E. coli Facilitates Murine Campylobacter Infection Figure 6. E. coli loads of adult mice following application of E. coli via drinking water. In conventionally colonized 3-months-old adult mice (n = 6) intestinal E. coli loads were raised by adding live E. coli into the drinking water (+ E. coli, until day 8) as described in Methods. At day 8, the E. coli suspension was withdrawn and replaced by regular tap water. Kinetic analyses of E. coli densities were performed by quantification of live bacteria in fecal samples by cultural analysis (CFU, colony forming units) until day 13 p.i. Medians (black bars) and levels of significance (P-values) as compared to the indicated groups (determined by Mann-Whitney-U test) are indicated. Data shown are representative for three independent experiments. doi:10.1371/journal.pone.0035988.g006 positive stool samples and mild diarrhea in the first week following infection, the clinical outcomes were similar in infant mice infected with C. jejuni strains B2 or ATCC 43431. Both strains originate from humans with severe enteritis, are invasive in vitro and induced distinct pro-inflammatory immune responses within the colon mucosa of C. jejuni infected infant mice. The elevated intestinal E. coli loads in mice suffering from intestinal inflammation and in infant mice provide evidence that E. coli is essentially involved in abrogation of CR. This is further underlined by a substantially increased colonization capacity of C. jejuni in adult mice in which the commensal intestinal microbiota was artificially modified towards supra-physiological intestinal E. coli loads. This finding demonstrates again that CR against C. jejuni is mediated by the microbiota and that shifts in the flora represent a risk for C. jejuni infection. We conclude that the murine models of C. jejuni infection presented here shed further lights into the mechanisms underlying CR against C. jejuni offering promising tools to further unravel the interplay between intestinal pathogens, gut microbiota and the innate and adaptive immune system. Figure 7. Time course of C. jejuni colonization in adult mice with increased E. coli loads. In conventionally colonized 3-months-old adult mice intestinal E. coli loads were raised up to 108 bacteria per gram feces by adding live E. coli into the drinking water (+ E. coli) as described in Methods. Mice were orally infected with C. jejuni strain ATCC 43431 (A) or B2 (B). Kinetic analyses of pathogen densities were performed by quantification of live C. jejuni in fecal samples derived from mice with elevated (Ec; solid circles) or normal (N; open circles) E. coli loads by cultural analysis (CFU, colony forming units) until day 12 p.i. (C) C. jejuni B2 densities in distinct compartments of the gastrointestinal tract were determined in luminal samples taken from stomach, duodenum, ileum, and colon of mice with elevated (Ec; solid circles) or normal (N; open circles) E. coli loads at day 12 p.i. Numbers of animals harboring C. jejuni out of the total number of analyzed animals are given in parentheses. Days post infection, medians (black bars) and levels of significance (P-values) as compared to the respective group (determined by Mann-Whitney-U test) are indicated. Data shown are pooled from at least three independent experiments. doi:10.1371/journal.pone.0035988.g007 PLoS ONE | www.plosone.org 9 May 2012 | Volume 7 | Issue 5 | e35988