Microbial metabolic exchange in 3D

Mono- and multispecies microbial populations alter the chemistry of their surrounding environments during colony development thereby influencing multicellular behavior and interspecies interactions of neighboring microbes. Here we present a methodology that enables the creation of three-dimensional (3D) models of a microbial chemotype that can be correlated to the colony phenotype through multimodal imaging analysis. 

These models are generated by performing matrix-assisted laser desorption ionization time-of-flight (MALDI-TOF) imaging mass spectrometry (IMS) on serial cross-sections of microbial colonies grown on 8 mm deep agar, registering data sets of each serial section in MATLAB to create a model, and then superimposing the model with a photograph of the colonies themselves. 

As proof-of-principle, 3D models were used to visualize metabolic exchange during microbial interactions between Bacillus subtilis and Streptomyces coelicolor, as well as, Candida albicans and Pseudomonas aeruginosa. The resulting models were able to capture the depth profile of secreted metabolites within the agar medium and revealed properties of certain mass signals that were previously not observable using two-dimensional MALDI-TOF IMS.

Enlace al trabajo

Observing the invisible through imaging mass spectrometry

Many microbes can be cultured as single-species communities. The microbial communities, or colonies, curate their environment via metabolic exchange factors such as released natural products. To date, there are very few tools available that can monitor, in a systematic and informative fashion, the metabolic release patterns by microbes grown in a pure or mixed culture. There are significant challenges in the ability to monitor the metabolic secretome from growing microbial colonies. For example, the chemistries of such molecules can be extremely diverse, ranging from polyketides (e.g. erythromycin), non-ribosomal peptides (e.g. penicillin), isoprenoids (e.g. artemisinin), fatty acids (e.g. octanoic acid), microcins (e.g. Nisin), to peptides (e.g. microcin C7), poly-nucleotides and proteins.  Because of this chemical diversity, most of these molecules are extracted prior to analysis and studied one at a time and apart from the native spatial context of a microbial colony. Thus, limited information is obtained about the metabolic output of colonies in a synergetic or multiplexed fashion.

Matrix-assisted laser desorption/ionization-time of flight (MALDI-TOF) imaging mass spectrometry (IMS) is a powerful tool for simultaneously investigating the spatial distribution of multiple different biological molecules. The technique offers a molecular view of the peptides, proteins, polymers and lipids produced by a microbial colony without the need of exogenous labels or radioactive trace material. 

Target compounds can be measured and visualized simultaneously and in a high throughput manner within a single experiment. IMS extends beyond techniques such as MALDI profiling or MALDI intact cell analysis. Although invaluable, these techniques give a broad view of the metabolites produced in reference to a growing colony, where discretely secreted low global concentration but high local concentration metabolites could be missed.

Enlace al trabajo 

Publicaciones de Pieter C. Dorrestein