BACKGROUND: Participatory health education interventions and/or community-based primary health care in remote regions can improve child survival. The most recent data from Guinea Bissau shows that the country ranks 5th from bottom globally with an under-five mortality rate of 198 per 1000 live births in 2007. EPICS (Enabling Parents to Increase Child Survival) is a cluster randomised trial, which is currently running in rural areas of southern Guinea Bissau. It aims to evaluate whether an intervention package can generate a rapid and cost-effective reduction in under-five child mortality. The purpose of the study described here was to understand levels of knowledge on child health and treatment-seeking and preventative behaviours in southern Guinea Bissau in order to develop an effective health education component for the EPICS trial. The study also aimed to assess the effect of gender and ethnicity on knowledge and behaviour. METHODS: Women and men were interviewed in their households using a structured questionnaire. Characteristics of the households and of the interviewed women and men were tabulated. The number of correct answers given to the health knowledge and practice questions and their percentage distribution were tabulated by items and by gender. An overall health knowledge score was derived. RESULTS: There are low levels of appropriate knowledge on child health, some inappropriate practices and generally low vaccination coverage. Health knowledge scores improve significantly amongst those who have accessed higher education. Differences in health knowledge between women and men become insignificant once age and education are accounted for. CONCLUSIONS: Health education activities should be an integral part of a package to improve child survival in rural Guinea Bissau. These activities should focus on diarrhoea, malaria, pneumonia, pregnancy, delivery, neonatal care and vaccination coverage, as these are areas where knowledge and practices were found to be inadequate in this study. Men as well as women should be involved in these activities. Prior to developing health education interventions in similar settings, studies to assess areas to be targeted should be conducted.