The present study investigated the effects of iron, iron chelators, and mutations of tonB or iroN fepA genes on the growth and virulence of Salmonella Typhimurium. Results indicated that organic iron (ferric citrate and ferrous-l-ascorbate) supported better growth of Salmonella compared to inorganic iron. Among tested chelators, 2,2'-bipyridyl at 500 μM showed the highest inhibition of Salmonella growth with 5 μM ferrous sulfate. Deletion of genes (tonB- and iroN- fepA- ) in the iron uptake system attenuated Salmonella invasion of Caco-2 cells and its ability to damage the epithelial monolayer. The expression of all tested host genes in Caco-2 was not affected under the iron-poor condition. However, claudin 3, tight junction protein 1, tumor necrosis factor α (TNF-α), and interleukin-8 (IL-8) were altered under the iron-rich condition depending on individual mutations. In Caenorhabditis elegans, a significant down-regulation of ferritin 1 expression was observed when the nematode was infected by the wild-type (WT) strain.