Phylogenetic and chemotypic diversity of Periglandula species in eight new morning glory hosts (Convolvulaceae)

Periglandula ipomoeae and P. turbinae (Ascomycota, Clavicipitaceae) are recently described fungi that form symbiotic associations with the morning glories (Convolvulaceae) Ipomoea asarifolia and Turbina corymbosa, respectively. These Periglandula species are vertically transmitted and produce bioactive ergot alkaloids in seeds of infected plants and ephemeral mycelia on the adaxial surface of young leaves. Whether other morning glories that contain ergot alkaloids also are infected by Periglandula fungi is a central question. Here we report on a survey of eight species of Convolvulaceae (Argyreia nervosa, amnicola, I argillicola, gracilis, hildebrandtii, leptophylla, muelleri, pes-caprae) for ergot alkaloids in seeds and associated clavicipitaceous fungi potentially responsible for their production. All host species contained ergot alkaloids in four distinct chemotypes with concentrations of 15.8-3223.0 mu g/g. Each chemotype was a combination of four or five ergot alkaloids out of seven alkaloids detected across all hosts. In addition, each host species exhibited characteristic epiphytic mycelia on adaxial surfaces of young leaves with considerable interspecific differences in mycelial density. We sequenced three loci from fungi infecting each host: the nuclear rDNA internal transcribed spacer region (ITS), introns of the translation factor 1-alpha gene (tefA) and the dimethylallyl-tryptophan synthase gene (dmaW), which codes for the enzyme that catalyzes the first step in ergot alkaloid biosynthesis. Phylogenetic analyses confirmed that these fungi are in the family Clavicipitaceae and form a monophyletic group with the two described Periglandula species. This study is the first to report Periglandula spp. from Asian, Australian, African and North American species of Convolvulaceae, including host species with a shrub growth form and host species occurring outside of the tropics. This study demonstrates that ergot alkaloids in morning glories always co-occur with Periglandula spp. and that closely related Periglandula spp. produce alkaloid chemotypes more similar than more distantly related species.