Ceratobasidium orchid mycorrhizal fungi reveal intraspecific variation and interaction with different nutrient media in symbiotic germination of Prasophyllum (Orchidaceae)

Understanding how nutrient requirements of orchid mycorrhizal fungi (OMF) affect symbiotic germination is essential for the ex situ conservation of threatened orchids and their mycorrhizal symbioses. Yet the influence of isolate-level variation in OMF nutrient preferences on orchid germination is unknown. We tested germination of Prasophyllum frenchii (Orchidaceae) on 15 different media of varying carbon and macronutrient compositions with three Ceratobasidium isolates of the same operational taxonomic unit (OTU) as determined with internal transcribed spacer locus sequencing. There was a significant interaction between media and fungal isolate on percentage germination, with each isolate recording its highest percentage germination on different nutrient media (Isolate 9.3: 5.2 +- 1.4% on MOM-S; Isolate 8.2: 5.4 +- 1.1% on MOM + S; Isolate 4.3: 2.2 +- 0.5% on 1.25 g/L wheat bran agar). Across all isolates, germination (percentage germination > 0) occurred more frequently on wheat bran agar media (39.7% of plates) than on oatmeal agar media (6.0% of plates). There was also an effect of media type on aerial hyphal growth behaviour of the OMF isolate. All isolates supported growth through to adult flowering plants. We demonstrated that symbiotic germination of Prasophyllum is affected by media composition. Further, percentage germination and aerial hyphal growth behaviour differed significantly among OMF isolates of the same OTU. This illustrates that a diversity of functionally significant fungal strains occurs within a single OTU, a previously unknown aspect of OMF research with important ecological and conservation implications.