Non-Replicating Mycobacterium tuberculosis Elicits a Reduced Infectivity Profile with Corresponding Modifications to the Cell Wall and Extracellular Matrix

Bacon, Joanna; Alderwick, Luke J.; Allnutt, Jon A.; Gabašová, Evelina; Watson, Robert J.; Hatch, Kim A.; Clark, Simon; Jeeves, Rose E.; Marriott, Alice; Rayner, Emma; Tolley, H. Dennis; Pearson, Geoffrey; Hall, G.A.; Besra, Gurdyal S.; Wernisch, Lorenz; Williams, Ann; Marsh, Philip

doi:10.1371/journal.pone.0087329

Cited by 72 publications

(64 citation statements)

References 62 publications

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“…1). The OD data were consistent with the cfu measurements and indicated a constant viable count of culturable cells, which is associated with data found for non-replicating persistence (NRP) 1 (Chao & Rubin, 2010;Shi et al, 2010;Bacon et al, 2014). The exception was M. phlei that showed a decline of growth from 1 day to 2 days after the log phase followed by constant values until 28 days suggesting a death phase before the switch to NRP (Bacon et al, 2014).…”

Section: Growth Properties Under Aerated and Hypoxic Cultivationsupporting

confidence: 86%

Metabolite analysis of Mycobacterium species under aerobic and hypoxic conditions reveals common metabolic traits

Drapal

Wheeler²,

Fraser

2016

Microbiology

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A metabolite profiling approach has been implemented to elucidate metabolic adaptation at set culture conditions in five Mycobacterium species (two fast-and three slow-growing) with the potential to act as model organisms for Mycobacterium tuberculosis (Mtb). Analysis has been performed over designated growth phases and under representative environments (nutrient and oxygen depletion) experienced by Mtb during infection. The procedure was useful in determining a range of metabolites (60-120 compounds) covering nucleotides, amino acids, organic acids, saccharides, fatty acids, glycerols, -esters, -phosphates and isoprenoids. Among these classes of compounds, key biomarker metabolites, which can act as indicators of pathway/process activity, were identified. In numerous cases, common metabolite traits were observed for all five species across the experimental conditions (e.g. uracil indicating DNA repair). Amino acid content, especially glutamic acid, highlighted the different properties between the fast-and slowgrowing mycobacteria studied (e.g. nitrogen assimilation). The greatest similarities in metabolite composition between fast-and slow-growing mycobacteria were apparent under hypoxic conditions. A comparison to previously reported transcriptomic data revealed a strong correlation between changes in transcription and metabolite content. Collectively, these data validate the changes in the transcription at the metabolite level, suggesting transcription exists as one of the predominant modes of cellular regulation in Mycobacterium. Sectors with restricted correlation between metabolites and transcription (e.g. hypoxic cultivation) warrant further study to elucidate and exploit post-transcriptional modes of regulation. The strong correlation between the laboratory conditions used and data derived from in vivo conditions, indicate that the approach applied is a valuable addition to our understanding of cell regulation in these Mycobacterium species.

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Section: Growth Properties Under Aerated and Hypoxic Cultivationsupporting

confidence: 86%

Metabolite analysis of Mycobacterium species under aerobic and hypoxic conditions reveals common metabolic traits

Drapal

Wheeler²,

Fraser

2016

Microbiology

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“…With a lipid-rich cell wall envelope, mycobacterial cell wall remodeling is also tightly associated with its lipid metabolism (45) and can be a possible adaptive mechanism of M. tuberculosis when coping with a constantly changing host environment (46). In line with the apparent nonreplicative state of iLivE M. tuberculosis, we detected a downregulation of pks2 and desA3 (Table 3), which are essential genes for the biosynthesis of main cell wall components of mycobacteria (47).…”

Section: Resultssupporting

confidence: 52%

Transcriptional Profiling of Mycobacterium tuberculosis Exposed toIn VitroLysosomal Stress

et al. 2016

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e Increasing experimental evidence supports the idea that Mycobacterium tuberculosis has evolved strategies to survive within lysosomes of activated macrophages. To further our knowledge of M. tuberculosis response to the hostile lysosomal environment, we profiled the global transcriptional activity of M. tuberculosis when exposed to the lysosomal soluble fraction (SF) prepared from activated macrophages. Transcriptome sequencing (RNA-seq) analysis was performed using various incubation conditions, ranging from noninhibitory to cidal based on the mycobacterial replication or killing profile. Under inhibitory conditions that led to the absence of apparent mycobacterial replication, M. tuberculosis expressed a unique transcriptome with modulation of genes involved in general stress response, metabolic reprogramming, respiration, oxidative stress, dormancy response, and virulence. The transcription pattern also indicates characteristic cell wall remodeling with the possible outcomes of increased infectivity, intrinsic resistance to antibiotics, and subversion of the host immune system. Among the lysosome-specific responses, we identified the glgE-mediated 1,4 ␣-glucan synthesis pathway and a defined group of VapBC toxin/anti-toxin systems, both of which represent toxicity mechanisms that potentially can be exploited for killing intracellular mycobacteria. A meta-analysis including previously reported transcriptomic studies in macrophage infection and in vitro stress models was conducted to identify overlapping and nonoverlapping pathways. Finally, the Tap efflux pump-encoding gene Rv1258c was selected for validation. An M. tuberculosis ⌬Rv1258c mutant was constructed and displayed increased susceptibility to killing by lysosomal SF and the antimicrobial peptide LL-37, as well as attenuated survival in primary murine macrophages and human macrophage cell line THP-1.

