Dbh−/− knockout mouse assays.

Dopamine β-hydroxylase knockout (Dbh^−/−) mice, maintained with a mixed C57BL/6J and 129 SvEv background, were generated at Emory University as previously described (82) and were shipped to the University of Texas (UT) Southwestern Medical School. Next, these 129x1/Svj mice were housed in a specific-pathogen-free facility at the UT Southwestern Medical Center. All experiments were performed under IACUC-approved protocols. At 6 to 11 weeks of age, the mice were orally infected by gavage (either mock infected with PBS or orally infected with 1.3 × 10⁹ CFU of the C. rodentium WT strain [ICC168] or of ΔqseC, ΔqseE, or ΔqseCE isogenic mutants). Mice were monitored daily for survival. The experiments were performed at least twice with a total of 6 mice per group. Colonization changes and virulence gene expression levels were measured directly from RNA extracted from fecal pellets collected in triplicate daily up to day 7 postinfection. The mice were also monitored for weight changes and colon macroscopic differences after euthanization. CFU change curves reflect the averages and standard deviations (SD) of the experiment results. Littermate Dbh^+/− heterozygous mice were considered controls because they had normal NE and Epi levels (83), as previously assayed with other enteric bacteria (8). The 2-way analysis-of-variance (ANOVA) multiple-comparison test was used to determine the statistical significance of the results of the comparisons of data determined each day and among the different mutants to WT levels.