Abstract
Single-cell decisions made in complex environments underlie many bacterial phenomena. Image-based transcriptomics approaches offer an avenue to study such behaviors, yet these approaches have been hindered by the massive density of bacterial mRNA. To overcome this challenge, we combine 1000-fold volumetric expansion with multiplexed error robust fluorescence in situ hybridization (MERFISH) to create bacterial-MERFISH. This method enables high-throughput, spatially resolved profiling of thousands of operons within individual bacteria. Using bacterial-MERFISH, we dissect the response of E. coli to carbon starvation, systematically map subcellular RNA organization, and chart the adaptation of a gut commensal B. thetaiotaomicron to micron-scale niches in the mammalian colon. We envision bacterial-MERFISH will be broadly applicable to the study of bacterial single-cell heterogeneity in diverse, spatially structured, and native environments.
Competing Interest Statement
JRM is a co-founder of, stakeholder in, and advisor for Vizgen, Inc. JRM is an inventor on patents associated with MERFISH applied for on his behalf by Harvard University and Boston Children's Hospital. JRM's interests were reviewed and are managed by Boston Children's Hospital in accordance with their conflict-of-interest policies. All other authors declare that they have no competing interests.