P317 - Mucosa-associated Escherichia coli and Faecalibacterium prausnitzii quantification by real-time PCR and its potential use as complementary diagnostic tool for inflammatory bowel diseases

Abstract

s of the 4th Congress of ECCO the European Crohn’s and Colitis Organisation S135 tissue trauma; IL-1b, IL-10 and IL-6, which together exhibited a significant positive correlation. Production of IL-1b, a key mediator of epithelial inflammation, correlated significantly with TNF-a production, but only in samples from CD or UC. TNF-a release was further associated with IL-12p70(Th1) production in UC, and IL-5(Th2) production in CD. UC patients who had probiotics showed significant decreases in IL-2 levels, suggesting general suppression of T-cell activity. Conclusions: Cytokines associated with tissue trauma were produced from all gut biopsies, but accompanied by TNFa only in tissue from IBD patients. TNF-a in IBD was further associated with production of Th1/Th2 cytokines of opposing bias to the inflammatory profile thought to predominate in these patients probably indicating the counter-balancing of pathologic immune responses in the gut. P317 Mucosa-associated Escherichia coli and Faecalibacterium prausnitzii quantification by real-time PCR and its potential use as complementary diagnostic tool for inflammatory bowel diseases M. Lopez-Siles1 *, M. Martinez-Medina1, X. Aldeguer2, L. Garcia-Gil1. 1Laboratori de Microbiologia Molecular, Departament de Biologia, Universitat de Girona, Girona, Spain, 2Departament de Gastroenterologia, Hospital Dr. Josep Trueta, Girona, Spain Background: It has been demonstrated that CD patients are distinguishable from C subjects according to their intestinal microbial composition, as revealed by qualitative molecular methods [1]. Escherichia coli and Faecalibacterium prausnitzii have been reported to be the main representatives of CD dysbiosis. Aim: The aim of this study was to determine if the quantification of E.coli and F.prausnitzii by real-time PCR could be used as complementary tool for diagnostic purposes in inflammatory bowel diseases. Materials and Methods: E.coli and F.prausnitzii were quantified using a previously reported real-time PCR assay [2] and a new real-time PCR assay designed in our laboratory, respectively. The abundance of E.coli was determined in 26 CD patients, 17 C subjects and 8 patients suffering from UC, whereas F.prausnitzii was quantified in 11 C subjects and 10 CD patients. Both studies were focused on the quantification of those bacteria intimately adhered to the mucosa. Thus, transient and loosely attached bacteria were discarded by mild sonication. Moreover, a quantification of human cells was performed in order to correct variability due to sample size and efficiency of DNA extraction (data are expressed as log bacterial cells/106 human cells). Results: Irrespectively of the zone sampled, CD patients carried higher quantity of E.coli (6.54±1.97 log E.coli /106 human cells) than C (4.97±2.29) and UC patients (4.49±0.71) (p = 0.010), which is in agreement with previous studies [3]. Among CD patients, those with I-CD showed highest E.coli abundance in comparison with those patients with other affectations (I-CD: 8.1±1.55, IC-CD: 4.96±0.72, C-CD: 5.97±1.32; p = 0.001). Concerning F.prausnitzii, CD patients harboured a lower quantity of F.prausnitzii (3.10±1.40 log F.prausnitzii /106 human cells) than C subjects (3.88±0.76) (p = 0.036). For this bacterial indicator, differences among distinct CD affectations were also observed (I-CD: 2.58±1.28, IC-CD: 2.80±1.33, C-CD: 4.64±0.49; p = 0.035). Conclusions: Our methodological approach revealed that in CD patients, E.coli numbers were higher than in C subjects, whereas F.prausnitzii numbers were lower, which makes this a potential diagnostic tool. Differences were especially clear for CD patients with ileal involvement, which is in agreement with previous reports [3]. Since current trends distinguish between I-CD and C-CD as two different pathological entities [4,5], additional bacterial indicators must be searched to find effective tools for C-CD and UC differential diagnose. Abbreviations: CD: Crohn’s disease, UC: ulcerative colitis, C: control, I-CD: Crohn’s ileitis, C-CD: Crohn’s colitis, IC-CD: Crohn’s disease with ileo-colonic affectation.