© 2020 International Society for Sexual MedicineBackground: Depression and anxiety levels, sexual dysfunctions, and affective temperament characteristics of women with lifelong vaginismus (LLV) and their male partners may have important effects on the development, maintenance, and exacerbation of LLV. Aim: We aimed to investigate depression and anxiety levels, sexual dysfunctions, and affective temperament characteristics of both women with LLV and their male partners. Methods: 56 women with LLV, their 56 male partners, and 44 couples with no complaints of any sexual function as a control group were included in this study. Dyadic data were analyzed using the Actor-Partner Interdependence Model. Outcomes: The Beck Depression Inventory, Beck Anxiety Inventory, Golombok Rust Inventory of Sexual Satisfaction, and Temperament Evaluation of Memphis, Pisa, Paris and San Diego Auto Questionnaire Scale were completed by all participants. Results: Women with LLV had higher levels of anxiety and depression and had more sexual dysfunctions except for avoidance than those of female controls. Temperament Evaluation of Memphis, Pisa, Paris and San Diego Auto Questionnaire Scale scores were significantly higher in women with LLV for depressive (odds ratio [OR] = 1.27, 95% CI = 1.09–1.49), cyclothymic (OR = 1.31, 95% CI = 1.15–1.49), anxious (OR = 1.22, 95% CI = 1.09–1.38), and irritable (OR = 1.22, 95% CI = 1.04–1.42) temperament than in female controls. It was found that anxiety levels of male partners of women with LLV were higher than those of male controls and that they experienced less sexual satisfaction. Depressive (OR = 1.31, 95% CI = 1.07–1.61) and cyclothymic (OR = 1.18, 95% CI = 1.04–1.34) temperament scores were significantly higher in male partners of women with LLV than in male controls. The Actor-Partner Interdependence Model analyses show that hyperthymic temperament in male partners of women with LLV and anxious and depressive temperament in women with LLV have a negative effect on their own sexual functions. In terms of partner effect, it was found that men with hyperthymic temperament had a negative effect on the sexual functions of women with LLV and men with depressive temperament had a positive effect. Clinical Implications: The individual characteristics of both the women and their male partners have an impact on LLV. Strengths & Limitations: The sample size was relatively small to assess affective temperaments. The inclusion of male partners in the study contributed to our understanding of couples with LLV. Conclusion: Our results indicate that affective temperaments detected in women with LLV (depressive, cyclothymic, anxious and irritable) and their male partners (depressive and cyclothymic) have an effect on the development, maintenance, and exacerbation of LLV, and affective temperaments have an effect on both their own and partner's sexual functions. Turan Ş, Usta Sağlam NG, Bakay H, et al. Levels of Depression and Anxiety, Sexual Functions, and Affective Temperaments in Women With Lifelong Vaginismus and Their Male Partners. J Sex Med 2020;17:2434–2445.