HIV and sexually transmitted infections is a major threat for humanity in the world, especially in developing countries. The goals of this study were to: (a) analyze differences between genders in knowledge, attitudes, and risk behaviors about HIV/AIDS; and (b) evaluate the association between inconsistent condom use and knowledge, attitudes and risk behaviors about HIV/AIDS in Portuguese university students. Data were collected through a self-administered questionnaire from the Portuguese sample of the Sexual and Reproductive Health in University Students study (SRHUS), which is an extension of the Health Behavior in School-aged Children (HBSC) study for university students. This cross-sectional study provides national representative data of 3278 Portuguese university students. Results show that the majority is sexually active; the usual contraceptive methods more reported are the oral contraceptive and the male condom. However the prevalence of consistent condom use in last 12 months is 34.5% (men) and 31.7% (women). Men more often than women report engaging in sexual risk behaviors (occasional partners, sex associated to alcohol and drugs, STIs). In general, university students have good knowledge about how to protect themselves from HIV and have positive attitudes towards people with HIV/AIDS. This survey could be a useful guide in the development of campaigns or programs designed to convey accurate information about HIV transmission routes and prevention strategies and to dispel erroneous beliefs about HIV.