Test the monophyly of Calostoma and resolve its higher-level phylogenetic position within the Sclerodermatineae. 3. Infer species-level relationships within the genus using phylogenetic analyses of a comprehensive sampling of Calostoma

Introduction The genus Calostoma comprises a morphologically unusual group of gasteroid fungi (puffballs). The genus name, "beautiful mouth", aptly describes this genus, which forms a brightly colored raised opening at its apex, from which it releases its spores (FIG 1a-b). The genus has been placed in multiple fungal groups including its own family, the Calostomataceae. Recent molecular studies have placed this genus in the suborder Sclerodermatineae of the Boletales, a large order of fungi that includes many ectomycorrhizal species, as well as a few saprotrophic species (Hughey et al 2001, Binder and Bresinsky 2002). Most Boletales have a characteristic cap and stalk morphology (boletoid), while others, including Calostoma, are puffballs that form spores internally (gasteroid) (FIG 1). Calostoma has been interpreted by some authors (Miller and Miller 1988, Bougher and Syme 1998) as being saprotrophic (litter/wood decomposing). However, based on its relationship to the Boletales, we suspect the genus to be ectomycorrhizal (ECM). While the molecular ecology and phylogeny of boletoid and some gasteroid Boletales have been well investigated (Dahlberg and Stenlid 1990; Kretzer et al 1996; Binder et al 1997; Kretzer and Bruns 1997, 1999; Grubisha et al 2001), recent studies in the Sclerodermatineae have been few. No studies have explored the ecology or taxonomic structure of Calostoma in depth. Twenty-nine species have been described in Calostoma, which are distributed from the Americas to Asia and Australasia, but so far only two species (C. cinnabarinum and C. ravenellii) have been included in any phylogenetic study (Hughey et al 2001, Binder and Bresinsky 2002). The proposed research has three major goals: 1. Describe the ecological role of Calostoma using molecular methods and analysis of stable isotopes. As part of this work, the plant hosts of temperate and tropical species will be identified and patterns of host switching will be resolved. 2. Test the monophyly of Calostoma and resolve its higher-level phylogenetic position within the Sclerodermatineae. 3. Infer species-level relationships within the genus using phylogenetic analyses of a comprehensive sampling of Calostoma taxa, including material collected in Southeast Asia (Thailand and Malaysia). Thus, the proposed research represents an integrated study of the phylogeny, taxonomy and ecology of an enigmatic group of basidiomycetes.