Cell-mediated immunity and production of type 1 cytokines are the main defenses against pathogenic fungi. Ligation of CD40 by CD40L on T cells is critical for the induction of these immune responses in vivo. We explored the role of CD40/CD40L interactions in vaccine immunity to Blastomyces dermatitidis by immunizing CD40−/− and CD40L−/− mice and analyzing their resistance to reinfection in a murine pulmonary model. In the absence of CD40 or CD40L, CD4+ cells failed to get primed or produce type 1 cytokine and impaired the generation of CD8+ T1 cells. The CD8+ T cell defect was not due to regulatory T cells or impaired APC maturation or Ag presentation to T cells. If CD4+ cells were first eliminated, vaccination of CD40−/− and CD40L−/− mice restored priming of CD8+ cells, type 1 cytokine production, and resistance. Hence, CD4+ and CD8+ cells differ sharply in their requirement for CD40/CD40L interaction during the generation of antifungal immunity. Despite the plasticity of T cell subsets in vaccine immunity, in absence of CD40/CD40L interaction, CD4+ cells may impede the priming of CD8+ cells at the cost of host survival against a lethal infectious disease.