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Sexual behaviors in human immunodeficiency virus (HIV)-infected pregnant women in South Africa are not well understood.Human immunodeficiency virus–infected pregnant women were recruited into a prospective cohort at first antenatal care visit. Sociodemographic information and self-collected vulvovaginal swab samples were collected from participants. Vulvovaginal swab samples were tested for Chlamydia trachomatis, Neisseria gonorrhoea, and Trichomonas vaginalis using GeneXpert. We investigated sexual behaviors, alcohol use, factors associated with condomless sex during pregnancy, and prevalent sexually transmitted infection (STI) among our cohort. We report descriptive, univariate and multivariable logistic regression results of sexual behaviors and alcohol use, factors associated with condomless sex at last sex, and having any STI during pregnancy adjusting for a priori confounders.We recruited and enrolled 430 HIV-infected pregnant women. Median age was 30 years; median gestational age was 20 weeks. Eighty-nine percent of women reported sex during pregnancy. At last sex, 68% reported condomless sex; 18% reported having more than 1 sex partner in the past 12 months. Adjusting for age, income and relationship status, condom use at last sex was associated with prior knowledge of HIV status (adjusted odds ratio [aOR], 2.46; 95% confidence interval [CI], 1.54–3.92) and being in a concordant HIV-positive (aOR, 3.17; 95% CI, 1.84–5.50), or serodiscordant relationship (aOR, 6.50; 95% CI, 3.59–11.80). The prevalence of any STI was 41% (95% CI, 36%–45%). Adjusting for mothers' age and employment, odds of having an STI increased if the woman reported alcohol use during pregnancy (aOR, 1.96; 95% CI, 1.06–3.64) or if the father of the child was a non-cohabiting or casual partner (aOR, 1.42; 95% CI, 0.97–2.03).Almost all HIV-infected pregnant women were sexually active during pregnancy and most women reported condomless sex at last sex. Condom use was associated with knowledge of serostatus and/or partner's serostatus before first antenatal care visit. Factors associated with having STIs included: alcohol use during pregnancy and father of child being a non-cohabiting partner.