AbstractEscherichia coli has a rich history as biology's ‘rock star’, driving advances across many fields. In the wild, E. coli resides innocuously in the gut of humans and animals but is also a versatile pathogen commonly associated with intestinal and extraintestinal infections and antimicrobial resistance—including large foodborne outbreaks such as the one that swept across Europe in 2011, killing 54 individuals and causing approximately 4000 infections and 900 cases of haemolytic uraemic syndrome. Given that most E. coli are harmless gut colonizers, an important ecological question plaguing microbiologists is what makes E. coli an occasionally devastating pathogen? To address this question requires an enhanced understanding of the ecology of the organism as a commensal. Here, we review how our knowledge of the ecology and within-host diversity of this organism in the vertebrate gut has progressed in the 137 years since E. coli was first described. We also review current approaches to the study of within-host bacterial diversity. In closing, we discuss some of the outstanding questions yet to be addressed and prospects for future research.