Low-input RNA-seq suggests metabolic specialization underlying morphological heterogeneity in a gut commensal bacterium.

Isogenic bacteria can be phenotypically diverse. This heterogeneity is evident in the Bacteroidota, a predominant phylum of the human gut microbiota. These bacteria adopt diverse morphologies, yet the molecular basis of their morphological heterogeneity is poorly understood. Here, we systematically characterize the variation in cellular morphology of Bacteroides thetaiotaomicron cells during laboratory growth and after isolation from different host niches. We develop a sensitive transcriptomics approach and apply it to B. thetaiotaomicron sorted into sub-populations of varying cell sizes. Differential expression analysis indicates metabolic specialization associated with morphology. Transcriptomic data also reveal morphological marker genes, whose size-dependent expression is validated through fluorescence in situ hybridization. Morphological characterization of deletion and overexpression mutants reveals that specific marker genes causally contribute to B. thetaiotaomicron cell-size determination. Since phenotypic heterogeneity is a common feature of microbial consortia, this study serves as a blueprint for understanding the role of bacterial genes in morphological variation.