Candidalysin is the first cytolytic peptide toxin identified in any human fungal pathogen. Candidalysin is secreted by Candida albicans and is critical for driving infection and host immune responses in several model systems. However, Candida infections are also caused by non-C. albicans species. Here, we identify and characterize orthologs of C. albicans candidalysin in C. dubliniensis and C. tropicalis. The candidalysins have different amino acid sequences, are amphipathic, and adopt a predominantly α-helical secondary structure in solution. Comparative functional analysis demonstrates that each candidalysin causes epithelial damage and calcium influx and activates intracellular signaling pathways and cytokine secretion. Importantly, C. dubliniensis and C. tropicalis candidalysins have higher damaging and activation potential than C. albicans candidalysin and exhibit more rapid membrane binding and disruption, although both fungal species cause less damage to epithelial cells than C. albicans. This study identifies the first family of peptide cytolysins in human-pathogenic fungi. IMPORTANCE Pathogenic fungi kill an estimated 1.5 million people every year. Recently, we discovered that the fungal pathogen Candida albicans secretes a peptide toxin called candidalysin during mucosal infection. Candidalysin causes damage to host cells, a process that supports disease progression. However, fungal infections are also caused by Candida species other than C. albicans. In this work, we identify and characterize two additional candidalysin toxins present in the related fungal pathogens C. dubliniensis and C. tropicalis. While the three candidalysins have different amino acid sequences, all three toxins are α-helical and amphipathic. Notably, the candidalysins from C. dubliniensis and C. tropicalis are more potent at inducing cell damage, calcium influx, mitogen-activated protein kinase signaling, and cytokine responses than C. albicans candidalysin, with the C. dubliniensis candidalysin having the most rapid membrane binding kinetics. These observations identify the candidalysins as the first family of peptide toxins in human-pathogenic fungi.