Elucidation of the life cycle of a saprotrophic inoperculate discomycete that is associated with pinesap, using a polyphasic taxonomic approach

Abstract

This study deals with the elucidation of the life cycle of an inoperculate discomycete that was eventually collected from conifer wood in South Germany and has become famous for the extraordinary diversity of its secondary metabolites when it was studied extensively during the 1990s. It had then been identified as Lachnum papyraceum (Lachnaceae, Helotiales) based on morphological traits, and extracts from its mycelial cultures were found to possess extraordinary nematicidal and antibiotic activities. Over 60 different secondary metabolites were finally identified from this fungus after extensive variation of culture media and scale up of production up to 100 litre scale. Among the main active principles were mycorrhizin A and chloromycorrhizin A, which had first been reported in 1987 from an unnamed "mycorrhizal" fungus of the hemiparasitic plant, Monotropa hypopitys (pinesap) that was isolated in Sweden. We noted that both the Lachnum strain and the original mycorrhizin producer were still available in the public domain, and decided to study them for comparison, using a multilocus phylogeny and also generated secondary metabolite profiles of both strains using analytical high performance liquid chromatography coupled to diode array and mass spectrometric detection (HPLC-DAD/MS). Surprisingly, the sequence data as well as the secondary metabolite profiles of both strains were highly similar, and it was also confirmed by phylogenetic methods that the strains are indeed nested within the genus Lachnum by comparison of their ITS, LSU and RBP2 sequences. The specimen called L. papyraceum in the old publications was tentatively re-identified by Hans-Otto Baral as L. cf. subvirgineum, but substantial further work on the taxonomy of the genus remains to be done, anyway. We conclude that some Lachnum species have a highly complex but all the more interesting life cycle, and the mycorrhizal symbiont partner may invade the host plant, where it may persist as an endophyte and finally turn saprotrophic on the wood of the senescent pine tree. The taxonomy of these fungi should also be further resolved in the future, using a polythetic concept that includes chemotaxonomic data and a multi-locus genealogy.