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Background: Toxoplasmosis is a major public health problem among immunocompromised individuals. This study aimed to determine the seroprevalence and associated risk factors of Toxoplasma gondii infection among pregnant women with and out HIV infections. Methods: This cross sectional study was conducted among consecutive 385 pregnant women attended Antenatal Clinic from May 2010 to October 2011 at the Gondar University Teaching Hospital, Northwest Ethiopia. Venous blood was collected from each pregnant woman for testing HIV-1/2 and anti- Toxoplasma antibodies using rapid test kits. Data were entered and analyzed using SPSS version 20 statistical package. Results: The overall magnitude of T. gondii and HIV was 88.6% (341/385) and 11.2% (43/385), respectively. The seroprevalence of T. gondii was not different among HIV infected and non-infected pregnant women (88.4%, 38/ 43 vs. 88.6%, 303/342). Keeping cats in house showed statistically significant association with seropositivity of toxoplasmosis (P