Low mycorrhizal diversity in the endangered and rare orchids Bipinnula volckmannii and B. apinnula of Central Chile

Several species of the family Orchidaceae have limited distribution and small populations, in which the probability of extinction is much higher than in broadly distributed species. As orchid seeds are very small and lack energy reserves, they require association with compatible mycorrhizal fungi that provide nutrients and carbon needed for germination and early development. Therefore, identification of putative orchid mycorrhizal partners is essential for the design of conservation strategies for these species. We investigated the mycorrhizal associations of two endangered rare species of orchids, Bipinnula apinnula and B. volckmannii, and compared our results with the mycorrhizal associations of two broadly distributed species of Bipinnula. We isolated mycorrhizal fungi and directly amplified DNA from colonized root segments and found an extremely low diversity of mycorrhizal fungi. The rarefaction curves of rare species populations saturated at a single OTU, showing a highly specialized association and considerably lower diversity than the other two species of Bipinnula. Nevertheless, symbiotic germination experiments with fungal strains of this specific fungal OTU only reached the stage of seed coat rupture in both orchid species, without formation of protocorms. Our results suggest that seeds may require additional treatment for successful germination and that fungal partners isolated from adult plants may not be the same that support the germination of seeds. Further studies are needed to improve germination of thiese orchid species and to contribute to their conservation.