Abstract
Adherent-invasive Escherichia coli (AIEC) is a pathovar linked to inflammatory bowel diseases (IBD), especially Crohn’s disease, and colorectal cancer. AIEC are genetically diverse, and in the absence of a universal molecular signature, are defined by in vitro functional attributes. The relative ability of difference AIEC strains to colonize, persist, and induce inflammation in an IBD-susceptible host is unresolved. To evaluate strain-level variation among tissue-associated E. coli in the intestines, we develop a long-read sequencing approach to identify AIEC by strain that excludes host DNA. We use this approach to distinguish genetically similar strains and assess their fitness in colonizing the intestine. Here we have assembled complete genomes using long-read nanopore sequencing for a model AIEC strain, NC101, and seven strains isolated from the intestinal mucosa of Crohn’s disease and non-Crohn’s tissues. We show these strains can colonize the intestine of IBD susceptible mice and induce inflammatory cytokines from cultured macrophages. We demonstrate that these strains can be quantified and distinguished in the presence of 99.5% mammalian DNA and from within a fecal population. Analysis of global genomic structure and specific sequence variation within the ribosomal RNA operon provides a framework for efficiently tracking strain-level variation of closely-related E. coli and likely other commensal/pathogenic bacteria impacting intestinal inflammation in experimental settings and IBD patients.
Highlights
Crohn’s disease (CD) is a type of Inflammatory bowel disease (IBD), a chronic inflammatory condition that is the result of an inappropriate immune response towards elements of the intestinal microbiota [1]
CD patients harbor microbiomes that differ from non-CD individuals in both composition and function, with reduced diversity and an increase of adherent/invasive E. coli (AIEC) [5,6,7,8,9,10,11]
We propose a full-length rRNA sequencing approach– enabled by longer reads produced by nanopore sequencing–that is capable of discriminating among closely-related strains of E. coli
Summary
Crohn’s disease (CD) is a type of Inflammatory bowel disease (IBD), a chronic inflammatory condition that is the result of an inappropriate immune response towards elements of the intestinal microbiota [1]. AIEC are closely associated with the intestinal mucosa, termed the mucosal niche, and induce inflammation in murine models of IBD, including the well-established, inflammation-susceptibe Il10-/- mouse model [3,12,13,14]. AIEC strains lack genes associated with virulence in diarrheagenic E. coli and are defined through the in vitro ability to adhere to and invade epithelial cells and survive in macrophages [5,7,15]. As AIEC are genetically diverse, there is no universal genomic feature that distinguishes the pathotype [16], limiting our ability to detect these pro-inflammatory strains from among patient microbiota
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