The present work investigated the effects of whey protein on Escherichia coli inoculated in yogurt. The results obtained indicated that the addition of whey protein (WP) led to complete inhibition of E.coli on the 3^rd day at a concentration of 3.13%, while at a concentration of 1.56%, the microorganism required 5 days for complete inhibition. The properties of yogurt were evaluated and the overall acceptability of those fortified with WP was very satisfying, more than the plain ones. 18 male rats were divided into three groups: 1) the control group, 2) the infected group (administered 2 mL of sterile phosphate-buffered saline containing 1 × 10⁹ colony-forming units of E. coli per mL on days 7 and 14), and 3) the treated group (which received the same protocol as the E. coli-infected animals, supplemented with whey protein at a dose of 2 mL/kg daily from day 7 to day 21, the end of the experiment). WP was investigated the effect of experimental infection with Escherichia coli and treatment with whey protein on various biochemical parameters and histopathological changes in rats. At the end of the experiment, blood samples were collected for biochemical analysis. Samples from the liver, kidney, spleen, and intestine were collected for histopathological examination. The obtained results revealed that whey protein supplementation has a strong antibacterial effect and can reduce the severity of E. coli infection at 1.56 and 3.13% and the histopathological alterations were observed in different organs.