prevent TB sterilization. In addition to increasing the risk of treatment failure, persisters may contribute to disease recurrence following treatment. The environmental stress experienced during infection and treatment may contribute to the formation of persister Mtb sub-populations.
Mtb ensures long-term survival within the host by secreting proteins which modulate the host immune response. We hypothesized that actively replicating and VBNR Mtb may secrete a different subset of proteins. Persister secreted proteins can potentially be serve as biomarkers or vaccine development candidates for latent TB infections.
A drug sensitive clinical isolate of Mtb obtained from a patient who had an unsuccessful treatment outcome, was subjected to low pH stress to induce a VBNR sub-population. A fluorescence dilution replication reporter plasmid confirmed the formation of a VBNR sub-population following low pH stress. The cell lysates and culture filtrates of actively replicating and VBNR Mtb was characterized using mass spectrometry.
In total we identified 2937 proteins in the cell lysates of actively replicating and acid-stressed Mtb, of these, 335 proteins were suggested to be significantly differentially abundant between actively replicating and VBNR enriched Mtb cultures. Investigation of differentially regulated proteins in VBNR enriched cultures may provide valuable insights into the formation of VBNR sub-populations in Mtb.
The characterization of Mtb culture filtrates resulted in the identification of 274 proteins from acid-stressed and VBNR enriched Mtb cultures, of which 44 were not identified in the culture filtrates of actively replicating Mtb. Proteins identified in the culture filtrates of VBNR enriched cultures can be investigated as modulators of the immune response and may serve as vaccine development candidates for TB vaccines aimed at the prevention of latent TB infections.
Project Outputs
Find out more about Dr Nastassja Kriel here.
