13C-Flux Spectral Analysis of Host-Pathogen Metabolism Reveals a Mixed Diet for Intracellular Mycobacterium tuberculosis

Comprehensive analysis of the gene expression profiles associated with human monocyte-to-macrophage differentiation and polarization toward M1 or M2 phenotypes led to the following main results: 1) M-CSF-driven monocyte-to-macrophage differentiation is associated with activation of cell cycle genes, substantiating the underestimated proliferation potential of monocytes. 2) M-CSF leads to expression of a substantial part of the M2 transcriptome, suggesting that under homeostatic conditions a default shift toward M2 occurs. 3) Modulation of genes involved in metabolic activities is a prominent feature of macrophage differentiation and polarization. 4) Lipid metabolism is a main category of modulated transcripts, with expected up-regulation of cyclo-oxygenase 2 in M1 cells and unexpected cyclo-oxygenase 1 up-regulation in M2 cells. 5) Each step is characterized by a different repertoire of G protein-coupled receptors, with five nucleotide receptors as novel M2-associated genes. 6) The chemokinome of polarized macrophages is profoundly diverse and new differentially expressed chemokines are reported. Thus, transcriptome profiling reveals novel molecules and signatures associated with human monocyte-to-macrophage differentiation and polarized activation which may represent candidate targets in pathophysiology.

The study documented the emergence of new forms of resistant bacilli (totally drug-resistant [TDR] or super extensively drug-resistant [XDR] tuberculosis [TB] strains) among patients with multidrug-resistant TB (MDR-TB). Susceptibility testing against first- and second-line drugs was performed on isolated Mycobacterium tuberculosis strains. Subsequently, the strains identified as XDR or TDR M tuberculosis were subjected to spoligotyping and variable numbers of tandem repeats (VNTR). Of 146 MDR-TB strains, 8 XDR isolates (5.4%) and 15 TDR isolates (10.3%) were identified. The remaining strains were either susceptible (67%) or had other resistant patterns (20%). Overall, the median of treatments and drugs previously received by MDR-TB patients was two courses of therapy of 15 months' duration with five drugs (isoniazid [INH], rifampicin [RF], streptomycin, ethambutol, and pyrazinamide). The median of in vitro drug resistance for all studied cases was INH and RF. The XDR or TDR strains were collected from both immigrants (Afghan, 30.4%; Azerbaijani, 8.6%; Iraqi, 4.3%) and Iranian (56.5%) MDR-TB cases. In such cases, the smear and cultures remained positive after 18 months of medium treatment with second-line drugs (ethionamide, para-aminosalicylic acid, cycloserine, ofloxacin, amikacin, and ciprofloxacin). Spoligotyping revealed Haarlem (39.1%), Beijing (21.7%), EAI (21.7%), and CAS (17.3%) superfamilies of M tuberculosis. These superfamilies had different VNTR profiles, which eliminated the recent transmission among MDR-TB cases. The isolation of TDR strains from MDR-TB patients from different regional countries is alarming and underlines the possible dissemination of such strains in Asian countries. Now the next question is how one should control and treat such cases.

Recent data indicate that the nutrients available to Mycobacterium tuberculosis (Mtb) inside its host cell are restricted in their diversity. Fatty acids and cholesterol appear more favored; however, their degradation can result in certain metabolic stresses. Their breakdown can generate propionyl-CoA, which gives rise to potentially toxic intermediates. Detoxification of propionyl-CoA relies on the activity of the methylcitrate cycle, the methylmalonyl pathway, or incorporation of the propionyl-CoA into methyl-branched lipids in the cell wall. The current work explores carbon flux through these pathways, focusing primarily on those pathways responsible for the incorporation of propionyl-CoA into virulence-associated cell wall lipids. Exploiting both genetic and biochemical rescue, we demonstrate that these metabolic pressures are experienced by Mtb inside its host macrophage and that the bacterium accesses host fatty acid stores. The metabolism of these host lipids expands the acetyl-CoA pool and alleviates the pressure from propionyl-CoA. These data have major implications for our appreciation of central metabolism of Mtb during the course of infection.