The aim of this study was to investigate knowledge of PMTCT and to describe potential barriers that might affect acceptability of interventions for PMTCT in a resource poor setting in South Africa. The sample included 1534 pregnant women recruited at first antenatal care visit from 5 clinics implementing PMTCT (61%) and from 5 communities around the 5 clinic areas (39%). In addition, the mothers or mothers-in-law (70.9%) and husbands or partners (58.2%) of the pregnant women were interviewed at their homes. Results indicate that major potential barriers in implementing PMTCT programmes in a resource poor setting included physical access to the health facility, PMTCT knowledge, stigma and support, HIV testing, and delivery preference.