PNNL

Abstract

Bacillus subtilis is a soil bacterium that can form biofilms, or communities of cells encased by an extracellular matrix. In these complex communities, cells perform numerous metabolic processes and undergo differentiation into functionally distinct phenotypes as a survival strategy. Because biofilms are often studied in bulk, it remains unclear how metabolite production spatially correlates with B. subtilis phenotypes within biofilm structures. In many cases, we still do not know where these biological processes are occurring in the biofilm. Here, we developed a method to analyze the localization of molecules within sagittal thin-sections of B. subtilis biofilms using high mass resolution mass spectrometry imaging. We related the organization of specific molecules to the localization of well-studied B. subtilis phenotypic reporters determined by confocal laser scanning microscopy within analogous biofilm thin sections. The correlations between these two datasets support the role of surfactin as a signal for extracellular matrix gene expression in the biofilm periphery and the role of bacillibactin as an iron-scavenging molecule. We then collected untargeted metabolomic data from biofilm thin sections to map the distribution of a wide range of molecules and speculate on their roles in phenotypic responses within B. subtilis biofilms. Taken together, this method will help us discover relationships between molecules and the phenotypic cell states in B. subtilis and other biofilm-forming bacteria.