Mycorrhizal associations and symbiotic propagation of two endangered Corunastylis (Orchidaceae)

Context To effectively translocate threatened orchids it is recommended that they are propagated symbiotically with their mycorrhizal fungi. The terrestrial orchid genus Corunastylis has over 25 threatened species, yet there is no information on the identity of the mycorrhizal fungi or symbiotic propagation. Aims To identify the mycorrhizal fungi associated with two endangered species, C. insignis and C. branwhiteorum, and determine effective symbiotic germination methods. Methods Fungi were cultured from roots of Corunastylis at two sites for each species, one of which was shared between species. To estimate taxonomic diversity, 25 fungal isolates were sequenced and identified by comparison with known operational taxonomic units (OTUs) through a combination of phylogenetic analysis and sequence divergence of the internal transcribed spacer (ITS) sequence locus. Further, we tested whether germination varies among growth media, seed accession, and isolates of the same OTU of mycorrhizal fungus. Key results Both species formed mycorrhizal associations with Rhizoctonia (Ceratobasidiaceae), C. insignis with OTU T and U and C. branwhiteorum only with OTU U. We found that modified oats medium without sucrose (MOM) led to significantly greater germination than did MOB medium where oatmeal was substitued with wheat bran or MOB plus coconut water medium (P < 0.05). There was significant variation in germination among mycorrhizal isolates within the same putative species (OTU). Conclusions Both species can be readily symbiotically germinated to adult plants by using Rhizoctonia OTU T and U, significantly improving conservation options for these species. Implications As isolates differ in their germination ability, we recommend ensuring that a range of isolates are included in symbiotic propagation for translocation to maintain diversity and potential to adapt to edaphic conditions.