Aims: This study aimed to investigate how confident and comfortable teachers at Tanzanian and South African urban and rural schools are in teaching HIV/AIDS and sexuality. It also aimed at identifying factors associated with teacher confidence and investigated how reported confidence was associated with the implementation of educational programmes on HIV/AIDS and sexuality. Methods: A survey was conducted among South African grade 8 and 9 Life Orientation teachers, and among science teachers for grade 5 to 7 in public primary schools in Tanzania. Teachers’ confidence levels were measured on a four-item scale (0–3). Results: A total number of 266 teachers participated in a survey in 86 schools in South Africa and Tanzania. Overall, teachers report to be rather confident in teaching HIV/AIDS and sexuality. Tanzanian teachers reported higher levels of confidence then did their South Africa colleagues (2.1 vs. 1.8; p <0.01). Confidence in teaching was significantly associated with the numbers of years teaching HIV/AIDS and sexuality, formal training in these subjects, experience in discussing the topics with others, school policy and priority given to teaching HIV/AIDS and sexuality at school. Finally, confidence in teaching remained positively associated with self-reported successful implementation of schoolbased programmes after adjusting for gender, age, religion and numbers of years teaching HIV/AIDS and sexuality. Conclusions: Across urban and rural sites in South Africa and Tanzania teachers reported to be fairly confident in teaching HIV/AIDS and sexuality. Further strengthening of their confidence levels could, however, be an important measure for improving the implementation of such programmes.