Control of Salmonella especially, Salmonella Enteritidis (SE) in mature laying hens is a major concern in the layer industry. The possible routes of Salmonella transmission include oral (OR) and intraclaocal (IC), and show no major differences between the recovery. We evaluated both OR and IC route of Salmonella by challenging White Leghorns using nalidixic acid resistant Salmonella Enteritidis (SENAR). After finding the OR route an easier and safer method to challenge hens, we carried out further research incorporating prebiotics, probiotics, bacteriophages and nitrogen containing compounds into the diets.Prebiotics, fructoligosaccharides (FOS) in the diet showed a trend towards decrease in the level of colonization of SENAR. Laying hens were challenged and fed two levels of FOS. Ceca, liver with gall bladder (L/GB), spleen and ovaries were analyzed for SENAR. The results showed FOS lowered ceca SENAR. Fecal shedding was significantly (P<0.05) lower in the 1.0% of FOS supplemented groups compared to the SENAR challenge control. There was a significant upregulation (P<0.05) of mRNA toll-like receptor (TLR)-4 and interferon gamma (IFN-) levels but no changes in other cytokines, such as interleukin (IL)- 1, IL-6 or IL-10 mRNA levels, bysupplementing FOS. Histology measurements of ileum showed no significant difference among treatments. Immunohistochemistry (IHC) analyses of ileal samples showed the numbers of immunoglobulin A (IgA) positive cells were higher in the Salmonella challenged and 1.0% FOS supplementation, whereas in challenged control and 0.5% FOS, the numbers were fewer.Supplementation with bacteriophages (BP), particularly at 0.2% in a hens diet showed greater efficacy in lower Salmonella in the ceca, L/GB, and spleen. Immune cytokine mRNA expression levels of IFN-, interleukin IL-6 and IL-10, were significantly higher (P<0.05) in the ileum of SENAR challenged hens as well as challenged and BP treated hens when compared to the negative control.Supplementation of 100 ppm NE and 200 ppm NP significantly reduced (P<0.05) cecal SENAR count, whereas 200 ppm NE and 100 ppm NP did not differ from the SENAR challenged control. Immune cytokines mRNA levels of IFN-, IL-1, IL-6, TLR-4 and IL-10, were significantly upregulated (P<0.05) by SENAR challenge. Supplementation at both levels of NE and NP showed significant immune gene expression responses in the ileum with reduction of IFN-, IL-6, TLR-4 and IL-10 mRNA levels.Supplementation of probiotics (Lactoplan-B) showed a significant upregulation of mRNA levels of IL-1, IL-6, IL-10, TLR-4 and IFN- by SENAR challenge. Highest probiotic level resulted in a significant decrease in IFN- and elevation of IL-10 gene expression in the ileum of chickens. The research conducted demonstrates that prebiotics (FOS), probiotics, BP and nitrocompounds can be included for use as intervention hurdles in the dietary strategies of the laying industry to reduce the SENAR infection on the farm and in the hens.