Toxoplasma gondii is one of the most prevalent zoonotic parasites being responsible for major economic losses in sheep and abortion in pregnant women. One hundred samples of cat feces were examined for T. gondii oocysts using sheather's sugar flotation. The prevalence of T. gondii oocysts was 2% at Sharkia Province, Egypt. For experimental infection, twelve kittens were randomly fed on the diaphragm meat (100 gm/ each kitten), collected from freshly slaughtered sheep at EL-Bassatein abattoir, Cairo, while other two kittens were left as a control group. Eleven out of twelve kittens shed unsporulated oocyst with an infection rate of 91.7%. The prepatent period was ranged from 4-7 days, while the patent period was within the range of 7- 11 days. After DNA extraction of T. gondii oocyst from feces of four experimentally infected kittens and two naturally infected cats, the B1 gene was amplified in all samples with a PCR product of 115 bp. Also, one hundred blood samples of pregnant sheep, with a history of previous abortions, were collected from three flocks in Sharkia Province. While, one hundred sera of pregnant women in their first trimester were collected from private labs after obtaining a comprehensive questionnaire that investigates the risk factors associated with prevalence of toxoplasmosis. The collected sera of sheep and pregnant women were serologically investigated for T. gondii antibodies by indirect hemagglutination test using Toxo-HAI Fumouze Kits.The seroprevalence rate of anti-T. gondii IgG antibodies was 85% in pregnant sheep, while IgM antibodies of Toxoplasma infection were negative. The anti-T. gondii IgG antibodies in seropositive sheep were evaluated with titers ranging from 1:160 to 1:2560. Moreover, the seroprevalence rates of anti-T. gonii IgG and IgM antibodies in pregnant women were 30 and 10%, respectively, but only 10% revealed a mixed seroprevalence for Toxoplasma IgG and IgM antibodies. The titers of anti-T. gondii IgG and IgM antibodies in seropositive women were ranged from1:160 to1: 2560 and 1:160 to 1:320, respectively. There were significant correlations between the seropositivity of T. gondii specific IgG antibodies in pregnant women and the most investigated risk factors including; knowledge about transmission modes, contact with cats, luncheon and sausage consumption, gardening or contact with soil, washing hands before meals and unwashed raw vegetables or fruits consumption. Meanwhile, no significant association between T. gondii seropositivity and ingestion of undercooked meat and viscera. This study emphasized that cats and sheep play a great role in epidemiology of T. gondii that having a public health hazard in pregnant women. Thereby it is recommended a further genotyping for T. gondii strains from different hosts to predict a recent strategy for prevention and control of such zoonotic parasite.