Phylogenetic tree and alignments

Hyperparasitic fungi associated with hypocrealean entomopathogens play a pivotal yet underexplored role in regulating fungal-arthropod interaction networks. Despite their frequent occurrence in nature, the taxonomic diversity and evolutionary relationships of these hyperparasites remain poorly resolved. In this study, we conducted an integrative taxonomic investigation of hyperparasites associated with hypocrealean entomopathogenic fungi, together with their closely allied entomopathogenic taxa. Multilocus phylogenetic analyses based on LSU, ITS, SSU, tef-1α, rpb1, and rpb2 were employed to reconstruct phylogenetic relationships, and molecular clock analyses were conducted independently to infer evolutionary timescales of families within Hypocrelaes. Morphological examinations were used to corroborate species delimitation and generic circumscription. Our results substantially expand the known diversity of this group, revealing one novel family, Chlorocilliaceae fam. nov., two new genera (Paralecanicillium gen. nov. and Neodingleyomyces gen. nov.), and 18 new species. In addition, three new combinations, seven new host, geographic, or asexual morph records, and two previously described species are documented, comprising a total of 30 species distributed across eighteen genera and six families. Chlorocilliaceae is proposed to accommodate Albacillium, Chlorocillium, Fiorinimazzantia, Husseyia, Neoaraneomyces, Paraneoaraneomyces, Rousseaua, Speluncomyces and Subuliphorum . Updated descriptions and emended generic concepts of Fiorinimazzantia and Husseyia are provided based on newly collected material. This study advances our understanding of the taxonomy, evolutionary history, and hidden diversity of hyperparasitic fungi allied to hypocrealean entomopathogens, and provides a robust framework for elucidating tripartite host-pathogen-hyperparasite interactions in fungal ecology.