Cross-sectional analyses have demonstrated an association between use of hormonal contraceptives and shedding of herpes simplex virus (HSV). This prospective study evaluated the effect of initiating use of hormonal contraception on cervical HSV detection. Two hundred women who were seropositive for HSV-2 and human immunodeficiency virus (HIV) type 1 were examined for cervical mucosal HSV by use of quantitative DNA polymerase chain reaction before and after beginning the use of hormonal contraceptives. Cervical HSV was detected in 32 women (16.0%) before initiating and in 25 women (12.5%) after initiating use of hormonal contraception (P=.4). There were no significant differences in HSV shedding among the subgroups of women starting combination oral contraceptives containing both estrogen and progesterone or progesterone-only contraceptives. Among the 54 women who shed HSV at least once, the median change in cervical HSV after initiation of hormonal contraception was -313 copies/swab. In this prospective study, use of hormonal contraceptives did not increase detection of cervical HSV.