Molecular identification of a mycorrhizal Sebacinaceae from the endangered Caladenia atroclavia (black clubbed spider orchid)

Caladenia atroclavia D. Jones & M. Clements is an endangered orchid found only in the Stanthorpe region of south east Queensland. As part of a recovery process for the species we aimed to isolate and identify the mycorrhizal fungus from the plant. Fungal coils were isolated from the orchid and grown into pure culture. DNA was extracted from these cultures and also from colonised plant stem collar regions. Following fungal ITS-DNA PCR amplification, cloning and BLAST similarity searches, we identified the main fungal endophyte within the orchid as a Sebacinaceae sp. Co-inoculation trials showed that isolates of the fungus were capable of germinating seed of the orchid. Neighbour-joining phylogenetic analysis of the ITS-DNA of this mycorrhizal fungus showed that it was related to Sebacinaceae spp. that have been isolated from other Australian Caladenia spp. This fungus will be used in ex situ propagation procedures to ultimately increase the number of plants in the natural state