Mycorrhizal specialization toward each distinct Oliveonia fungus in two closely related photosynthetic Dactylostalix orchids

Abstract

Although rhizoctonias from Ceratobasidiaceae, Tulasnellaceae and Serendipitaceae are typical orchid mycobionts, orchid mycorrhizal fungi exhibit vast taxonomic and ecological diversity. This diversity stems from the high specificity of orchid mycorrhizal associations and the remarkable diversity of over 28,000 orchid species. The subtribe Calypsoinae is particularly notable for its diverse mycorrhizal partnerships, including rhizoctonias, ectomycorrhizal and saprotrophic non‐rhizoctonia fungi. However, the mycobionts within certain Calypsoinae lineages, such as the genus Dactylostalix, remain understudied. This study explores the physiological ecology of two photosynthetic Calypsoinae species, Dactylostalix ringens and Dactylostalix uniflora, to gain insight into potentially novel associations and their ecological implications. We analysed the mycorrhizal communities of both Dactylostalix species using high‐throughput ITS metabarcoding of root samples collected from multiple locations. Additionally, we measured the natural abundances of 13C and 15N isotopes in the leaves of the two Dactylostalix species and their co‐occurring autotrophic reference plants, as well as in fungal pelotons isolated from D. ringens, to assess the potential for partial mycoheterotrophy. Our findings revealed that D. ringens and D. uniflora form specialized mycorrhizal associations predominantly with distinct lineages of Oliveonia (Oliveoniaceae, Auriculariales), even in sympatric populations. Stable isotope analysis showed that both Dactylostalix species exhibited conflicting isotopic signals: elevated δ15N values, supporting partial mycoheterotrophy, but lower δ13C values compared to autotrophic plants, suggesting autotrophy. Peloton samples from D. ringens displayed only modest 13C enrichment relative to autotrophic references. These conflicting isotopic signals make it difficult to precisely determine whether both Dactylostalix species are autotrophic or partially mycoheterotrophic. Intriguingly, the 13C and 15N signatures of Dactylostalix species and their pelotons resemble those of many rhizoctonia‐associated orchids. This isotopic evidence implies a niche overlap with endophytic tendencies between rhizoctonias and Oliveonia, suggesting that potential endophytic traits may have facilitated the recruitment of Oliveonia as novel mycorrhizal partners. Furthermore, the mycorrhizal segregation between D. ringens and D. uniflora likely promotes their sympatric coexistence and may contribute to reproductive isolation through ecological specialization. Read the free Plain Language Summary for this article on the Journal blog.