Colibacillosis, caused by avian pathogenic Escherichia coli (APEC), is a multimillion dollar annual problem for the U.S. poultry industry. Recent recognition that APEC may be a food-borne source of E. coli causing human disease indicates that control of avian colibacillosis is highly desirable for reasons of both animal and human health. Unfortunately, control of this disease has been elusive. RNAseq technology offers a new avenue for comprehensive analysis of the transcriptome. The transcriptomic response of primary lymphoid tissues (bone marrow, thymus, and bursa) each displayed differential expression between different phenotype birds associated with days post-infection (dpi) under systemic APEC infection. Within bone marrow, thymus, and bursa, large numbers of significantly differentially expressed genes were identified in susceptible birds against resistant and non-challenged birds at 5 dpi and 5 dpi against 1 dpi susceptible birds. Moreover, innate immune response (phagosome, lysosome, toll like receptor) is the major response mechanism for host resistant to systemic APEC infection. However, the T and B lymphocytes differentiation and proliferation were significantly impaired in susceptible birds compared to resistant and non-challenged birds at 5 dpi. There were large differences in gene expression in bone marrow versus thymus and bursa across all combination time points in susceptible birds. However, differences of gene expression were not detectable in bone marrow versus thymus and bursa across all combination time points in resistant birds. Choice of the primary lymphoid tissues provide novel insight into the earliest developmental changes that occur in the immune system cells, as they are stimulated to commit to pathways of defense. Additionally, this study also offers an opportunity to access the T and B cells development in the separate primary lymphoid organs. Bioinformatic analysis of differentially expressed genes from each tissue and treatment group, and integration of data across tissues, gave insight into the large subset (networks) of differentially regulated genes in functional genomics of host immune response to systemic APEC infection. This new knowledge can provide the essential preliminary information toward the control of colibacillosis by enhancing host resistance genetics in commercial stocks.