The gut is home to diverse microbial communities that protect against pathogens, with mucus-lined epithelia being a particularly dense area of colonization. But how are these communities organised at the single-cell level, and how does this organisation determine community function?

Here, we investigate how the spatial ecology of host-associated microbiota affects their ability to protect us from pathogens. Using humanzied mice, we systematically study the spatial organisation of gut symbionts in relation to each other, the mucosa, and the invading pathogen Salmonella enterica serovar Typhimurium. We established a new multiplexed staining and imaging approach, which allows for unprecedented strain-level, community-wide positional mapping of pathogens and commensals in the gut landscape. To reveal functional niches in the gut, we augmented these single-cell maps of taxa with genomic phenotypes.

This approach has provided valuable insights into critical changes in the spatial ecology of the gut during infection. We believe that a new frontier in host-microbe symbiosis lies in understanding the spatial ecology of the microbiota, and we provide a framework that will be applicable to a range of systems.