Stationary phase gene expression of Mycobacterium tuberculosis following a progressive nutrient depletion: a model for persistent organisms?

Abstract

SummaryThe majority of individuals infected with TB develop a latent infection, in which organisms survive within the body while evading the host immune system. Such persistent bacilli are capable of surviving several months of combinatorial antibiotic treatment. Evidence suggests that stationary phase bacteria adapt to increase their tolerance to environmental stresses. We have developed a unique in vitro model of dormancy based on the characterization of a single, large volume fermenter culture of M. tuberculosis, as it adapts to stationary phase. Cells are maintained in controlled and defined aerobic conditions (50% dissolved oxygen tension), using probes that measure dissolved oxygen tension, temperature, and pH. Microarray analysis has been used in conjunction with viability and nutrient depletion assays to dissect differential gene expression. Following exponential phase growth the gradual depletion of glucose/glycerol resulted in a small population of survivors that were characterized for periods in excess of 100 days. Bacilli adapting to nutrient depletion displayed characteristics associated with persistence in vivo, including entry into a non-replicative state and the up-regulation of genes involved in β-oxidation of fatty acids and virulence. A reduced population of non-replicating bacilli went on to adapt sufficiently to reinitiate cellular division.

Global transcriptional response to vancomycin in Mycobacterium tuberculosis

“…Alternatively, the sulfuryl moiety of PAPS can be reduced to sulfide and incorporated into cysteine and, from there, into other reduced-sulfur metabolites. It has been shown previously that the cysDNC operon is induced upon oxidative stress, in stationary phase and in the intra-phagosomal milieu, suggesting regulation of sulfur assimilation in response to toxic oxidants (Hampshire et al, 2004;Pinto et al, 2004). Interestingly sulfolipids, important mycobacterial surface components able to inhibit phagosome-lysosome fusion, are sulfated by PAPS (Pinto et al, 2004), suggesting a role of the cysDNC operon also in determining surface composition.…”

The pleiotropic transcriptional response of Mycobacterium tuberculosis to vitamin C is robust and overlaps with the bacterial response to multiple intracellular stresses

“…Mtb H37Rv genes differentially regulated in response to vit C treatment were compared to the data from other dormancy models. These datasets were obtained from NCBI GEO (http://www.ncbi.nlm.nih.gov/gds/) and included responses observed under hypoxia [Wayne hypoxia model wherein hypoxia develops gradually by the consumption of oxygen in sealed culture tubes (Voskuil et al, 2004) and the enduring hypoxia model wherein severe hypoxia (0.2 % oxygen with N 2 balance) is developed rapidly (Rustad et al, 2008)], low pH (Fisher et al, 2002, Rohde et al, 2007, nutrient starvation (Betts et al, 2002;Hampshire et al, 2004), oxidative stress Voskuil et al, 2011), nitrosative stress (Voskuil et al, 2011) and in response to both vit C (Vilchèze et al, 2013) and macrophage infection . The datasets were filtered for up-and downregulated genes and then compared to vit C response using PERL programs for the commonly upand downregulated genes.…”

“…The survival mechanisms adopted by intracellular Mtb under conditions of nutrient limitation were delineated in an in vitro nutrient starvation model (Loebel et al, 1933;Betts et al, 2002). An aerobic nutrient depletion adaptation model was described where the dissolved oxygen tension was maintained at 50 %, which allowed the maintenance of long-term stationary phase culture (Hampshire et al, 2004). The model was based on the hypothesis that a bacterial subpopulation of Mtb that survives in vitro during the stationary phase may represent the population of the bacilli that persist during infection.…”

Characterization of the mycobacterial chromosome segregation protein ParB and identification of its target in Mycobacterium smegmatis

“…A unique feature of M. tuberculosis is its ability to maintain a dormant, non-replicating state for extended periods of time under unfavourable conditions. The mechanism (or mechanisms) by which this bacterium shifts to a dormant state and reverts to active growth is not well understood (Hampshire et al, 2004;Gó mez & Smith, 2000;Wayne & Hayes, 1996: Wayne & Sohaskey, 2001. Knowledge regarding the steps of the mycobacterial cell cycle (replication, chromosome segregation and cell division) seems to be critical for understanding the mechanisms that are responsible for the transition from an active to a non-replicative persistent state (and vice versa) of pathogenic mycobacteria, particularly M. tuberculosis.…”