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“…Finally, the increased infectivity of StarvM.tb in this study is in contrast to another recent study (39) (using the guinea pig TB model), where nonreplicating M. tuberculosis was established by the gradual depletion of nutrients in an oxygenreplete environment. In this study, it was found that the nonreplicating M. tuberculosis elicited a significantly reduced infectivity (39). Taken together, these contrasting results may reflect the different methods used to establish a dormant form of M. tuberculosis, and exactly how these different dormant forms of M. tuberculosis are present in humans during a latent (or chronic) infection is important to establish in future studies.…”

Section: Discussionmentioning

confidence: 67%

Differential Influence of Nutrient-Starved Mycobacterium tuberculosis on Adaptive Immunity Results in Progressive Tuberculosis Disease and Pathology

et al. 2015

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When infected with Mycobacterium tuberculosis, most individuals will remain clinically healthy but latently infected. Latent infection has been proposed to partially involve M. tuberculosis in a nonreplicating stage, which therefore represents an M. tuberculosis phenotype that the immune system most likely will encounter during latency. It is therefore relevant to examine how this particular nonreplicating form of M. tuberculosis interacts with the host immune system. To study this, we first induced a state of nonreplication through prolonged nutrient starvation of M. tuberculosis in vitro. This resulted in nonreplicating persistence even after prolonged culture in phosphate-buffered saline. Infection with either exponentially growing M. tuberculosis or nutrient-starved M. tuberculosis resulted in similar lung CFU levels in the first phase of the infection. However, between week 3 and 6 postinfection, there was a very pronounced increase in bacterial levels and associated lung pathology in nutrientstarved-M. tuberculosis-infected mice. This was associated with a shift from CD4 T cells that coexpressed gamma interferon (IFN-␥) and tumor necrosis factor alpha (TNF-␣) or IFN-␥, TNF-␣, and interleukin-2 to T cells that only expressed IFN-␥. Thus, nonreplicating M. tuberculosis induced through nutrient starvation promotes a bacterial form that is genetically identical to exponentially growing M. tuberculosis yet characterized by a differential impact on the immune system that may be involved in undermining host antimycobacterial immunity and facilitate increased pathology and transmission. Mycobacteria are known to adapt to harsh environments by regulating their metabolism, protein expression, and replication. Mycobacterium smegmatis starved of carbon, nitrogen, or phosphorus has been shown to remain viable for over 650 days after an initial 2-to 3-log drop in number of CFU and has displayed increased stress resistance, increased mRNA stability, and an overall decrease in protein synthesis (1). Another example is latent Mycobacterium tuberculosis, which represents a continuum of M. tuberculosis in different growth stages, some of which involve M. tuberculosis in a low-replicating or nonreplicating dormant condition (dormant M. tuberculosis is defined as "nonreplicating bacilli maintaining full viability at a very low metabolic rate" [2]). As one-third of the world's population is infected with latent tuberculosis (TB), these individuals therefore constitute an enormous reservoir for TB disease and transmission (3).Thus, new strategies for the eradication of the low-replicating or nonreplicating dormant mycobacteria that are present during a latent infection are required. We believe that such strategies will arise from a better understanding of the immune response against this particular form of M. tuberculosis, as well as increased insight into the process of reactivation. It has been suggested that dormant mycobacteria reside in phagosomes inside macrophages without free access to oxygen and nutrients. To understand...

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Non-Replicating Mycobacterium tuberculosis Elicits a Reduced Infectivity Profile with Corresponding Modifications to the Cell Wall and Extracellular Matrix

Cited by 72 publications

References 62 publications

Metabolite analysis of Mycobacterium species under aerobic and hypoxic conditions reveals common metabolic traits

Metabolite analysis of Mycobacterium species under aerobic and hypoxic conditions reveals common metabolic traits

Transcriptional Profiling of Mycobacterium tuberculosis Exposed toIn VitroLysosomal Stress

Differential Influence of Nutrient-Starved Mycobacterium tuberculosis on Adaptive Immunity Results in Progressive Tuberculosis Disease and Pathology

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