Background: Congenital syphilis is associated with perinatal deaths, preterm births and congenital malformations. Low rates of syphilis screening during pregnancy and treatment of those found seropositive have been reported in the Democratic Republic of the Congo (DRC) and Zambia. We report the rates on antenatal syphilis screening, the seroprevalence of syphilis infection, and the frequency of antibiotic treatment in pregnant women screened positive for syphilis during their attendance at antenatal care (ANC) clinics in Kinshasa, DRC and Lusaka, Zambia. Methods: Women attending their first ANC were enrolled consecutively during a 9-month period in 16 and 13 ANC clinics in Kinshasa and Lusaka respectively, in the context of the baseline period of a cluster trial. Study personnel collected data on women’s characteristics, the syphilis screening practices, the test results, and the frequency of treatment, that were done under routine ANC conditions and registered in the clinic records. Results: 4,153 women in Kinshasa and 18,097 women in Lusaka were enrolled. The frequency of screening at the first visit was 59.7% (n= 2,479) in Kinshasa, and 27.8% (n=5,025) in Lusaka. Screening test availability varied. In the periods in which tests were available the screening rates were 92.8% in Kinshasa and 52.0% in Lusaka. The frequency of women screened seropositive was 0.4% (n=10) in Kinshasa and 2.2% (n=109) in Lusaka. Respectively, 10% (n=1) and 11.9% (n= 13) among seropositive women received treatment at the first visit. Conclusions: The results of the study show that screening for syphilis in pregnancy is not universal even when supplies are available. Our ongoing trial will evaluate the impact of a behavioral intervention on changing health providers’ practices to increase screening and treatment rates when supplies are available.