In the framework of HIV serosurveillance, serosurveys in pregnant women are a good alternative to population-based surveys, which are more difficult to implement. In 2002 and 2006, surveys were conducted in Niger to assess the HIV seroprevalence in pregnant women and to evaluate the trend of the HIV epidemic. The overall seroprevalence was 0.96% (95% CI 0.5-1.7%) and 1.3% (95% CI 0.9-1.8%) in 2002 and 2006, respectively, showing no significant change. In the 2006 overall sample, women living in urban areas were significantly more infected than those in rural environments, with prevalences of 1.9% and 0.7%, respectively (P=0.006). Women with higher school attainment were more often HIV-positive than other women (4.6% vs. 1.7%; P<0.001). The 2006 prevalence, which is among the lowest of the sub-Saharan region, was not significantly different from the national seroprevalence measured in adults in 2002 (0.87%, 95% CI 0.5-1.3%). Close monitoring of the epidemic must be continued.