After removal of spots with a signal to noise ratio less than 3, a total of 24,387 genes were included in the statistical analysis. Effects of treatment were analyzed through contrasts of treatment groups. Contrasts measured tested effect of time for each treatment group, comparisons of each group to control, effect of vaccination, and effect of pathology. Large numbers (>100) of significantly

differentially learn more expressed genes (q value<0.05) were detected in 4 contrasts of interest, all involving the NV–C severe groups. The largest number of differentially expressed genes (1914 genes) was found in the contrast of NV–C severe day 5 vs. NV–C mild day 5. The second largest number of differentially expressed genes (1097 genes) was found in the contrast of NV–C severe group vs. NV–NC control group on day 1. This number was reduced to 506 differentially expressed genes for the same contrast on day 5. The number of differentially expressed Apoptosis inhibitor genes in the contrast of NV–C severe day

1 vs. NV–C severe 5 was 107 genes. Numbers of shared and unique differentially expressed genes between 3 contrasts analyzing differences in treatment group on the same day are displayed in Fig. 2. A total of 417 differentially expressed genes found in the contrast of NV–C severe day 5 vs. NV–NC control day 5 were also differentially expressed in the NV–C severe day 5 vs. NV–C mild day 5 contrast; 280 genes had increased expressed

in the severe group in both contrasts, 137 genes had decreased expressed in the severe group in both contrasts. In all of the shared genes between the contrasts represented in Fig. 2, 99% of genes were expressed in the same direction relative to the severe group; either all more highly expressed in the severe group across all contrasts or more lowly expressed among all contrasts. The differences between internal lesion scores of vaccinated and non-vaccinated, challenged birds were tested through 6-phosphogluconolactonase two-sample t-tests for each day. There was a significant (P value<0.001) reduction in internal lesions among vaccinated, challenged birds compared to those non-vaccinated, challenged birds on both days, however, no differentially expressed genes due to vaccination effect were detected. On day 1, the mean±standard deviation of lesion scores for vaccinated birds was 1.50±1.27 (N=76) and unvaccinated birds 3.06±1.68 (N=87). On day 5, the mean±standard deviation of lesion scores for vaccinated birds was 2.94±2.08 (N=80) and unvaccinated birds 4.28±1.94 (N=85). Vaccination effect on gene expression was tested both through the V–NC group vs. NV–NC contrast on both days and through a contrast utilizing a combination of all treatment groups, non-challenged and challenged, on both days. Samples from individuals with severe pathology or collected 5 day post-infection exhibited more gene induction than repression.