Clostridium perfringens is one of the main pathogens causing necrotic enteritis which is a highly contagious illness of poultry farm. The disease causes severe economic losses in poultry industry due to bird losses and the expensive of treatment and preventative measures. This study was investigated the effect of betaine on Hemato-biochemical parameters, histopathological picture and intestinal integrity in broiler chickens challenged with C. perfringens. Chickens of various ages (22-40 -day of age) they were collected from different commercial broiler farms at Assiut city. 110 intestinal contents from newly dead and diseased broilers with signs of necrotic enteritis and brownish diarrhea, the collected samples were subjected to bacteriological examination where the total percent of the isolated C. perfringens was 43.6%. Applied experimental study on total of 120, one-day-old broiler chicks (Rose 308) were randomly allotted for three group: 1) negative control (NC), 2) infected group (IG) and 3) treated group (TG), which were dosed with 1.0 ml betaine/L drinking water, from day one until the end of experiment. At 14 days of age, both the infected and treated groups were orally inoculated with 1×10 4 sporulated oocyte of mixed Eimeria species. Five days later, the same groups were orally challenged with a toxigenic strain of C. perfringens by inoculating each bird with 2 ml of broth culture (1×10 8 CFU) per bird. Betaine decreased the severity of necrotic enteritis in (TG) by decreasing the severity of brown diarrhea resulting in a significant (P< 0.05) reduction in the total count of C. perfringens (CFU/g) in the cecum in comparison to the infected and control groups. Betaine supplementation improves blood parameters. Additionally, betaine treatment diminishes pathological lesions in comparison to the infected group.