BACKGROUND: In recent years, increasing evidence has accumulated that suggests the majority of cases of genital ulcer disease in sub-Saharan Africa are due to viral and not bacterial infections. Although many cross-sectional studies support such a trend, few serial cross-sectional data are available to show the evolution of genital ulcer disease over time. METHODS: We surveyed the prevalence of sexually transmitted diseases (STDs) among patients with STD symptoms and women recruited from family planning clinics in 3 cities in Botswana in 2002 and compared our findings with those from a survey of a similar population conducted in 1993. RESULTS: The observed proportion of cases of genital ulcer disease due to chancroid decreased from 25% in 1993 to 1% in 2002, whereas the proportion of ulcers due to herpes simplex virus increased from 23% in 1993 to 58% in 2002. Although the proportion of ulcers due to syphilis was similar for both surveys, the rate of positive serologic test results for syphilis among patients with genital ulcer disease decreased from 52% in 1993 to 5% in 2002. During this period, decreases in the prevalence of gonorrhea, syphilis-reactive serologic findings, chlamydial infection, and trichomoniasis were also detected among patients with STDs and women from family planning clinics. These changes remained significant after estimates were adjusted for the sensitivity and specificity of diagnostic tests. CONCLUSIONS: Our findings suggest a decrease in the prevalence of bacterial STDs and trichomoniasis, a reduction in the proportion of ulcers due to bacterial causes, and an increase in the proportion of ulcers due to herpes simplex virus during the period 1993-2002. These changes should be taken into consideration when defining new guidelines for the syndromic management of genital ulcer disease.