Fifty 1-2-month-old Guinea pigs were divided into 5 equal groups, 10 each. Control (Gp1) did receive neither viable bacteria nor treatment. Each animal from the other groups (Gp2-5) was challenged with (1-2 × 10(8)) viable E. coli in 200 μ L normal saline (0.9%) through IP route. GP2 infected group was treated with 200 μ L saline IP and kept as positive control group. Gp3-4 are infected and treated with Withania somnifera (ethanol root extract) with doses 50 and 100 mg/kg. BW, respectively. Gp5 infected treated group was treated with cefoperazone antibiotic at dose 35 mg/Kg BW. The treatment by drug or the extracted medicinal plant was started 72 h after infection for 7 successive days. Serum and whole blood sample were collected from all groups 14 days after treatment to evaluate some hematological and biochemical changes as well as immunomodulatory cytokine tumor necrosis factor-alpha (TNF- α ). Oral treatment of the plant extract caused significant benefit results in infected Guinea pig appeared in the correction of some hematological and biochemical parameters also try to suppressed inflammatory cytokine response represent in TNF- α . It could be concluded that W. somnifera extract has potent antibacterial activity, and this appears in the correction with hematological, biochemical, and immunological results.