Low uptake of prevention of mother-to-child transmission of HIV (PMTCT) services in resource-limited settings requires new approaches to prevent missed opportunities. Routine HIV testing ("opt-out" testing) in antenatal care (ANC) should be considered. An exploratory cross-sectional survey was conducted in 6 PMTCT sites in rural Zimbabwe. Women who had attended ANC in health centers where PMTCT was provided were surveyed in postnatal services. Of 520 women sampled, 285 (55%) had been HIV tested during their last pregnancy. Primary education or no education (P = 0.02), reporting receiving neither group education in the ANC clinic (P < 0.001) nor individual pretest counseling (P < 0.001), and having attended <6 ANC visits (P < 0.03) were associated with not having been HIV tested. Among the 235 women not HIV tested in ANC, 79% would accept HIV testing if opt-out testing was introduced. Factors associated with accepting the opt-out approach were being <20 years old (P = 0.005), having secondary education or more (P = 0.03), living with a partner (P = 0.001), and the existence of a PMTCT service where the untested women delivered. Thirty-seven women of 235 (16%) would decline routine HIV testing, mainly because of their fear of knowing their HIV status and the need to have their partner's consent. Among the women already tested in ANC (n = 285), 97% would accept the opt-out approach. In Zimbabwe, where 25% of pregnant women are HIV infected, introducing the opt-out strategy for HIV testing may have a far-reaching public health impact on PMTCT. Issues regarding, stigma, quality of post-testing counseling and staffing must be considered, however.