Abstract
Adherent-invasive Escherichia coli (AIEC) strains are detected more frequently within mucosal lesions of patients with Crohn’s disease (CD). The AIEC phenotype consists of adherence and invasion of intestinal epithelial cells and survival within macrophages of these bacteria in vitro. Our aim was to identify candidate transcripts that distinguish AIEC from non-invasive E. coli (NIEC) strains and might be useful for rapid and accurate identification of AIEC by culture-independent technology. We performed comparative RNA-Sequence (RNASeq) analysis using AIEC strain LF82 and NIEC strain HS during exponential and stationary growth. Differential expression analysis of coding sequences (CDS) homologous to both strains demonstrated 224 and 241 genes with increased and decreased expression, respectively, in LF82 relative to HS. Transition metal transport and siderophore metabolism related pathway genes were up-regulated, while glycogen metabolic and oxidation-reduction related pathway genes were down-regulated, in LF82. Chemotaxis related transcripts were up-regulated in LF82 during the exponential phase, but flagellum-dependent motility pathway genes were down-regulated in LF82 during the stationary phase. CDS that mapped only to the LF82 genome accounted for 747 genes. We applied an in silico subtractive genomics approach to identify CDS specific to AIEC by incorporating the genomes of 10 other previously phenotyped NIEC. From this analysis, 166 CDS mapped to the LF82 genome and lacked homology to any of the 11 human NIEC strains. We compared these CDS across 13 AIEC, but none were homologous in each. Four LF82 gene loci belonging to clustered regularly interspaced short palindromic repeats region (CRISPR)—CRISPR-associated (Cas) genes were identified in 4 to 6 AIEC and absent from all non-pathogenic bacteria. As previously reported, AIEC strains were enriched for pdu operon genes. One CDS, encoding an excisionase, was shared by 9 AIEC strains. Reverse transcription quantitative polymerase chain reaction assays for 6 genes were conducted on fecal and ileal RNA samples from 22 inflammatory bowel disease (IBD), and 32 patients without IBD (non-IBD). The expression of Cas loci was detected in a higher proportion of CD than non-IBD fecal and ileal RNA samples (p <0.05). These results support a comparative genomic/transcriptomic approach towards identifying candidate AIEC signature transcripts.
Highlights
Crohn’s disease (CD) is a form of inflammatory bowel disease (IBD) that is characterized by skip lesions of transmural inflammation, and can occur at multiple sites in the digestive tract
A greater relative abundance of E. coli has been associated with CD, and in active disease compared to patients in remission [17]
Expressed Genes (DEGs) in LF82 compared to HS
Summary
Crohn’s disease (CD) is a form of inflammatory bowel disease (IBD) that is characterized by skip lesions of transmural inflammation, and can occur at multiple sites in the digestive tract. Inflammation can be found anywhere in the gastrointestinal tract from the mouth to the anus, but in most (60–80%) CD patients, the distal small intestine is frequently involved [1, 2]. Factors implicated in the pathogenesis of IBD include host genetic predisposition, and continual activation of the mucosal immune system by luminal bacteria and their products [3, 4]. A greater relative abundance of E. coli has been associated with CD, and in active disease compared to patients in remission [17]. Mucosa-associated E. coli in particular are more abundant in CD [18] and in several small studies were isolated from inflamed tissue that include areas with ulcers and granulomas [19, 20]. In addition E. coli from the neoterminal ileum in post-surgical CD patients are linked to early recurrence of the disease [2]
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