In situ single-cell activities of microbial populations revealed by spatial transcriptomics

ABSTRACT

Microbial populations and communities are heterogeneous, yet capturing their diverse activities has proven challenging at the relevant spatiotemporal scales. Here we present par-seqFISH, a targeted transcriptome-imaging approach that records both gene-expression and spatial context within microscale assemblies at a single-cell and molecule resolution. We apply this approach to the opportunistic bacterial pathogen, Pseudomonas aeruginosa, analyzing ∼600,000 individuals across dozens of physiological conditions in planktonic and biofilm cultures. We explore the phenotypic landscape of this bacterium and identify metabolic and virulence related cell-states that emerge dynamically during growth. We chart the spatial context of biofilm-related processes including motility and kin-exclusion mechanisms and identify extensive and highly spatially-resolved metabolic heterogeneity. We find that distinct physiological states can co-exist within the same biofilm, just a few microns away, underscoring the importance of the microenvironment. Together, our results illustrate the complexity of microbial populations and present a new way of studying them at high-resolution.