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

Abstract

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.

Non-Tuberculous Mycobacteria: Molecular and Physiological Bases of Virulence and Adaptation to Ecological Niches

“…The characteristic slow growth and the underlying metabolism of NTM enable adaptation to change conditions, turning them less susceptible to antimicrobial agents and disinfectants [ 31 , 69 ] and/or to environmental stresses such as anaerobiosis [ 70 ], starvation [ 71 ], low pH [ 72 ], high temperature [ 73 ], and osmotic stress [ 74 ]. The general adaptation of mycobacteria to extremely stressful conditions is characterized by the entrance of the cell into a dormant state, slowing the metabolism and protecting the cell from life-threatening conditions [ 75 ].…”

“…Additionally, several other genomic mutations, plasmid transfer of resistance determinants, and the production of enzymes that metabolize drugs to a less active form have been described as resistance mechanisms [ 133 , 134 ]. Furthermore, the intracellular growth of NTM [ 10 ] combined with survival in caseum under a non-replicative state of bacterial persistence [ 70 , 135 ], mucus growth [ 135 ], and biofilm growth [ 136 ], make antibiotic delivery and action particularly difficult towards intrinsically resistant NTM [ 137 ]. Although all NTM possess some level of intrinsic resistance to antimicrobials, differences across NTM species have been demonstrated, with, for example, M. kansasii being susceptible to multiple antibiotics, while M. avium is susceptible mainly to macrolides [ 138 ].…”

Metabolite Profiling: A Tool for the Biochemical Characterisation of Mycobacterium sp.

“…The culture conditions mimicked by many studies involve a partially aerated or hypoxic NRP state, typically designated on the basis of protocols from Wayne and Hayes [44], which documented an orderly downregulation of the intermediary metabolism, e.g., [27]. Other studies comparing changes of metabolite levels reported a transcriptional mode of regulation under aerated conditions and a more complex metabolic regulation in the presence of hypoxic conditions, e.g., [45]. Despite evident discrepancies about the mode of regulation in previous studies, overall data identified compounds utilised as storage compounds (e.g., succinic acid, triacylglycerides and poly glutamic acid) and arising during the immediate response to resuscitation.…”

“…Despite evident discrepancies about the mode of regulation in previous studies, overall data identified compounds utilised as storage compounds (e.g., succinic acid, triacylglycerides and poly glutamic acid) and arising during the immediate response to resuscitation. Most of these compounds accumulate intracellularly, but some are transported outside the cell, to reduce toxicity and resulting cellular dysfunction [45,46,47]. The process of resuscitation, which can occur quite suddenly [39], is still poorly understood, as it is difficult to establish the exogenous factors influencing this process [7].…”

“…They share ~99.9% of the DNA and, for unknown reasons, M. bovis BCG never regained virulence [4,65]. A comparison of five Mycobacterium species ( M. bovis BCG, Mycobacterium avium , Mycobacterium intracellulare , M. smegmatis and Mycobacterium phlei ) with different growth rates and pathogenicity, showed that: (i) Their only common trait was the metabolic processes related to the cell wall and (ii) the metabolic changes under aerobic and hypoxic conditions emphasise differences on a genetic level between fast- and slow-growing mycobacteria [45]. Conclusions from other studies highlighted that the model Mycobacterium species for Mtb depends on the purpose of the metabolic study, and the use of phenotypically closer-related species (e.g., M. bovis BCG) is recommended for specialised pathways.…”