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Abstract Sphaerellopsisspecies are putative hyperparasites of rust fungi and may be promising biological control agents (BCA) of rust diseases. However, few detailed studies limit potential BCA development inSphaerellopsis. Here, we explored the biogeography, host-specificity, and species diversity ofSphaerellopsisand examined the early infection stage of one species, S. macroconidialis,to infer its trophic status. We randomly screened 5,621 rust specimens spanning 99 genera at the Arthur Fungarium for the presence ofSphaerellopsis. We identified 199 rust specimens infected withSphaerellopsisspecies on which we conducted morphological and multi-locus phylogenetic analyses. FiveSphaerellopsisspecies were recovered, infecting a total of 122 rust species in 18 genera from 34 countries.Sphaerellopsis melampsorinearumsp. nov. is described as a new species based on molecular phylogenetic data and morphological features of the sexual and asexual morphs.Sphaerellopsis paraphysatawas the most commonly encountered species, found on 77 rust specimens, followed bySphaerellopsis macroconidialison 56 andS. melampsorinearumon 55 examined specimens. The type species,Sphaerellopsis filum, was found on 12 rust specimens andSphaerellopsis hakeaeon a single specimen. We also recovered and documented for the first time, the sexual morph ofS. macroconidialis, from a specimen collected in Brazil. Our data indicate thatSphaerellopsisspecies are not host specific and furthermore that most species are cosmopolitan in distribution. However,S. paraphysatais more abundant in the tropics, andS. hakeaemay be restricted to Australia. Finally, we confirm the mycoparasitic strategy ofS. macroconidialisthroughin-vitrointeraction tests with the urediniospores ofPuccinia polysora. Shortly after germination, hyphae ofS. macroconidialisbegan growing along the germ tubes ofP. polysoraand coiling around them. After 12 days of co-cultivation, turgor loss was evident in the germ tubes ofP. polysora, and appressorium-like structures had formed on urediniospores. The interaction studies indicate thatSphaerellopsisspecies may be more effective as a BCA during the initial stages of rust establishment. 

Abstract Rust fungi (Pucciniales, Basidiomycota) are a species-rich (ca. 8000 species), globally distributed order of obligate plant pathogens. Rust species are host-specific, and as a group they cause disease on many of our most economically and/or ecologically significant plants. As such, the ability to accurately and rapidly identify these fungi is of particular interest to mycologists, botanists, agricultural scientists, farmers, quarantine officials, and associated stakeholders. However, the complexities of the rust life cycle, which may include production of up to five different spore types and alternation between two unrelated host species, have made standard identifications, especially of less-documented spore states or alternate hosts, extremely difficult. The Arthur Fungarium (PUR) at Purdue University is home to one of the most comprehensive collections of rust fungi in the world. Using material vouchered in PUR supplemented with fresh collections we generated DNA barcodes of the 28S ribosomal repeat from > 3700 rust fungal specimens. Barcoded material spans 120 genera and > 1100 species, most represented by several replicate sequences. Barcodes and associated metadata are hosted in a publicly accessible, BLAST searchable database called Rust HUBB (Herbarium-based Universal Barcode Blast) and will be continuously updated.