E. coli Facilitates Murine Campylobacter Infection Figure 1. C. jejuni colonization along the gastrointestinal tract of mice with T. gondii-induced acute ileitis. Acute ileitis was induced in conventionally colonized 3-months-old C57BL/6 mice by oral infection with 100 cysts of Toxoplasma gondii ME49 strain. Four days following ileitis induction, mice were orally infected with C. jejuni strain ATCC 43431 as described in Methods. (A) At day 7 p.i. when T. gondii infected mice display severe ileitis (ILE) C. jejuni densities in distinct compartments of the gastrointestinal tract were determined by quantification of live C. jejuni in luminal samples taken from stomach, duodenum, ileum, and colon by cultural analysis (CFU, colony forming units; solid circles) and compared to control mice without ileitis (Con). (B) E. coli loads in luminal ileum samples of mice with T. gondii-induced acute ileitis (day 7 p.i.; open circles) were compared to na¨ıve controls (open diamonds). (C) Relative translocation frequencies (%) of live C. jejuni 43431 ATCC (Cj, black bars) and E. coli (Ec, open bars) were determined in ex vivo biopsies of mesenteric lymphnodes (MLN), spleen, liver, and blood derived from mice with T. gondii-induced acute ileitis at day 7 p.i. by culture in enrichment broths. (D) A complex molecular analysis of the ileal microbiota composition was assessed in mice with T. gondii- induced acute ileitis at day 7 p.i. (ILE; solid circles) and compared to na¨ıve control animals (N; open circles). Main gut bacterial groups were quantified by Real-Time-PCR analyses amplifying bacterial 16S rRNA variable regions and 16S rRNA gene numbers/ng DNA derived from luminal ileal samples: Enterobacteriaceae (EB), Enterococci (EC), Lactic acid bacteria (LB), Bifidobacteria (BIF), Bacteroides/Prevotella spp. (BP), Clostridium leptum group (CLEP), Clostridium coccoides group (CLOCC), Mouse intestinal bacteroidetes (MIB), and total eubacterial load (TL). Numbers of animals harboring C. jejuni or E. coli out of the total number of analyzed animals are given in parentheses. Medians (black bars), standard deviations and significance levels (P- values) determined by Mann-Whitney-U test are indicated. Data shown were pooled from at least three independent experiments. doi:10.1371/journal.pone.0035988.g001 IL-10 Deficient Mice with Chronic Colitis are Susceptible to C. jejuni Colonization Next, we were interested whether colonization resistance against C. jejuni is also abrogated in mice with chronic intestinal inflammation accompanied with gut microbiota shifts towards higher commensal enterobacteria loads [34]. To address this question we infected 6-months-old IL-102/2 mice with chronic colitis harboring a conventional gut microbiota with C. jejuni ATCC 43431. Twelve days following oral infection, C. jejuni could be detected in the ileum and colon (in 55.2% and 69.0% of infected mice, respectively), but not in the stomach or duodenum of IL-102/2 mice with colitis, whereas healthy wildtype control animals remained free of C. jejuni (Fig. 2A). Furthermore, the median C. jejuni loads in infected IL-102/2 mice were more than 4 orders of magnitude lower (105 CFU/g luminal content) when compared to mice with T. gondii-induced acute ileitis (Fig. 1A, 2A). Of note, uninfected IL-102/2 mice with colitis harbored 4 orders of magnitude higher E. coli counts in their large intestines when compared to healthy wildtype mice (Fig. 2B). Interestingly, live C. jejuni could be cultured only from less than 5% (3 out of 67) of MLNs derived from infected IL-102/2 mice, whereas no C. jejuni at all translocated into spleen, liver, kidney or cardiac blood at day 12 p.i. (not shown). A molecular survey of the main bacterial groups within the colonic lumen revealed that IL-102/2 PLoS ONE | www.plosone.org 3 May 2012 | Volume 7 | Issue 5 | e35988