The emergence of new fungal pathogens threatens sustainable crop production worldwide. One mechanism by which new pathogens may arise is hybridization. To investigate hybridization, the related smut fungi, Ustilago maydis and Sporisorium reilianum, were selected because they both infect Zea mays, can hybridize, and tools are available for their analysis. The hybrid dikaryons of these fungi grew as filaments on plates but their colonization and virulence in Z. mays were reduced compared to the parental dikaryons. The anthocyanin induction caused by the hybrid dikaryon infections was distinct, suggesting its interaction with the host was different from that of the parental dikaryons. Selected virulence genes previously characterized in U. maydis and their predicted S. reilianum orthologs had altered transcript levels during hybrid infection of Z. mays. The downregulated U. maydis effectors, tin2, pit2, and cce1, and transcription factors, rbf1, hdp2, and nlt1, were constitutively expressed in the hybrid. Little impact was observed with increased effector expression; however, increased expression of rbf1 and hdp2, which regulate early pathogenic development by U. maydis, increased the hybrid’s capacity to induce symptoms including the rare induction of small leaf tumors. These results establish a base for investigating molecular aspects of smut fungal hybrid pathogen emergence.

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As a biotrophic fungus, Ustilago maydis, the causal agent of common smut of maize, must establish and maintain a relationship with the host throughout the pathogenic cycle. Functional characterization of the U. maydis transcription factor Zfp1 suggests a role in modulating pathogenic development. Deletion of zfp1 resulted in attenuated pathogenic hyphal growth, reduced infection frequency, arrested disease symptom development, and decreased anthocyanin production. Complementation with Zfp1 constructs lacking conserved domains indicated it may function with and without DNA binding and suggests a role for the Fungal_trans domain in enabling full virulence. RNA‐sequencing revealed the transcription of approximately 30% of U. maydis genes, including those encoding predicted and confirmed effectors, were altered in the zfp1 deletion strain during pathogenic growth. Together, the results suggest that Zfp1 is a Zn(II)2Cys6 transcription factor that is required for the full induction of infection, anthocyanin production, and virulence.