Congenital toxoplasmosis is a potentially severe infection and its prevention is most often based on serological screening in pregnant women. Many cases could be prevented by simple precautions during pregnancy. Aiming to assess the knowledge about toxoplasmosis among professionals working in antenatal care in a high prevalent region, a questionnaire was administered to 118 obstetric nurses and physicians attending at primary care units and hospitals. The questionnaire was self-completed and included questions on diagnosis, clinical issues, and prevention. Only 44% of total answers were corrected. Lower scores were observed among those with over 10 years of graduation, working in primary care units, and nurses. Errors were mainly observed in questions of prevention and diagnosis. As congenital toxoplasmosis is a mother-to-child (MTC) transmitted disease, early diagnosis and treatment can prevent serious and irreversible fetal damage. Thus, doctors and nurses who provide prenatal care must be appropriately trained on prophylactic, diagnostic, and clinical aspects of toxoplasmosis. The authors suggest that measures should be taken for continuing education regarding toxoplasmosis in pregnancy.