Cell-to-cell communication is a universal phenomenon that involves exchange of signals that take many chemical forms. Bacterial, archeal, and eukaryal cells produce and respond to numerous different signals. Yet despite the universality of cell-to-cell communication, there are no tools available to study events involving multiple signals in a spatial fashion.
Here we develop and use thin-layer agar natural product MALDI-TOF imaging (npMALDI-I) mass spectrometry of intact bacterial colonies grown on top of the MALDI target plate. This approach enables real-time visualization of many natural products from bacteria in culture. The chemical complexity of microbial communication, usually depicted as interspecies chemical warfare, is often inferred. Thin-layer agar npMALDI-I captures complex molecular output by an organism and resolves metabolite production both temporally and spatially during colony growth. We developed the thin-layer agar npMALDI-I approach by observing several known signals from a growing colony of Bacillus subtilis and by visualizing interspecies communication between B. subtilis and Streptomyces coelicolor. Amidst all the signals observed from B. subtilis are surfactin, plipastatin, subtilosin, and extracellular polyglutamate. These signals were tracked and correlated with landmark events of colony development and sporulation. Using mixed-species cultures of B. subtilis and S. coelicolor, thin layer agar npMALDI-I provided direct evidence that a wild-type B. subtilis silences the defensive arsenal of Streptomyces coelicolor. We provide direct evidence that bacillaene and surfacin, natural products from B. subtilis, silence the defensive metabolic arsenal of S. coelicolor. These results establish the efficacy of thin layer agar npMALDI-I as a gateway tool to improve our understanding of multifactorial interspecies signaling.