Sexism, heteronormativity and homo/transphobia are pervasive in every social institutions. In sport such culture is evident because the cultural frame is represented by a very strong sex-segregation. The marked difference between males and females can produce dangerous gender stereotypes facilitating the development of a heterosexist culture. Heterosexism is an integral part of the male identity and seems to be particularly evident in team sports. Men socialize their own identity in opposition to the feminine one perceiving themselves as stronger and more powerful. This dynamic creates serious problems for sexual minority youths who are affected by homophobia and genderism. The purpose of this study was to explore knowledge, opinions and attitudes related to gender and sexual issues in sport among 111 undergraduates attending the faculty of Movement Science at Parthenope University of Naples. Participants completed a questionnaire containing many questions about gender and sexual issues and homophobia and transphobia in sports field. In addition they completed also the Genderism and Transphobia Scale (GTS), the Homophobia Scale (HS) and the Ambivalent Sexism Inventory (ASI). Forty% of respondents have wrong notions about gender and sexual issues. Despite that and even though most of them have not directly experienced or witnessed homophobic or transphobic episodes in sport, homophobia and transphobia were considered as significant barriers to participation in sport. Furthermore, more than one third of both males and females showed high levels of sexism, although males had higher levels than females in sexist attitudes. As in sexism, males showed high levels than females in transphobia and homophobia too. Our results suggest the need of introducing specific training in the degree courses of Movement Science aimed to deconstruct gender and sexual stereotypes and prejudices.