A Comparative Lipidomics Platform for Chemotaxonomic Analysis of Mycobacterium tuberculosis

Abstract

Summary The lipidic envelope of Mycobacterium tuberculosis promotes virulence in many ways, so we developed a lipidomics platform for broad survey of cell walls. Here we report two new databases (MycoMass, MycoMap), 30 lipid fine maps and mass spectrometry datasets that comprise a static lipidome. Further, by rapidly regenerating lipidomic datasets during biological processes, comparative lipidomics provides statistically valid, organism-wide comparisons that broadly assess lipid changes during infection or among clinical strains of mycobacteria. Using stringent data filters, we tracked more than 5,000 molecular features in parallel with few or no false positive molecular discoveries. The low error rates allowed the first chemotaxonomic analyses of mycobacteria, which describe the extent of chemical change in each strain and identified particular strain-specific molecules for use as biomarkers.

Self-poisoning of Mycobacterium tuberculosis by interrupting siderophore recycling

“…To quantify the level of endogenous siderophores in the ΔmmpS4/S5 mutant and to examine whether these mutations changed the siderophore profile, the whole-cell lipidome of the ΔmmpS4/S5 mutant was analyzed under low-iron conditions. Cellular lipids were analyzed from chloroform/ methanol extracts of these strains grown on low-iron plates, as previously described (19,24). The phospholipid levels were identical for all strains ( Fig.…”

Molecular profiling of Mycobacterium tuberculosis identifies tuberculosinyl nucleoside products of the virulence-associated enzyme Rv3378c

“…To identify the genes necessary for 1-TbAd production, an existing library of random transposon insertional mutants (25) was screened in high throughput (4,196 mutants) for 1-TbAd production using a simplified 3-min HPLC-MS method (Fig. 3A).…”

“…Bacteria were cultured and extracted by chloroform/methanol mixtures or ethyl acetate, respectively, as described (4,37). Lipid extracts were analyzed using an Agilent 6520 Accurate-Mass Q-Tof and a 1200 series HPLC system with a Varian MonoChrom Diol column (4, 37) with data output from XCMS and MultiplotPreprocess and Multiplot modules of GenePattern (Broad Institute) (38).…”

“…Despite study of this pathogen for more than a century, the spectrum of natural lipids within M. tuberculosis membranes is not yet fully defined. For example, the products of many genes annotated as lipid synthases remain unknown (3), and mass spectrometry detects hundreds of ions that do not correspond to known lipids in the MycoMass and LipidDB databases (4,5).…”

“…To broadly compare the lipid profiles of virulent and avirulent mycobacteria, we took advantage of a recently validated metabolomics platform (4). This high performance liquid chromatography-mass spectrometry (HPLC-MS) system uses methods of extraction, chromatography, and databases that are specialized for mycobacteria.…”