<b><i>Introduction:</i></b> The aim of this study was to evaluate (i) the knowledge about different dimensions of sexual identity in a group of family pediatricians and (ii) the efficacy of a training program to improve knowledge and reduce genderism and heteronormativity. <b><i>Methods:</i></b> A pre-post-follow-up study was conducted with 96 Italian pediatricians (48 men and 48 women) who participated in a 6-h training program and divided into 2 sections. The first section was theoretical and focused on the conceptual foundations of sexual identity, the depathologizing approach to gender diversity, and the role of pediatricians as the first contacts of children’s or adolescents’ family. The second part was experiential and included the presentation of a clinical case and the activation of a group reflection on the management of gender-diverse youth. Knowledge about sexual identity, genderism, and heteronormativity was measured. <b><i>Results:</i></b> Pre-training questionnaires revealed that the mean score of knowledge about sexual identity was 7.13 ± 3.21. One-way within-subject ANOVA revealed significant effects from pre- to post-training and from pre- to follow-up assessment but not from post-training to follow-up assessment, suggesting that significant changes in the knowledge about sexual identity (<i>F</i> = 39.75, <i>p</i> < 0.001), in personal biases related to genderism (<i>F</i> = 7.46, <i>p</i> < 0.01), and in heteronormative attitudes (<i>F</i> = 44.99, <i>p</i> < 0.001) and behaviors (<i>F</i> = 79.29, <i>p</i> < 0.001) were achieved through the training and maintained at follow-up. <b><i>Conclusion:</i></b> These findings indicate the importance of training pediatricians to work with gender-diverse youth and provide them with the best clinical interventions.